Laser Desorption Ionization Mass Spectrometry Based on Nanophotonic Structure: From Material Design to Mechanistic Understanding (Springer Theses) 9819968771, 9789819968770

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Springer Theses Recognizing Outstanding Ph.D. Research

Moon-Ju Kim

Laser Desorption Ionization Mass Spectrometry Based on Nanophotonic Structure: From Material Design to Mechanistic Understanding

Springer Theses Recognizing Outstanding Ph.D. Research

Aims and Scope The series “Springer Theses” brings together a selection of the very best Ph.D. theses from around the world and across the physical sciences. Nominated and endorsed by two recognized specialists, each published volume has been selected for its scientific excellence and the high impact of its contents for the pertinent field of research. For greater accessibility to non-specialists, the published versions include an extended introduction, as well as a foreword by the student’s supervisor explaining the special relevance of the work for the field. As a whole, the series will provide a valuable resource both for newcomers to the research fields described, and for other scientists seeking detailed background information on special questions. Finally, it provides an accredited documentation of the valuable contributions made by today’s younger generation of scientists.

Theses may be nominated for publication in this series by heads of department at internationally leading universities or institutes and should fulfill all of the following criteria ● They must be written in good English. ● The topic should fall within the confines of Chemistry, Physics, Earth Sciences, Engineering and related interdisciplinary fields such as Materials, Nanoscience, Chemical Engineering, Complex Systems and Biophysics. ● The work reported in the thesis must represent a significant scientific advance. ● If the thesis includes previously published material, permission to reproduce this must be gained from the respective copyright holder (a maximum 30% of the thesis should be a verbatim reproduction from the author’s previous publications). ● They must have been examined and passed during the 12 months prior to nomination. ● Each thesis should include a foreword by the supervisor outlining the significance of its content. ● The theses should have a clearly defined structure including an introduction accessible to new PhD students and scientists not expert in the relevant field. Indexed by zbMATH.

Moon-Ju Kim

Laser Desorption Ionization Mass Spectrometry Based on Nanophotonic Structure: From Material Design to Mechanistic Understanding Doctoral Thesis accepted by Yonsei University, Seoul, Korea (Republic of)

Author Dr. Moon-Ju Kim Nano Bio System Laboratory, Department of Materials Science and Engineering Yonsei University Seoul, Korea (Republic of)

Supervior Prof. Dr. Jae-Chul Pyun Nano Bio System Laboratory, Department of Materials Science and Engineering Yonsei University Seoul, Korea (Republic of)

ISSN 2190-5053 ISSN 2190-5061 (electronic) Springer Theses ISBN 978-981-99-6877-0 ISBN 978-981-99-6878-7 (eBook) https://doi.org/10.1007/978-981-99-6878-7 © The Editor(s) (if applicable) and The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 This work is subject to copyright. All rights are solely and exclusively licensed by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed. The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use. The publisher, the authors, and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty, expressed or implied, with respect to the material contained herein or for any errors or omissions that may have been made. The publisher remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. This Springer imprint is published by the registered company Springer Nature Singapore Pte Ltd. The registered company address is: 152 Beach Road, #21-01/04 Gateway East, Singapore 189721, Singapore Paper in this product is recyclable.

Supervisor’s Foreword

Mass Spectrometry (MS) is a key analytical technique for establishing molecular identification, providing qualitative as well as quantitative information about the molecule. Particularly, MS methods are of significant interest to researchers in metabolomics because decoding of the molecular identity is directly realizable by MS analysis, whereby potential diagnostic and/or prognostic biomarkers can be determined. Among the several MS techniques available, matrix-assisted laser desorption/ ionization (MALDI)-MS, which employs an organic matrix, has been acknowledged for its great analytical usefulness, detection sensitivity, and soft ionization since its first advent in 1988. Despite its benefits, critical challenges mainly originate from the organic matrix, which is inherently fragile in response to laser pulses, producing intense, unreproducible fragments in the low-mass region and thus impeding small molecule detection. MALDI is also incapable of yielding reproducible and quantitative MS results because of the “sweet spot”. To address these challenges, researchers suggested an organic-matrix-free methodology by making use of inorganic nanophotonic structures, which is referred to as laser desorption/ionization (LDI)-MS. However, in most cases, the nanostructure-based LDI-MS is still less efficient than the conventional MALDI-MS. In this doctoral thesis, Dr. Moon-Ju Kim designed and developed novel nanophotonic platforms for LDI-MS to ensure that the innovative LDI-active platforms can help meet the analytical demands on sensitivity, reproducibility, and quantification. Dr. Kim presented multiple strategies for improving desorption and ionization efficiencies through systematic explorations of the photo-thermal, electronic, and structural characteristics of the nanostructures, along with DFT-based computational analyses. The findings of this thesis further provide groundbreaking insights that shed light on LDI mechanisms. The thesis also contains proof-of-concept studies showing that newly developed nanostructures are effective in detecting small metabolites with low polarity and poor ionizability, suggesting that they have a great deal of potential for resolving the challenges that (MA)LDI-MS is currently facing.

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Supervisor’s Foreword

As her supervisor, I believe that this thesis will provide broad readers with a more comprehensive as well as deeper understanding of fundamental nanostructure properties, LDI mechanisms, and design rules of nanophotonic structures for high LDI-MS performance, which will contribute to the advancement of the LDI-MS technique. Seoul, Korea (Republic of) May 2023

Prof. Dr. Jae-Chul Pyun

Parts of this thesis have been published in the following journal articles: 1. Moon-Ju Kim, Tae Gyeong Yun, Joo-Yoon Noh, Jong-Min Park, Min-Jung Kang, Jae-Chul Pyun, Synergistic Effect of the Heterostructure of Au Nanoislands on TiO2 Nanowires for Efficient Ionization in Laser Desorption/Ionization Mass Spectrometry, ACS Appl. Mater. Interfaces 2019, 11, 22, 20509–20520. (Reproduced with Permission) 2. Moon-Ju Kim, Tae Gyeong Yun, Joo-Yoon Noh, Zhiquan Song, Hong-Rae Kim, Min-Jung Kang, Jae-Chul Pyun, Laser-Induced Surface Reconstruction of Nanoporous Au-Modified TiO2 Nanowires for In-Situ Performance Enhancement in Desorption and Ionization Mass Spectrometry, Adv. Funct. Mater. 2021, 31, 29, 2102475. (Reproduced with Permission) 3. Moon-Ju Kim, Tae Gyeong Yun, Joo-Yoon Noh, Min-Jung Kang, Jae-Chul Pyun, Photothermal Structural Dynamics of Au Nanofurnace for In-Situ Enhancement in Desorption and Ionization, Small 2021, 17, 49, 2103745. (Reproduced with Permission) 4. Moon-Ju Kim, Joo-Yoon Noh, Tae Gyeong Yun, Min-Jung Kang, Dong Hee Son, Jae-Chul Pyun, Laser-Shock-Driven In Situ Evolution of Atomic Defect and Piezoelectricity in Graphitic Carbon Nitride for the Ionization in Mass Spectrometry, ACS Nano 2022, 16, 11, 18284–18297. (Reproduced with Permission)

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Acknowledgements

First and foremost, I would like to express my heartfelt gratitude and admiration to my supervisor, Prof. Jae-Chul Pyun, for his unwavering support and encouragement throughout my PhD. At the beginning of my graduate studies, the lesson I learned from him about the significance of critical thinking has helped me tremendously in developing my capacity to think clearly and logically and in becoming a better problem solver in the scientific field, and for that, I am very grateful. Also, I do appreciate his tolerance and flexibility in letting me pursue studies and experiments that I found intriguing. All these experiences I have had with him have allowed me to mature as an independent researcher. I would like to thank the members of my advisory committee, Prof. HyungHo Park, Prof. Heon-Jin Choi, Prof. Seong-Ju Hwang at Yonsei University, and Prof. Dong Hee Son at Texas A&M University. I truly appreciate their time and effort in reviewing my PhD research and providing constructive feedback, which enabled me to complete and strengthen my thesis. I also want to thank all faculty members in the Department of Materials Science and Engineering at Yonsei University for their knowledge and teaching since my undergraduate days. I wish to express my sincere appreciation to everyone I have worked with and met at the NanoBio System Laboratory over the years. I was able to live a successful research life thanks to my colleagues, who comforted me in the hard times and celebrated with me in the fruitful moments. And thanks to all my friends in other labs who helped me out by lending me their lab’s tools and instruments. Finally, I would like to attribute all this glory to my parents and my sweet older sister for their endless love and support throughout the years. I am so fortunate to be a part of such a wonderful family. With all of my heart, I appreciate my parents for always doing their best for me.

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Acknowledgements

Once again, thanks to all of you who have helped shape me into the person I am today. Always remembering the warmth and kindness shown to me, I will try to grow as a better person who can give more love and help to others around me. May 2023

Moon-Ju Kim

Contents

1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1.1 Overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1.2 Motivation and Outline . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1 1 2 3

2 Backgrounds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 2.1 Organic Matrix-Assisted Laser Desorption Ionization (MALDI)-MS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 2.2 Nanostructure-Based Laser Desorption Ionization (LDI)-MS . . . . . . 7 2.3 Research Trends in Developing LDI-Active Nanophotonic Structure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13 3 Synergistic Effect of the Hybrid of Au Nanoislands on TiO2 Nanowires (Au-TNW) in Laser Desorption and Ionization . . . . . . . . . . . 3.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2 Experimental Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3 Results and Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.1 Fabrication and Characterization of Au-TNW Nanohybrid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.2 LDI-MS Analysis Results of Immunosuppressive Drugs . . . . 3.3.3 Mechanistic Insights into the Synergistic Ion Production . . . . 3.4 Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

15 15 16 17 17 18 22 25 27

4 In Situ Surface Reconstruction-Driven Desorption and Ionization Enhancement in Nanoporous Au-Modified TiO2 Nanowires Hybrid (npAu-TNW) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29 4.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29 4.2 Experimental Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30

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4.3 Results and Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.3.1 Fabrication and Characterization of npAu-TNW Nanohybrid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.3.2 Origin of the Enhancement in Desorption and Ionization . . . . 4.3.3 Application of npAu-TNW Nanohybrid for Neurotransmitter LDI-MS Analysis . . . . . . . . . . . . . . . . . . . 4.4 Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized ZnO Nanotube (AuNI-ZNT) for In Situ Enhancement in Desorption and Ionization . . . . . . . . . . . . . . 5.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.2 Experimental Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.3 Results and Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.3.1 Fabrication and Characterization of AuNI-ZNT Nanohybrid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.3.2 Strategies for Improving Desorption and Ionization Efficiencies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.3.3 LDI-MS Performance Enhancement Based on AuNI-ZNT Nanohybrid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.4 Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 In Situ Evolution of Atomic Defect and Piezoelectricity in Graphitic Carbon Nitride Nanosheet (G-C3 N4 NS) by Short Laser Pulse for the Ionization in Mass Spectrometry . . . . . . . . . . . . . . . . 6.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6.2 Experimental Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6.3 Results and Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6.3.1 Fabrication and Characterization of G-C3 N4 NS . . . . . . . . . . . 6.3.2 Influence of Nanosecond Laser-Pulse-Driven Shock Pressure on LDI-MS Performance . . . . . . . . . . . . . . . . . . . . . . . 6.3.3 G-C3 N4 NS as Efficient LDI-MS Substrate . . . . . . . . . . . . . . . 6.4 Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

31 31 34 44 45 46

49 49 50 51 51 53 63 66 68

69 69 70 72 72 76 89 90 92

7 Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 95

Chapter 1

Introduction

1.1 Overview Mass spectrometry (MS) is a method used for biomolecular identification based on the mass-to-charge ratio (m/z) of the analyte ion [1]. Among various MS methods, matrix-assisted laser desorption/ionization mass spectrometry (MALDIMS) is recognized as an analytical tool showing high detection sensitivity, which enables the rapid detection of intact molecules (soft ionization), leading to an easy interpretation of the MS results [2]. Thus, the m/z measurements obtained from experiments may be used to reliably identify molecules. MALDI-MS makes use of ultraviolet (UV) nanosecond laser pulses, and an organic matrix is employed to aid the analyte desorption and ionization. Matrix molecules absorb UV light very efficiently, leading to the simultaneous release of matrix and analyte species into the gas phase and, ultimately, the production of analyte ions [3–5]. However, MALDI-MS has a severe problem in that the organic matrix is so unstable to the UV laser that fragmented ions are always produced in the low m/ z range, and they are very powerful and unpredictable. This restricts its application only to large molecules, e.g., DNAs and proteins [2, 6, 7]. Moreover, non-uniform crystallization of the analyte-embedded matrix crystals produces large variances in mass peak intensities depending on the laser-irradiated position, resulting in poor data reproducibility and quantification [8, 9]. In order to resolve these issues, the organicmatrix-free methodology known as laser desorption/ionization mass spectrometry (LDI-MS) was introduced. This technique employs nanostructures for the desorption and ionization of small molecules. In LDI-MS, the laser-irradiated nanostructure rarely produces background interferences in the low mass range, which allows for a mass analysis of small molecules. In addition, data reproducibility and quantification in LDI-MS are considerably improved compared to MALDI-MS. As LDI-MS is currently less efficient than traditional MALDI-MS, developing novel nanostructures is urgently needed to improve the analytic performance.

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 M.-J. Kim, Laser Desorption Ionization Mass Spectrometry Based on Nanophotonic Structure: From Material Design to Mechanistic Understanding, Springer Theses, https://doi.org/10.1007/978-981-99-6878-7_1

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1 Introduction

1.2 Motivation and Outline In this dissertation, numerous different types of nanostructures were designed to attain less background noise, good reproducibility, and enhanced LDI efficiency. The characteristics of nanostructures, including photoabsorption, photo-thermal conversion, heat transfer, and photo-induced carrier generation, play a crucial role in the LDI process [10–17]. However, underlying LDI mechanisms are only limitedly available in the literature. The motivation of this dissertation is to investigate the origin of the enhancement in desorption and ionization with regard to nanostructure properties. It is anticipated that this approach will provide a deeper understanding of the mechanism behind the nanostructure-based LDI-MS. The dissertation is structured as follows: Chap. 2 introduces the academic background involving the basic principles of MALDI and nanostructure-based LDI essential for this dissertation. In Chap. 3, a nanohybrid composed of Au nanoislands and TiO2 nanowires (Au-TNW) is presented as an LDI nanostructure. A synergistic effect is attained between Au and TiO2 in desorption and ionization. We investigate the reason for the synergy with respect to the thermal and electronic properties of the nanohybrid, whereby nanohybrid contributions to individual desorption and ionization are assessed. In Chap. 4, nanoporous Au-modified TiO2 nanowires (npAu-TNW) nanohybrid is proposed as an LDI substrate for the LDI enhancement of neurotransmitters. We attribute the laser-induced in situ surface restructuring/ melting phenomenon that occurs in npAu-TNW to the origin of enhanced desorption and ionization. This structural dynamics as a result of laser exposure positively affect internal energy transfer between npAu and analytes and also lead to efficient analyte ionization by improved charge transfer reactions. Investigations of the laser-induced structural dynamics lead to new insights on the use of piezoelectric nanomaterials as a strategy for improving desorption and ionization efficiencies. Chapter 5 presents a nanohybrid of Au nanoislands and ZnO nanotubes (AuNI-ZNT) that contains a piezoelectric ZnO material. Photothermal structural dynamics of Au nanoislands are validated to be effective in enhancing desorption and ionization efficiencies. To the best of our knowledge, this study is the first report correlating laser-induced structural changes and piezoelectricity development in LDI-MS. High LDI-MS performance is verified by the analysis results of small metabolites of fatty acids and saccharides, which have been extremely difficult to detect in traditional MALDI-MS. In Chap. 6, we develop two-dimensional graphitic carbon nitride nanosheets (2D g-C3 N4 NS) as a promising, efficient LDI nanostructure. This chapter provides a systematic explanation of how laser shock waves affect increased ionization, focusing on the in-situ formation of atomic defects and piezoelectricity. Density functional theory (DFT) studies on Mulliken charges and dipole moments corroborate the roles played by defects and piezoelectricity in the enhanced ionization, complementing the results of the experimental investigations. The dissertation will conclude with Chap. 7, which summarizes the experimental results and findings. I believe that this dissertation will extend our knowledge, giving us more insights as to the design and development of efficient LDI nanostructures.

References

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References 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17.

Kumar N, Hoque M, Shahjaman M, Islam S, Mollah M, Haque N (2017) Biomed Res Int 2017 Wei J, Buriak JM, Siuzdak G (1999) Nature 399:243–246 Dreisewerd K (2003) Chem Rev 103:395–426 Knochenmuss R (2006) Analyst 131:966–986 Niehaus M, Soltwisch J (2018) Sci Rep 8:1–10 Siuzdak G (2004) J Lab Autom 9:50–63 Kim MJ, Park JM, Noh JY, Yun TG, Kang MJ, Pyun JC (2019) Rapid Commun Mass Spectrom 33:527–538 Park J-M, Kim M-J, Noh J-Y, Yun TG, Kang M-J, Lee S-G, Yoo BC, Pyun J-C (2020) J Am Soc Mass Spectrom 31:917–926 Kang MJ, Pyun JC, Lee JC, Choi YJ, Park JH, Park JG, Lee JG, Choi HJ (2005) Rapid Commun Mass Spectrom 19:3166–3170 Kim M-J, Park J-M, Yun TG, Noh J-Y, Kang M-J, Pyun J-C (2020) Chem Commun 56:4420– 4423 Kim M-J, Yun TG, Noh J-Y, Park J-M, Kang M-J, Pyun J-C (2019) ACS Appl Mater Interfaces 11:20509–20520 Wang J, Sun J, Wang J, Liu H, Xue J, Nie Z (2017) H Chem Commun 53:8114–8117 Ng K-M, Chau S-L, Tang H-W, Wei X-G, Lau K-C, Ye F, Ng AM-C (2015) J Phys Chem C 119:23708–23720 Kim MJ, Yun TG, Noh JY, Song Z, Kim HR, Kang MJ, Pyun JC (2021) Adv Funct Mater 31:2102475 Kim MJ, Yun TG, Noh JY, Kang MJ, Pyun JC (2021) Small 17:2103745 Song K, Cheng Q (2020) Appl Spectrosc Rev 55:220–242 Liu Q, Shi J, Jiang G (2012) TrAC. Trends Anal Chem 37:1–11

Chapter 2

Backgrounds

2.1 Organic Matrix-Assisted Laser Desorption Ionization (MALDI)-MS Mass spectrometry (MS) is an indispensable analytical technique whereby identification of the analytes of interest is accomplished by the detection of their ions under electric and/or magnetic fields. Molecular identities and structures are revealed by the separation of analyte ions based on their mass-to-charge ratios (m/z). In addition, the relative amount is informed by the mass peak intensity of the detected analyte ions proportional to their abundance. Therefore, the first step is the formation of analyte ions in MS analysis. Various ionization methods have been developed in the MS research field, for example, fast atom bombardment (FAB) [1], electrospray ionization (ESI) [2], and matrix-assisted laser desorption/ionization (MALDI) [3]. MALDI is a potent ionization method for detecting intact molecules without causing fragmentation and is used for mass spectrometric investigation of large molecules [4]. In MALDI, a UV nanosecond laser is used to desorb and ionize the analytes embedded in abundant organic matrix molecules. The MALDI process involves several cascading events, the first being initiated when rapid absorption of light energy from a laser pulse occurs (Fig. 2.1). Numerous complicated events such as energy absorption, distribution, and transfer, simultaneous release of matrix and analyte, and primary/secondary ionization are accompanied when the analyteembedded organic matrix is exposed to laser light, producing the analyte ions [5, 6]. However, due to the very intricate processes, the fundamental MALDI mechanisms are yet to be established and are still under debate. Several researchers proposed potential MALDI mechanisms, which typically involve two phases: primary and secondary ionization [7]. Neutral matrix molecules produce their initial ions through primary ionization, while analyte ions are obtained through secondary ionization. A widely recognized theory for primary ionization is excited-state proton transfer (ESPT). According to the ESPT model, photo-excited

© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 M.-J. Kim, Laser Desorption Ionization Mass Spectrometry Based on Nanophotonic Structure: From Material Design to Mechanistic Understanding, Springer Theses, https://doi.org/10.1007/978-981-99-6878-7_2

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Fig. 2.1 MALDI-MS. a Schematic representations of MALDI-MS. Reprinted with permission from Cho, Y. T. et al. Adv. Clin. Chem. 2015, 69, 209–254. Copyright 2015 Elsevier Inc. [6]. b Typical organic matrices used in MALDI-MS

matrix molecules are believed to transfer protons to analytes nearby and/or to ground-state matrices (Eqs. 2.1–2.3) [8]. M + hν → M ∗

(2.1)

M ∗ + A → (M − H )− + AH +

(2.2)

M ∗ + M → (M − H )− + M H +

(2.3)

where M and A are the matrix and the analyte, respectively. Excited matrix molecules are present in a hot, dense bath of neutral matrices, and they interact and collide with the analyte and matrix. During the collision, matrixdriven primary ions are formed inside a plume as intermediates that can further react with the remaining analyte, creating the analyte ions (i.e., secondary ionization). Thus, reactions between matrix and analyte and analyte ion formation are involved in the secondary ionization step, wherein gas-phase proton transfer between them mainly occurs. That is, MH + and (M–H)− intermediates after primary ionization ionize the analyte by protonating and deprotonating the analytes, respectively (Eqs. 2.4 and 2.5) [9].

2.2 Nanostructure-Based Laser Desorption Ionization (LDI)-MS

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M H + + A → M + AH +

(2.4)

(M − H )− + A → M + (A − H )−

(2.5)

Analyte protonation is thermodynamically favored when the proton transfer reaction from the protonated matrix to the analyte (Eq. 2.4) has a negative Gibbs free energy change (ΔG) < 0. In contrast, an analyte can be deprotonated by matrix ions when ΔG of the reaction in Eq. (2.5) is negative. The matrix-analyte proton transfer process is therefore dependent on the gas-phase proton affinity and basicity values of matrices and analytes (Table 2.1).

2.2 Nanostructure-Based Laser Desorption Ionization (LDI)-MS MALDI-MS is recognized for having a high detection sensitivity on the order of femtomole-picomole, together with a wide detection range. Analytes can be softly ionized by UV laser pulses, resulting in minimal analyte fragmentation and thus enabling molecular identification. Additionally, the ease of sample preparation, quick analysis (≈1–2 s), and high salt tolerance of the MALDI-MS make it a desirable analytical technique. However, MALDI-MS has some challenges. UV laser pulses produce intense and unpredictable fragmented ions of the organic matrix, particularly in the low mass area (3. We determined the LOD of the analyte whose concentration resulted in a mass peak with an S/N of 3.

3.3 Results and Discussion 3.3.1 Fabrication and Characterization of Au-TNW Nanohybrid The preparation process of the Au-TNW nanohybrid is depicted in Fig. 3.1, along with crystallographic structures for each step. KOH treatment etched the Ti surface, causing Ti–O–Ti bonds in TiO6 units to break and producing layered titanates of K2 Ti2 O4 (OH)2 [7, 8]. Water treatment removed excessive K+ and OH− ions that caused some of the Ti–O-K and Ti–OH bonds in layered titanate K2 Ti2 O4 (OH)2 to revert to Ti–O-Ti bonds. Some K+ ions in layered titanate K2 Ti2 O4 (OH)2 structures were also replaced by H+ ions. H2 Ti2 O4 (OH)2 was finally dehydrated through thermal annealing at 600 °C, resulting in the formation of TiO2 anatase and rutile phases. We controlled the Au nanoisland size by altering the Au film thickness: 1, 5, 10, and 15 nm, which generated the nanoislands with an average diameter of 34.6, 40.7, 91.5, and 123.5 nm, respectively, after solid-state dewetting (Fig. 3.2). Uniform spatial distributions of Au nanoislands were noticed without aggregates on the TNW, whereas severe aggregates of colloidal Au nanoparticles larger than 700 nm were observed that inevitably screened the photocatalytic-active TNW surface (Fig. 3.3). We obtained anatase, rutile, and layered titanate phases in TNW and Au-TNW, as seen in the HR-XRD patterns of Fig. 3.4. Note that the three crystal structures of anatase, rutile, and layered titanate were achievable only after thermal annealing. The presence of Au metal was confirmed in Au-TNW by the appearance of its

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3 Synergistic Effect of the Hybrid of Au Nanoislands on TiO2 Nanowires …

Fig. 3.1 Schematic depiction of the Au-TNW nanohybrid production process. At every stage of the synthesis, the crystal structure is shown

characteristic XRD peaks. We estimated a bandgap energy by using the Kubelka– Munk function [9], and pristine TNW and Au-TNW had a bandgap energy of 3.11 and 3.12 eV, respectively (Fig. 3.5). A notable absorption peak around 2.34 eV of Au-TNW supports the formation of Au nanoislands.

3.3.2 LDI-MS Analysis Results of Immunosuppressive Drugs We first selected the optimal morphology of the Au-TNW for high LDI-MS using four kinds of immunosuppressive drugs. We prepared the Au-TNW substrates at different Au film thicknesses of 1, 5, 10, and 15 nm, and their LDI efficiency was tested with four drugs. As observed in Fig. 3.6, we obtained the highest mass peak intensities of the four drugs with the use of Au-TNW synthesized with an Au thickness of 5 nm. Poor LDI results using the Au-TNW synthesized with 1-nm-thick Au film might arise from the insufficient Au amount for the LDI. Even though the Au-TNW synthesized with 10-nm-thick and 15-nm-thick Au films had a larger amount of Au, the large-sized Au screened UV laser radiation to the photocatalytic-active TNW (referred to as a shadow effect), which reduced the LDI efficiency. Thus, we here employed the Au-TNW nanohybrid prepared with 5-nm-thick Au film for efficient LDI-MS.

3.3 Results and Discussion

19

Fig. 3.2 Surface morphologies of Au-TNW that were created using Au films of 1, 5, 10, and 15 nm in thickness

20

3 Synergistic Effect of the Hybrid of Au Nanoislands on TiO2 Nanowires …

Fig. 3.3 Colloidal Au nanoparticles on a TNW substrate arranged in an irregular spatial pattern Fig. 3.4 HR-XRD patterns of the following substrates: Ti treated with KOH, Ti treated with water, pure TNW, and Au-TNW

Notable enhancement in LDI-MS was achieved with Au-TNW when comparing the results with organic matrix (CHCA) and nanostructures (Au nanoparticles, pristine TNW, Au nanoparticles-TNW, and Au-TNW) (Fig. 3.7). CHCA hardly protonated all four drugs, and Au nanoparticles were also ineffective in ionizing the drugs. Thermal energy generated by UV laser irradiation of the Au nanoparticles barely contributed to the drug ionization. On the contrary, four drugs were ionized as K+ adducts with TNW, indicating that TiO2 had a stronger ionizing capability than Au

3.3 Results and Discussion

21

Fig. 3.5 Tauc plots of the substrates. The bandgap was calculated by applying the Kubelka–Munk transformation on the UV–vis diffuse reflectance spectra

Fig. 3.6 The LDI-MS data for four immunosuppressive drugs based on Au-TNW produced with varying Au film deposition thickness. The excellent LDI-MS findings for the four drugs were achieved using Au-TNW produced with an Au film thickness of 5 nm

owing to the photo-excited electrons and holes. Au nanoparticle-TNW exhibited increased mass peak intensities when compared with pristine TNW, suggesting that the interface between Au nanoparticle and TNW resulted in a positive effect on the drug ionization. Further enhancement was noticed for the Au-TNW. The S/N ratio of the mass peaks increased when using Au-TNW compared with TNW and Au nanoparticle-TNW, respectively. Contrary to Au-TNW, Au nanoparticle-TNW

22

3 Synergistic Effect of the Hybrid of Au Nanoislands on TiO2 Nanowires …

Fig. 3.7 Outcomes of LDI-MS analysis on four immunosuppressants. Organic matrix of a CHCA and nanostructures of b Au nanoparticles, c TNW, d Au nanoparticle-TNW, and e Au-TNW

constrained the TNW surface by Au nanoparticle aggregates and led to poor LDI-MS results. These mass analysis results indicate that semiconducting TNW was crucial for the analyte ionization, while metallic Au alone was not that effective in LDIMS. However, we observed the LDI improvement with Au-TNW compared with pristine TNW, revealing that Au contributed more to the analyte desorption rather than ionization, and thus the nanohybrid of Au and TNW enhanced both desorption and ionization processes, leading to synergy in LDI-MS performance. We also demonstrated the Au-TNW-based quantitative mass analysis (Fig. 3.8), supporting the analytical validity.

3.3.3 Mechanistic Insights into the Synergistic Ion Production Improved photocatalytic ionization due to Schottky heterojunction between TiO2 and Au and higher heat release from Au nanoislands are the two main reasons we gave for the LDI improvement with the Au-TNW nanohybrid, as depicted in Fig. 3.9.

3.3.3.1

Photocatalytic Behavior of Au-TNW Nanohybrid

We estimated the ionizing capability of nanostructures by assessing their photocatalytic efficiencies. Chromophoric group in methylene blue can be destroyed by the electrophilic attack of a hydroxyl radical (·OH) generated by water oxidation

3.3 Results and Discussion

23

Fig. 3.8 Quantitative LDI-MS using Au-TNW. The four drugs were measured with great linearity and sensitivity

Fig. 3.9 Positive synergistic impact of Au-TNW nanohybrid on LDI performance

by hole under UV light [10] (Fig. 3.10). The photocatalytic efficiency of Au-TNW was compared with that of Au under UV irradiation (365 nm). Au was prepared by annealing a thin Au film with a 5 nm thickness sputtered on a Ti substrate, which exhibited negligible photocatalytic efficiency (Fig. 3.11). As opposed to this, TNW effectively degraded methylene blue, and further notable photocatalytic degradation was obtained with Au-TNW. This might be attributed to the presence of a Schottky barrier at the Au/TNW interface that consequently prevented recombination of photoexcited charge carriers [11–15]. As depicted in Fig. 3.12, a Schottky barrier developed at the Au/TNW heterojunction due to the difference in Fermi energy levels between Au and TiO2 , which in turn bent TiO2 band edges upward toward the interface and modulated the carrier motions. As a result, the Au-TNW nanohybrid improved photocatalytic activity by impeding charge carrier recombination.

24

3 Synergistic Effect of the Hybrid of Au Nanoislands on TiO2 Nanowires …

Fig. 3.10 Methylene blue photodegradation using an Au-TNW nanohybrid photocatalyst

Fig. 3.11 Comparison of photocatalytic activities of the Au, TNW, and Au-TNW

Fig. 3.12 The creation of a Schottky barrier and the segregation of charge carriers in Au-TNW

3.3.3.2

Thermal Behavior of the Au-TNW Nanohybrid

Using a DSC instrument and histidine as a phase transition indicator, we examined the heat transfer efficiency of TNW, Au nanoparticles, and Au-TNW. Heat flow as

3.4 Summary

25

a function of temperature was observed in relation to the thermal transition of histidine. As shown in Fig. 3.13, histidine started to melt at 278.1 °C and had a melting enthalpy of 598.4 mJ. When histidine was mixed with the TNW, changes in melting temperature and enthalpy were barely observable, which is a stark contrast with the case where histidine was mixed with Au nanostructures (Au nanoparticles and AuTNW). Melting temperature and enthalpy notably decreased when metallic Au was added to a sample, suggesting that Au was able to rapidly trigger the endothermic process of histidine melting. Reduced melting temperature and enthalpy occurred from the heat being transmitted from the Au surface to the histidine analyte much more quickly than from the TiO2 surface because Au had a higher thermal conductivity than TiO2 . These efficient heat transfers imply that analyte desorption could be much preferred with Au metal. We verified the synergistic effect of the Au-TNW nanohybrid on the LDI-MS (Fig. 3.14). With Au nanoparticles and TNW, the S/N ratio was respectively calculated as 55.0 and 431.8, demonstrating that TNW was more effective in the analyte ionization than Au. Remarkable thermal properties of Au thus made it effective in analyte desorption rather than ionization [16]. Nanohybrid structures of Au nanoparticles-TNW and Au-TNW resulted in increased peak intensities compared to pristine TNW, confirming the synergy between Au and TNW. Further LDI enhancement was achievable with Au-TNW, which was attributed to improved analyte desorption owing to Au and also improved analyte ionization owing to TNW and a Schottky heterojunction at the Au/TNW interface.

3.4 Summary We developed an Au-TNW nanohybrid for the LDI enhancement, which was verified by analyzing four immunosuppressors. S/N ratios of mass peaks based on Au-TNW were much higher as compared to pure TNW and Au nanoparticle-TNW. We also confirmed the quantification, with Au-TNW showing a high linearity. As evidenced by photocatalytic results, semiconducting TNW contributed to analyte ionization, while Au assisted analyte desorption by enhancing heat transmission, as demonstrated in DSC thermal analysis. Further, we attributed the LDI enhancement of the Au-TNW nanohybrid to the effective charge separation and thereby reduced recombination by a Schottky heterojunction at the Au/TNW interface.

26

3 Synergistic Effect of the Hybrid of Au Nanoislands on TiO2 Nanowires …

Fig. 3.13 DSC thermal study of histidine in the presence of several nanostructures: TNW, Au nanoparticles, and Au-TNW

References

27

Fig. 3.14 LDI-MS of histidine with distinct nanostructures of the TNW, Au nanoparticles, and Au-TNW

References 1. Yonezawa T, Kawasaki H, Tarui A, Watanabe T, Arakawa R, Shimada T, Mafune F (2009) Anal Sci 25:339–346 2. Schürenberg M, Dreisewerd K, Hillenkamp F (1999) Anal Chem 71:221–229 3. Joh S, Na H-K, Son JG, Lee AY, Ahn C-H, Ji D-J, Wi J-S, Jeong MS, Lee S-G, Lee TG (2021) ACS Nano 15:10141–10152 4. Kim M, Park J-M, Yun TG, Noh J-Y, Kang M-J, Pyun J-C (2018) ACS Appl Mater Interfaces 10:33790–33802 5. Müller M, Schiller J, Petkovi´c M, Oehrl W, Heinze R, Wetzker R, Arnold K, Arnhold J (2001) Chem Phys Lipids 110:151–164 6. Braun S, Kalinowski H-O, Berger S (1996) VCH Weinheim 7. Xie J, Wang X, Zhou Y (2012) J Mater Sci Technol 28:488–494

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8. Kim J-I, Park J-M, Hwang S-J, Kang M-J, Pyun J-C (2014) Anal Chim Acta 836:53–60 9. Lin H, Huang C, Li W, Ni C, Shah SI, Tseng Y-H (2006) Appl Catal 68:1–11 10. Yang C, Dong W, Cui G, Zhao Y, Shi X, Xia X, Tang B, Wang W (2017) RSC Adv 7:23699– 23708 11. Lin Z, Wang X, Liu J, Tian Z, Dai L, He B, Han C, Wu Y, Zeng Z, Hu Z (2015) Nanoscale 7:4114–4123 12. Tian J, Zhao Z, Kumar A, Boughton RI, Liu H (2014) Chem Soc Rev 43:6920–6937 13. Devi LG, Kavitha R (2016) Appl Surf Sci 360:601–622 14. Moon SY, Song HC, Gwag EH, Nedrygailov II, Lee C, Kim JJ, Doh WH, Park JY (2018) Nanoscale 10:22180–22188 15. Jung H, Jung J, Kim Y-H, Kwon D, Kim B-G, Na HB, Lee HH (2018) BioChip J 12:249–256 16. McLean JA, Stumpo KA, Russell DH (2005) J Am Chem Soc 127:5304–5305

Chapter 4

In Situ Surface Reconstruction-Driven Desorption and Ionization Enhancement in Nanoporous Au-Modified TiO2 Nanowires Hybrid (npAu-TNW)

4.1 Introduction The three classes of inorganic materials have inspired the creation of nanostructures for LDI-MS: (1) carbon-based nanomaterial [1–4], (2) semiconductor [5–7], and (3) metal [8–10]. They do this by engaging in a number of laser-induced processes— including photoabsorption, photothermal conversion, heat transfer, and the production of photo-induced charge carriers—that ultimately help in the desorption and ionization of the analyte [11–14]. TiO2 has been frequently employed for LDI-MS among other nanostructures as its bandgap energy is 3.2 eV, which is in tune with the UV laser’s (355 or 337 nm) operating wavelength in an LDI-MS device. Although promising, TiO2 application to LDI-MS has been hampered by two major obstacles: photoexcited carriers recombine at a rapid rate, and the photothermal effect is low. A hybrid system composed of TiO2 and metal has been recently designed as a substitute method for attaining an effective LDI, which has an impact on the (1) electronic and (2) thermal characteristics: a Schottky barrier at the TiO2 /metal interface lengthens the lifespan of photo-excited carriers, and (2) the outstanding thermal conductivity and low specific heat of metals improve the efficiency of heat transfer. In this chapter, we present nanoporous Au-modified TiO2 nanowires (npAuTNW) nanohybrid for the advancement of LDI-MS. High LDI-MS was revealed by analyzing the three types of neurotransmitters (norepinephrine, serotonin, and dopamine). Notable LDI improvement was achievable with npAu-TNW compared with TNW and non-porous Au nanoisland-modified TNW (Au-TNW) nanohybrid, which was explained by the surface restructuring/melting phenomenon upon laser pulses. Laser-induced structural changes in the nanostructures reportedly originated from their photothermal effect, greatly affected by their surface characteristics like This chapter is an edited version of the following published journal article: Moon-Ju Kim, Tae Gyeong Yun, Joo-Yoon Noh, Zhiquan Song, Hong-Rae Kim, Min-Jung Kang, and Jae-Chul Pyun, Adv. Funct. Mater. 2021, 31, 2102475. Copyright 2021 Wiley–VCH GmbH. © The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 M.-J. Kim, Laser Desorption Ionization Mass Spectrometry Based on Nanophotonic Structure: From Material Design to Mechanistic Understanding, Springer Theses, https://doi.org/10.1007/978-981-99-6878-7_4

29

30

4 In Situ Surface Reconstruction-Driven Desorption and Ionization …

as porosity and area. A highly porous npAu structure with an enlarged surface area enabled efficient photothermal conversion of the npAu surface, which caused the surface to be noticeably reconstructed by laser exposure. Adsorbed analytes received more of the internal energy released from the npAu surface at the time of structural rearrangement of npAu, promoting analyte desorption. Additionally, defect-related trap states were introduced in situ inside the TNW bandgap due to the distortion of the TNW lattice interfaced with restructuring npAu. These defects caused carriers to be trapped, lowering their recombination and thus increasing ionization efficiency.

4.2 Experimental Methods Synthesis of npAu-TNW Nanohybrid Wet-corrosion synthesis was used to create TNW, as has been described before [8]. Sequential sputtering of Au and Ag thin films on the TNW was followed by thermal dewetting to create Au–Ag nanoalloys on the TNW. In order to create the npAu structure, the dewetted Au–Ag nanoalloys were then submerged in strong nitric acid for a chemical dealloying procedure for 5 min at ambient temperature. After dealloying, to eliminate the acid, the nanostructure was given a thorough water wash. Nanostructure Characterization The surface morphology of the npAu-TNW hybrid was characterized using FE-SEM. Element analysis via EDS was carried out. Using HR-XRD, we were able to learn about the crystal structure. The photocatalytic efficiency of the substrates (TNW, Au-TNW, and npAuTNW) was compared in terms of surface wettability change. Substrates were modified to be hydrophobic by immersing them in a solution of trimethoxy(3,3,3trifluoropropyl)silane (FAS, 2 vol%) for 2 h at room temperature. Thereafter, substrate surfaces were UV-irradiated using a UV lamp (365 nm) to analyze their photocatalytic behaviors. Variation in surface hydrophobicity was evaluated by measuring the water contact angle (WCA) at room temperature using a Smartdrop instrument. We analyzed the thermal properties using a DSC instrument. At a heating rate of 10 °C min−1 , DSC was used to observe the phase change of tyrosine in the presence of nanostructures heated from 175 to 400 °C. Laser-Induced Surface Restructuring/Melting Laser-induced structural alterations of the nanostructures were driven in an LDI-MS instrument (Autoflex Max mass spectrometer, Bruker Daltonics, MA, USA) equipped with a nanosecond laser (smartbeam laser). The laser attenuator, offset, and range were all adjusted to 40%, 40%, and 10%, respectively, to provide an estimated 28 μJ of total laser energy. Flexcontrol software was used to programmatically adjust the relative laser intensity from 0 to 100% while staying inside the safe operating range.

4.3 Results and Discussion

31

FE-SEM was used to assess the degree to which the nanostructures’ surfaces (TNW, Au-TNW, and npAu-TNW) were remodeled or melted by the laser. The resultant lattice distortions were analyzed using HR-XRD. Raman spectroscopy using an Ar-ion laser was used to investigate the surface state variations. Using a UV–vis spectrometer, we measured the diffuse reflectance, and then we converted the data using the Kubelka–Munk function to determine the optical bandgap. Using an Al-Kα X-ray source, XPS was used to investigate the chemical composition and valence state of surfaces. LDI-MS Nanostructures (TNW, Au-TNW, and npAu-TNW) and organic matrices (α-cyano4-hydroxycinnamic acid (CHCA) and 2,5-dihyroxybenzoic acid (DHB)) were subjected to mass spectrometry. The neurochemicals were mass-analyzed using an Autoflex Max mass spectrometer (Bruker Daltonics, MA, USA). We also prepared artificial cerebrospinal fluid (aCSF) to examine the practical feasibility of using npAu-TNW as an LDI-active nanostructure. The aqueous aCSF contained NaCl (125 mM), KCl (2.5 mM), MgCl2 (1 mM), NaH2 PO4 (1.25 mM), CaCl2 (2 mM), NaHCO3 (25 mM), and D-glucose (25 mM), wherein the neurochemical analytes were spiked.

4.3 Results and Discussion 4.3.1 Fabrication and Characterization of npAu-TNW Nanohybrid Figure 4.1 illustrates the fabrication procedure of the npAu-TNW nanohybrid. TNW was prepared via a wet-corrosion process, as previously reported [8], and its surface was functionalized with npAu through an alloying and dealloying process [15, 16]. Au–Ag alloy nanoislands were created by thermally dewetting thin sputtered coatings of Au and Ag. Note that Au and Ag are completely miscible at any composition due to the same crystal structure of face-centered cubic, similar lattice constants, and high interdiffusion rates, resulting in an Au–Ag alloy phase. The extremely corrosive HNO3 was used to selectively dissolve away the Ag component of the alloy, forming a sponge-like open-cell foam morphology of npAu. The surface morphology of the npAu-TNW product at various Au/Ag ratios was observed (Fig. 4.2). A 5 nm Au film and 2.5, 5, 10, and 15 nm Ag films were sputtered onto TNW to create the various nanostructures, followed by heat annealing (alloying) and then dealloying. The nanoporous morphology of npAu was rarely discernible when created with a small amount of Ag (2.5 and 5 nm), which might be attributable to the pore coalescence [17]. Thus, it was decided to make Ag thicker than 10 nm since its amount was essential for npAu creation. As seen in Fig. 4.3a, the npAu with nanoscale pores and Au ligament was created after dealloying the alloy nanoislands of Au (thickness of

32

4 In Situ Surface Reconstruction-Driven Desorption and Ionization …

5 nm)–Ag (thickness of 10 nm). EDS elemental analysis verified that only Ag was selectively dissolved from the alloy nanoislands (Fig. 4.3b). For the alloy nanoislands fabricated on the TNW surface (AuAg-TNW; before dealloying), the Au and Ag concentrations were 0.16 and 0.33 at%, respectively, with a composition ratio of almost 1:2. After dealloying with HNO3 , npAu-TNW contained Au of 0.14 at% and Ag of 0.06 at%, revealing that highly reactive Ag was selectively etched without suffering a massive loss of Au. Selective Ag loss from the Au–Ag alloy nanoislands was further corroborated by EDS maps of npAu-TNW (Fig. 4.3c). We estimated the Au surface area of non-porous Au nanoislands and the npAu supported on the TNW (Au-TNW and npAu-TNW, respectively) with the use of 4-aminothiophenol (4-ATP) which could be chemisorbed on the Au surface through Au-thiol interaction [18]. Similar to npAu-TNW, Au-TNW was fabricated, but only without chemical dealloying. The 4-ATP molecules (30 nmol) were incubated with

Fig. 4.1 Process flow diagram for producing npAu-TNW nanohybrid

Fig. 4.2 Surface morphologies of synthetic npAu-TNW made with varying amounts of Au and Ag. Deposits of Au (5 nm thick) and varying concentrations of Ag were used to fabricate all of the nanostructures

4.3 Results and Discussion

33

Fig. 4.3 Substrate characterization. a Surface morphologies and b EDS elemental analyses before and after dealloying. c EDS maps of the npAu-TNW. d Absorption of UV–vis light by unbound 4-ATP on the substrates of TNW, Au-TNW, and npAu-TNW produced with varying Au/Ag ratios. e Coverage of 4-ATP on the Au surface of Au-TNW and npAu-TNW produced with varying Au/ Ag ratios. f HR-XRD patterns of the substrates

nanostructure substrates of TNW (control), Au-TNW, and npAu-TNW with a crosssectional area of 0.20 cm2 . The amount of 4-ATP unbound onto the substrate surface was estimated by measuring the absorbance at the 4-ATP-characteristic λmax of 258 nm with UV–vis spectrometry. As indicated in Fig. 4.3d, the absorbance of 4-ATP unbound on the TNW was nearly identical to that of initially added 4-ATP, revealing little adsorption of 4-ATP onto the TNW surface. As opposed to this, the absorbance decreased for Au-TNW and npAu-TNW, demonstrating the chemisorption of 4-ATP molecules on the Au surface. Notably, a larger amount of 4-ATP covered the surface of npAu prepared at an Au/ Ag ratio of 1:2 and 1:3 that had a greater npAu surface area. Using the Beer-Lambert law, we determined the quantity of 4-ATP that was adsorbed onto the substrate (Table 4.1). Given the 4-ATP molecular footprint of 0.20 nm2 per molecule [19, 20], the Au surface area was estimated to be 8.47 cm2 (Au-TNW), 9.91 cm2 (npAu-TNW synthesized at Au:Ag = 1:1), 13.7 cm2 (npAu-TNW synthesized at Au:Ag = 2:1), 21.7 cm2 (npAu-TNW synthesized at Au:Ag = 1:2), and 24.1 cm2 (npAu-TNW synthesized at Au:Ag = 1:3), which were markedly increased in comparison to the cross-sectional substrate area of 0.20 cm2 (Fig. 4.3e). In addition, npAu (synthesized at Au:Ag = 1:2 and 1:3) had a even greater Au surface area than non-porous Au

34

4 In Situ Surface Reconstruction-Driven Desorption and Ionization …

Table 4.1 Quantity of 4-ATP molecules adsorbed on a surface

Substrate

Substrate-adsorbed 4-ATP (nmol)

TNW

0.72

Au-TNW

7.03

npAu-TNW (Au:Ag = 1:1)

8.23

npAu-TNW (Au:Ag = 2:1) 11.4 npAu-TNW (Au:Ag = 1:2) 17.9 npAu-TNW (Au:Ag = 1:3) 20.1

nanoislands (Au) and the npAu (synthesized at Au:Ag = 1:1 and 2:1). The HR-XRD patterns in Fig. 4.3f corroborated that TNW, Au-TNW, and npAu-TNW contained both anatase and rutile TiO2 phases.

4.3.2 Origin of the Enhancement in Desorption and Ionization 4.3.2.1

Photocatalytic Behavior of npAu-TNW Nanohybrid

We estimated the ionizing capability of the nanostructures in aspect of their photocatalytic activity [21]. Photocatalytic efficiency was determined by measuring the WCA variations on the FAS-functionalized hydrophobic substrates of TNW, AuTNW, and npAu-TNW as the UV-irradiation time increased [22]. The hydrophobic fluoroalkyl chains of FAS on the substrates were cleaved by radicals generated by photocatalytic TiO2 under UV light, resulting in a change in the surface wettability (Fig. 4.4a). As presented in Fig. 4.4b, the WCAs on the TNW, Au-TNW, and npAuTNW were 159.1°, 161.3°, and 163.4°, respectively, immediately after FAS modification. After exposing FAS-modified substrates to UV light, the fluoroalkyl chain was photocatalytically cleaved, rendering the surfaces more hydrophilic. With increasing UV-irradiation time, the water droplets on npAu-TNW were progressively wetted, whereas drops on TNW and Au-TNW almost perfectly preserved their spherical forms (Fig. 4.5). We further compared the photocatalytic efficiencies by analyzing the photocatalytic reaction kinetics, assuming a pseudo-first-order reaction. As observed in Fig. 4.4c, the reaction rate constant (kr ) for npAu-TNW photocatalyst was much greater than that for TNW and Au-TNW. Schottky barrier formation at the Au/ TiO2 interface and the prohibition of carrier recombination contributed to the higher kr of Au-TNW relative to TNW [8]. Moreover, npAu-TNW had a higher photocatalytic efficiency than Au-TNW, which can be explained by an enlarged surface area of npAu (Fig. 4.4d) [23, 24]. Interconnected npAu also facilitated the transport of the photo-induced electrons to the exposed surface where FAS molecules were

4.3 Results and Discussion

35

Fig. 4.4 Activity of TNW, Au-TNW, and npAu-TNW as a photocatalyst. a Surface wettability alterations caused by TiO2 photocatalysis on FAS-treated surfaces. b WCA variations in TNW, Au-TNW, and npAu-TNW. c Kinetics of fluoroalkyl chain cleavage from FAS molecules using photocatalysis. d Enhanced photocatalysis with npAu-TNW compared to Au-TNW

Fig. 4.5 Changes in WCA as a function of UV irradiation duration for TNW, Au-TNW, and npAuTNW substrates. The FAS hydrophobic agent was applied to all the nanostructures’ surfaces

deposited. Accordingly, numerous FAS molecules encountered the npAu surface more frequently, resulting in the hydrophobic groups being efficiently cleaved from the npAu surface.

4.3.2.2

Thermal Behavior of npAu-TNW Nanohybrid

According to several published papers, heat is the primary factor in initiating the desorption of analytes from the nanostructure surface in LDI-MS [25]. Considering that metal has outstanding thermal properties, including high photoabsorption, efficient photo-thermal conversion, low specific heat, and high thermal conductivity, efficient analyte desorption from the nanometal surface is highly anticipated [26,

36

4 In Situ Surface Reconstruction-Driven Desorption and Ionization …

27]. The surface of a nanometal may be heated to an appropriate level by laser irradiation, which can be swiftly transferred to the surface-adsorbed analytes and thus lead to their desorption. In this section, before considering the laser application, we first investigated the heat transfer efficiency of nanostructures utilizing a DSC instrument to determine their contribution to the thermally driven desorption process. Figure 4.6a presents the DSC thermograms of the tyrosine and the tyrosine mixed with nanostructures. The peak temperature associated with the tyrosine melting was 311.98 °C and the melting enthalpy was 539.77 mJ (Fig. 4.6b). Tyrosine, when combined with TNW, had almost identical melting temperature and enthalpy values to those of tyrosine alone. With the Au-containing TNW (Au-TNW and npAu-TNW), a clear difference in the melting response was observed in contrast to the prior result of the TNW. The enthalpy change and melting transition temperature of tyrosine were both decreased, revealing that the endothermic melting event was promoted by including Au in the sample. Because the thermal conductivity of the Au is higher than that of the TiO2 , heat transfer from the Au to the tyrosine analyte occurred more rapidly than that from the TiO2 to the analyte, favoring the melting reaction. Moreover, the increased surface area of the npAu was responsible for the additional reduction in tyrosine melting temperature (273.95 °C) and enthalpy (285.72 mJ) with the npAuTNW as compared with the Au-TNW. Thus, the melting transition was aided by the open-cell foam, sponge-like npAu morphology, which enabled the tyrosine to be repeatedly adsorbed on the surface.

Fig. 4.6 Tyrosine in TNW, Au-TNW, and npAu-TNW mixtures subjected to DSC. a DSC thermograms. b Changes in the melting point of tyrosine and the enthalpy of its initiation. c Activation energy for tyrosine melting estimated using Arrhenius plots. d Extent of the melting reaction

4.3 Results and Discussion

37

We studied the kinetics of the tyrosine melting process with the use of an DSC instrument to further compare the heat transfer efficiency of the nanostructures [28, 29]. The kinetics of the thermally driven tyrosine melting were investigated based on the Borchardt and Daniels method for calculating thermal parameters [30]. This method is based on the assumption that the reaction kinetics are described by the general rate equation and the n-th-order kinetics (Eq. 4.1): dα = k(T )[1 − α]n dt

(4.1)

is the reaction rate (s−1 ), α is the fractional conversion, k(T ) is the rate where dα dt constant at temperature T, and n is the reaction order. This method also assumes the Arrhenius behavior (Eq. 4.2): ) ( Ea k(T ) = Z exp − kB T

(4.2)

where E a is the activation energy (J), Z is the pre-exponential factor (s−1 ), and k B is the Boltzmann constant. Kinetic parameters were calculated by solving the aforementioned equations with a multiple linear regression. In Fig. 4.6c, logarithms of k(T ) were plotted against the temperature reciprocal, 1/T, to obtain the activation energy E a , which was represented as the slope of the straight line in a plot. The E a barrier for the tyrosine melting process was greatly reduced by npAu-TNW, which allowed for more direct and efficient heat transport to the tyrosine. In addition, the extent of reaction (%) was estimated, and the 50% melting temperature (T50% ) was far lower than that of other nanostructures (Fig. 4.6d). The thermal parameters are summarized in Table 4.2. These findings from the thermal study show that npAu is capable of efficient heat transmission, making it a powerful tool in analyte desorption. Table 4.2 Measured thermal parameters of the tyrosine melting with TNW, Au-TNW, and npAuTNW Tyrosine only

TNW with Tyrosine

Au-TNW with Tyrosine

npAu-TNW with Tyrosine

Tonset (o C)

303.57

302.15

286.51

273.95

Heat of reaction (ΔH, mJ)

539.77

530.91

338.99

285.72

Activation energy (Ea , kJ/mol)

1031.75 ± 109.86

1136.82 ± 184.21

713.71 ± 26.96

567.79 ± 10.03

T50% (o C)

300.41

300.59

283.06

267.79

38

4.3.2.3

4 In Situ Surface Reconstruction-Driven Desorption and Ionization …

Laser-Induced in Situ Structural Changes in npAu-TNW Nanohybrid

Short laser pulses may induce surface restructuring/melting of the nanostructures, which reportedly aids in analyte desorption [31–33] (Fig. 4.7a). Photoabsorption and photothermal conversion of nanometals give rise to their structural alterations following laser pulses [34]. As can be seen in Fig. 4.7b, the degree of structural changes differed in nanostructures, and the npAu of the npAu-TNW was severely distorted while the TNW retained its original shape. As the laser energy was increased, we could see distinct structural differences between the TNW, AuTNW, and npAu-TNW (Fig. 4.8). Structural changes in the TNW were noticed at a significantly high energy of >25.2 μJ, while a lower energy of 18.2 μJ was required to result in surface reconstruction of the non-porous Au in the Au-TNW. Even at the weakest laser power, the npAu in the npAu-TNW was readily deformed. At a laser energy of 15.4 μJ, the pores in the npAu started to collapse, and at higher laser levels, the npAu melted and resolidified, creating severe aggregates. As the npAu frame had a lesser thickness than the diameter of the non-porous Au, uniform heating was easily achieved in the npAu by laser pulses, which in turn reduced the amount of laser energy required to induce restructuring or melting in the material [35].

Fig. 4.7 Structural alterations in the npAu-TNW by laser pulses. a Schematic depiction of the laserinduced surface restructuring/melting in npAu-TNW as a result of laser heating. b Laser-induced structural modifications in TNW, Au-TNW, and npAu-TNW. c Evaluation of TNW, Au-TNW, and npAu-TNW desorption efficiency. Methylthioninium served as a reliable thermometer for us

4.3 Results and Discussion

39

Fig. 4.8 Laser-induced surface restructuring/melting of the TNW, Au-TNW, and npAu-TNW under increasing laser energy

Laser-induced surface reconstruction reportedly exerts a positive effect on the analyte desorption, which can be explained in the aspect of thermodynamics. The npAu-TNW (system) is able to effectively and spontaneously transfer heat to the analytes (surroundings) when subjected to an external laser because of the strong photoabsorption and high photo-thermal conversion of the npAu (ΔStotal > 0, ΔS denotes the entropy change, and total denotes both system and surroundings). Moreover, the npAu instantaneously works on the analytes (surroundings) during npAu restructuring; that is, the internal energy of the npAu-TNW (system) is transferred to the analytes (surroundings). Therefore, analyte desorption is aided by the efficient transfer of heat as well as internal energy from the npAu-TNW (system) to the analytes (surroundings). Laser-induced structural changes in npAu were the most noticeable and hence facilitated analyte desorption more than in other nanostructures. To validate the enhanced desorption in the npAu-TNW, we evaluated the desorption efficiency using methylene blue (methylthioninium chloride) as a probe [36]. We used methylthioninium as a thermometer cation to gauge the desorption efficiency. As shown in Fig. 4.7c, methylthioninium mass peak intensity was dramatically increased with npAu-TNW compared to TNW and Au-TNW. These desorption results clearly verify that the npAu showing the most effective laser-induced surface restructuring/ melting behavior accelerated the desorption in LDI-MS. Laser-induced surface restructuring/melting event also benefited the ionization procedure in LDI-MS. Thermal deformation of the npAu brought to distortions in the surrounding TNW lattice during surface alterations in the npAu. Figure 4.9a shows the HR-XRD patterns of the npAu-TNW in accordance with the laser energy, and the strain developed in the TNW was investigated using the XRD data. The variation in the crystallographic parameters is summarized in Table 4.3. As shown in Fig. 4.9b, Compressive volumetric strain (−7.32%) was detected in tetragonalstructured anatase following 25.2 μJ-laser irradiation, with compression along the caxis (−7.89%) and expansion in the ab-plane (0.264%). However, increasing the laser

40

4 In Situ Surface Reconstruction-Driven Desorption and Ionization …

power did not cause any discernible lattice deformation in the rutile. The basic unit of both anatase and rutile is the TiO6 octahedron, which consists of four equatorial Ti–O links (which are shorter) and two axial Ti–O bonds (which are longer). As a result, the TiO6 unit has a softer (more deformable) axial orientation than an equatorial one. [37]. Rutile has two orthogonal soft axes: a half of soft axis along the [110] direction and the other half along the [1–10] direction. Thus, no overall soft axis exists in rutile TiO2 . The [001] direction is the only orientation in which anatase’s soft axis may be found, which makes the anatase possess a larger void volume along the [001]-axis and therefore possess excellent compressibility in that direction [38]. Thus, lattice distortion was more prominent in the deformable anatase than in the rutile. Oxygen vacancy production in metal oxides is said to be the consequence of lattice strain linked with the atomic and electronic structural features [39, 40]. Removal of an oxygen atom from the TiO2 leaves behind an undercoordinated Ti atom, which causes the Ti atom to move away from the oxygen vacancy and the neighboring O atom to move toward the defect. These atomic movements naturally affect the TiO2 structure [41, 42]. On the basis of the XRD data, we determined the degree of distortion of TiO6 octahedra in anatase and rutile as a function of laser energy, and the octahedra structure was visualized using the visualization for electronic and structural analysis (VESTA) program. Crystallographic structural factors like bond lengths and bond angles vary as a function of the laser energy, as shown in Table 4.4. Variations in rutile’s lattice characteristics predicted no correlation between increased laser energy and altered bond lengths (Fig. 4.9c). On the contrary, we

Fig. 4.9 Strain formation in TNW by laser-induced surface restructuring/melting of the npAuTNW. a HR-XRD patterns. b Variations in volumetric lattice strain of the TNW anatase and rutile in npAu-TNW as a function of laser energy. c Variations in the Ti–O bond lengths of the anatase and rutile. d Schematics comparing pre- and post-laser distortion of anatase

4.3 Results and Discussion Table 4.3 Changes in the lattice parameters and volumes of rutile and anatase as a function of incoming laser energy

41

TiO2 rutile Lattice volume (Å3 )

Laser energy (μJ)

Lattice constant (Å) a

c

0

4.56

2.99

62.17

15.4

4.57

2.96

61.81

18.2

4.57

2.98

62.24

21.7

4.61

2.94

62.48

25.2

4.57

2.95

61.61

TiO2 anatase Laser energy (μJ)

Lattice constant (Å) a

c

0

3.79

9.85

Lattice volume (Å3 )

141.5

15.4

3.79

9.84

141.3

18.2

3.79

9.78

140.5

21.7

3.80

9.35

135.0

25.2

3.80

9.13

131.8

clearly observed changes in bond lengths in anatase. As the laser energy was raised, the Ti–O bond length of the anatase was substantially lowered, but that in the abplane was not noticeable. Ti–O bond lengths in anatase are progressively shortened. In addition, the O–Ti–O bond angle in the ab-plane was found to drop by 0.43% as laser energy was raised. In contrast, the O–Ti–O bond angle perpendicular to the ab-plane increased. This suggests that the O atom sharing a corner in the TiO6 octahedra was predominantly squeezed along the c-axis (i.e., soft axis) and that the O atoms in the ab-plane were moved away from the Ti4+ center. Figure 4.9d shows a laser-irradiated anatase lattice, reflecting changes in bond lengths and angles. Thus, laser-induced surface restructuring/melting of the npAu led to the development of TNW lattice distortion, most notably in the TiO2 anatase, and this was potentially associated with the production of oxygen vacancies in the anatase by laser pulses. We confirmed oxygen vacancy formation in the npAu-TNW by using Raman spectroscopy (Fig. 4.10a). The bending vibration of the O-Ti–O bond was identified as the source of the 146.5 cm−1 Raman signal, which corresponded to the Eg(1) active anatase mode. In response to laser pulses, the dominant Eg(1) active anatase mode migrated upward in wavenumber (Fig. 4.10b). In addition, we observed the broadening in full width at half maximum of the Eg(1) peak with increasing laser energy. It is widely accepted that variations in peak position and peak broadening in a Raman spectrum are caused by oxygen non-stoichiometry, which also explains the bandgap narrowing by trapping states within the bandgap of TiO2 [43–45]. The oxygen vacancy with a +2 positive charge repels the Ti4+ away from the oxygen vacancy and pulls it toward the adjacent oxygen atom, eventually shortening the

42

4 In Situ Surface Reconstruction-Driven Desorption and Ionization …

Table 4.4 Variations in the crystallographic structural parameters of rutile and anatase as a function of incoming laser energy TiO2 rutile Laser energy (μJ)

Ti–O bond length (Å) Equatorial

Axial

Axial

Axial

0

1.97

1.96

80.0

130.0

15.4

1.97

1.95

80.8

130.4

18.2

1.97

1.95

80.5

130.3

21.7

1.99

1.95

81.8

130.9

25.2

1.97

1.94

81.1

130.5

O-Ti–O bond angle (o )

Ti–O-Ti bond angle (o )

TiO2 anatase O-Ti–O bond angle (o )

Laser energy (μJ)

Ti–O bond length (Å) Equatorial

Axial

Equatorial

Axial

0

1.94

2.05

92.6

77.6

102.4

15.4

1.94

2.04

92.5

77.7

102.3

18.2

1.94

2.03

92.5

77.8

102.2

21.7

1.93

1.95

92.3

78.3

101.7

25.2

1.93

1.89

92.2

78.6

101.4

Ti–O bond length and thus blue-shifting the Eg(1) Raman peak. This agrees with the findings above in Fig. 4.9c, which described the shortening of the anatase’s Ti–O bond length in response to an increase in laser energy. The non-crystallinity also impacted the width of the Eg(1) Raman peak. As shown in Fig. 4.10c, the bandgap energy of the pristine npAu-TNW was calculated to be 3.29 eV, which gradually narrowed down to 2.49 eV with increasing laser energy, corroborating the presence of trap states below the conduction band generated by the oxygen vacancies (Fig. 4.10d). These trap states lowered the probability of carrier recombination, which thereby increased the ionization efficiency. The blue-shift was not seen in the TNW, unlike the npAu-TNW (Fig. 4.10e), and the Eg(1) peak in the Au-TNW was blue-shifted as a consequence of a laser pulse with an energy greater than 18.2 μJ (Fig. 4.10f). These findings reveal that the laser-induced restructuring/melting of the npAu distorted the adjacent TNW anatase lattice, inducing trap levels for electrons and thus lowering the recombination rate of the photo-excited carriers. XPS analysis was used to analyze changes in the surface chemical compositions and valence states of the npAu-TNW in response to UV irradiation (Fig. 4.10g). The Ti 2p XPS spectra exhibited characteristic peaks centered at 459.0 eV (Ti 2p3/2 ) and 464.8 eV (Ti 2p1/2 ), which originated from the Ti4+ valence states in the lattice oxygen. Two further shoulder peaks, at 458.1 eV (Ti 2p3/2 ) and 463.9 eV (Ti 2p1/2 ), were seen when the laser energy was increased, associated with the Ti3+ valence state and oxygen vacancy defect [46, 47]. The O 1 s XPS spectra showed two peaks, one at 530.2 eV (lattice oxygen in the Ti–O bond) and the other at 532.3 eV (non-lattice

4.3 Results and Discussion

43

Fig. 4.10 Increased desorption and ionization due to laser-induced surface restructuring/melting. a Raman spectra of the npAu-TNW as a function of incoming laser energy. b Raman Eg(1) peak shift and broadening in the npAu-TNW. c Diffuse reflectance and Tauc plot for the npAu-TNW. d Trap states formation. Raman spectra of e TNW and f Au-TNW as a function of incoming laser energy

oxygen; surface hydroxyl group (Ti–OH) or oxygen vacancy). The 532.3 eV peak became more prominent with increasing laser energy. As indicated in Fig. 4.10h, the ratios of Ti3+ /Ti4+ (2p3/2 and 2p1/2 ) and of Onon-lattice /Olattice gradually increased with the laser energy, verifying the loss of lattice oxygen atoms from the stoichiometric TiO2 . With increasing laser power, the Ti/O atomic ratio increased in Fig. 4.10i, also supporting oxygen vacancy formation. In summary, increased desorption and ionization with npAu-TNW may be attributed to structural alterations generated by laser light. (1) The desorption efficiency was enhanced by the entropy effect and internal energy transfer. (2) The ionization efficiency was improved by lowering the rate of charge recombination, which was caused by lattice distortions created in the anatase of npAu-TNW.

44

4 In Situ Surface Reconstruction-Driven Desorption and Ionization …

4.3.3 Application of npAu-TNW Nanohybrid for Neurotransmitter LDI-MS Analysis We determined the optimum structural condition of the npAu-TNW for high LDIMS using three neurotransmitters: norepinephrine hydrochloride, serotonin, and dopamine hydrochloride. As a first step, the optimal Au/Ag ratio was selected with the Au thickness held constant at 5 nm, and we chose an Au/Ag ratio of 1:2 as the optimal one, which exhibited the highest mass peak intensities of the three neurochemicals (Fig. 4.11). The npAu-TNW synthesized at an Au/Ag ratio of 2:1 and 1:1 performed poorly in LDI-MS because of their decreased porosity and small surface area. Despite having a greater npAu surface area of the npAu-TNW synthesized at an Au/Ag ratio of 1:3, its performance was lower than that of the npAu-TNW synthesized at an Au/Ag ratio of 1:2 because the bigger npAu hindered the UV laser from reaching the photocatalytic-active TNW. With this optimized Au/Ag ratio of 1:2, we then optimized the Au amount. We prepared the npAu-TNW with Au (Ag) layers of 1 (2), 2 (4), 5 (10), and 10 (20) nm. As indicated in Fig. 4.12, the best LDI-MS results were obtained with npAu-TNW with an Au (Ag) of 5 (10) nm. The resultant npAuTNW had an inadequate Au surface area for efficient LDI-MS performance when the Au (Ag) layer thickness was reduced to 1 (2) and 2 (4) nm. Therefore, thicker Au and Ag films were required to increase the surface area and, thereby, the amount of adsorbed analytes on the npAu-TNW surface. However, the excessive abundance of Au (Ag) limited the UV laser’s access to the TNW surface. Thus, npAu-TNW synthesized with Au (5 nm) and Ag (10 nm) was selected as an LDI-MS substrate. We compared the npAu-TNW’s LDI-MS performance to that of other nanostructures and organic matrices (such as CHCA and DHB), as presented in Fig. 4.13a.

Fig. 4.11 Three neurotransmitters were analyzed using npAu-TNW-based LDI-MS. Deposits of 5 nm Au and varying concentrations of Ag were used to manufacture all of the substrates

4.4 Summary

45

Fig. 4.12 Three neurotransmitters were analyzed using npAu-TNW-based LDI-MS. The npAuTNW was synthesized at different Au amounts at a given Au/Ag ratio of 1:2 (optimized)

The neurotransmitters were protonated in the presence of CHCA and DHB, both of which include a carboxylic acid group. Although the three analytes were ionized by these acidic organic matrices, the S/N ratio was extremely low because of interference from matrix fragments. On the other hand, LDI-MS applications using TNW, Au-TNW, and npAu-TNW nanostructures resulted in much lower levels of background noise. Significantly improved LDI-MS performance was seen with npAuTNW. When comparing npAu-TNW to TNW and Au-TNW, the S/N ratio for norepinephrine (serotonin, dopamine) is 22.8 (–, 42.4)-fold and 10.3 (71.8, 26.7)-fold, respectively. Three neurotransmitters were further added to aCSF to mimic their natural concentrations in the brain. Without any preliminary sample extraction and purification procedures, spike samples were directly tested. As observed in Fig. 4.13b, even in the very salty biological fluid, every neurotransmitter was detected and identified. These findings from mass spectrometry prove the npAu-TNW, which has exceptional photo-induced thermal and electrical characteristics, improved the LDI efficiency of low-molecular-weight analytes, highlighting its potential as a tool for achieving superior LDI-MS results.

4.4 Summary We developed an npAu-TNW nanohybrid to improve LDI efficiencies. The enhancement with our nanohybrid was achievable owing to (1) increased local heating and heat transfer, (2) improved photocatalytic activities, and (3) the occurrence of laserinduced surface restructuring/melting. Particularly, laser-induced structural alterations in the npAu-TNW improved the thermal desorption, as evidenced by the

46

4 In Situ Surface Reconstruction-Driven Desorption and Ionization …

Fig. 4.13 LDI-MS of the three neurotransmitters. a Mass spectra of the three neurotransmitters compared with those of organic matrices including CHCA and DHB and nanostructures containing TNW, Au-TNW, and npAu-TNW. b Neurotransmitter detection with npAu-TNW in the biologically relevant medium of aCSF

DSC thermal analysis results. Moreover, restructuring npAu distorted the adjacent TNW anatase lattice, whereby oxygen vacancies were developed and trap levels were induced within the bandgap, as confirmed by HR-XRD and Raman spectroscopic measurements. With our npAu-TNW nanohybrid, three neurotransmitters, norepinephrine hydrochloride, serotonin, and dopamine, were effectively detected in the aCSF biofluid.

References 1. 2. 3. 4.

Lee S-H, Park JH, Kim SM (2021) J Korean Ceram Soc 58:148–159 Ren S-F, Guo Y-L (2006) J Am Soc Mass Spectrom 17:1023–1027 Liu Q, Shi J, Jiang G (2012) TRAC-Trend. Anal Chem 37:1–11 Sricharoen P, Chanthai S, Lamaiphan N, Sakaew C, Limchoowong N, Nuengmatcha P, Oh W-C (2021) J Korean Ceram Soc 58:201–211 5. Ke Y, Kailasa SK, Wu H-F, Chen Z-Y (2010) Talanta 83:178–184

References

47

6. Kim J-I, Park J-M, Noh J-Y, Hwang S-J, Kang M-J, Pyun J-C (2016) Chemosphere 143:64–70 7. Du J, Zhu Q, Teng F, Wang Y, Lu N (2019) Talanta 192:79–85 8. Kim M-J, Yun TG, Noh J-Y, Park J-M, Kang M-J, Pyun J-C (2019) ACS Appl Mater Interfaces 11:20509–20520 9. Nayak R, Knapp DR (2010) Anal Chem 82:7772–7778 10. Sherrod SD, Diaz AJ, Russell WK, Cremer PS, Russell DH (2008) Anal Chem 80:6796–6799 11. Wang J, Sun J, Wang J, Liu H, Xue J, Nie Z (2017) Chem Commun 53:8114–8117 12. Ng K-M, Chau S-L, Tang H-W, Wei X-G, Lau K-C, Ye F, Ng AM-C (2015) J Phys Chem C 119:23708–23720 13. Song K, Cheng Q (2020) Appl Spectrosc Rev 55:220–242 14. Silina YE, Volmer DA (2013) Analyst 138:7053–7065 15. Nguyen NT, Altomare M, Yoo J, Schmuki P (2015) Adv Mater 27:3208–3215 16. Altomare M, Nguyen NT, Schmuki P (2016) Chem Sci 7:6865–6886 17. Lu X, Balk T, Spolenak R, Arzt E (2007) Thin Solid Films 515:7122–7126 18. Xue Y, Li X, Li H, Zhang W (2014) Nat Commun 5:1–9 19. DeVetter BM, Mukherjee P, Murphy CJ, Bhargava R (2015) Nanoscale 7:8766–8775 20. Jiang C, Elliott JM, Cardin DJ, Tsang SC (2009) Langmuir 25:534–541 21. Kim M-J, Park J-M, Yun TG, Noh J-Y, Kang M-J, Pyun J-C (2020) Chem Commun 56:4420– 4423 22. Liang Q, Chen Y, Fan Y, Hu Y, Wu Y, Zhao Z, Meng Q (2012) Appl Surf Sci 258:2266–2269 23. Lin Z, Wang X, Liu J, Tian Z, Dai L, He B, Han C, Wu Y, Zeng Z, Hu Z (2015) Nanoscale 7:4114–4123 24. Hyun G, Song JT, Ahn C, Ham Y, Cho D, Oh J, Jeon S (2020) Proc Natl Acad Sci 117:5680–5685 25. Yonezawa T, Kawasaki H, Tarui A, Watanabe T, Arakawa R, Shimada T, Mafune F (2009) Anal Sci 25:339–346 26. Jiang K, Smith DA, Pinchuk A (2013) J Phys Chem C 117:27073–27080 27. Almada M, Leal-Martínez BH, Hassan N, Kogan MJ, Burboa MG, Topete A, Valdez MA, Juárez J (2017) Mater Sci Eng C 77:583–593 28. Javdanitehran M, Berg DC, Duemichen E, Ziegmann G (2016) Thermochim Acta 623:72–79 29. Vyazovkin S (2020) Polymers 12:1070 30. Borchardt HJ, Daniels F (1957) J Am Chem Soc 79:41–46 31. Xu C, Ravi Anusuyadevi P, Aymonier C, Luque R, Marre S (2019) Chem Soc Rev 48:3868–3902 32. Northen TR, Woo H-K, Northen MT, Nordström A, Uritboonthail W, Turner KL, Siuzdak G (2007) J Am Soc Mass Spectrom 18:1945–1949 33. Timoshenko VY, Dittrich T, Sieber I, Rappich J, Kamenev BV, Kashkarov PK (2000) Phys Status Solidi 182:325–330 34. Mascaretti L, Naldoni A (2020) J Appl Phys 128:041101 35. Hopkins PE, Serrano JR, Phinney LM, Kearney SP, Grasser TW, Harris CT (2010) J Heat Transf 132 36. Feng Y, Cheng Y, Chang Y, Jian H, Zheng R, Wu X, Xu K, Wang L, Ma X, Li X, Zhang H (2019) Biomaterials 217:119327 37. Panizon E, Ferrando R (2016) Nanoscale 8:15911–15919 38. Yin W-J, Chen S, Yang J-H, Gong X-G, Yan Y, Wei S-H (2010) Appl Phys Lett 96:221901 39. Pang CL (2020) Semicond Sci Technol 35:113001 40. Hary´nski Ł, Grochowska K, Karczewski J, Ryl J, Siuzdak K (2020) ACS Appl Mater Interfaces 12:3225–3235 41. Ma D, Lu Z, Tang Y, Li T, Tang Z, Yang Z (2014) Phys Lett A 378:2570–2575 42. Wawrzyniak J, Karczewski J, Kupracz P, Grochowska K, Zał˛eski K, Pshyk O, Coy E, Bartma´nski M, Szkodo M, Siuzdak K (2020) Appl Surf Sci 508:145143 43. Li H, Guo Y, Robertson J (2015) J Phys Chem C 119:18160–18166

48

4 In Situ Surface Reconstruction-Driven Desorption and Ionization …

44. Silva Junior E, La Porta FA, Liu MS, Andrés J, Varela JA, Longo E (2015) Dalton Trans 44:3159–3175 45. Chen S, Xiao Y, Wang Y, Hu Z, Zhao H, Xie W (2018) Nanomaterials 8:245 46. Jin X, Agyeman DA, Kim S, Kim YH, Kim MG, Kang Y-M, Hwang S-J (2020) Nano Energy 67:104192 47. Mekasuwandumrong O, Chaitaworn S, Panpranot J, Praserthdam P (2019) Catalysts 9:329

Chapter 5

Photothermal Structural Dynamics of Au Nanofurnace-Functionalized ZnO Nanotube (AuNI-ZNT) for In Situ Enhancement in Desorption and Ionization

5.1 Introduction In previous Chap. 4, we demonstrated that laser-induced surface restructuring/ melting of the Au exerted a beneficial effect on the LDI process, wherein the Auadjacent TiO2 lattice was mechanically distorted by restructuring Au. Inspired by the findings, in this chapter, we propose a hybrid nanostructure of Au nanoislandsfunctionalized ZnO nanotubes (AuNI-ZNT) to attain high LDI-MS results. We rationally designed the nanohybrid materials to include piezoelectric ZnO and metallic Au. ZnO has been used for LDI-MS because its bandgap energy of 3.37 eV allows the generation of free carriers in the material through a UV laser pulse (355 or 337 nm), thereby aiding in the analyte ionization [1, 2]. Moreover, ZnO can be very useful as an LDI-MS substrate from an electromechanical perspective. The hexagonal wurtzite ZnO crystal is characterized by the absence of an inversion center. Because of this non-centrosymmetry, ZnO exhibits piezoelectricity along the c-axis with a piezoelectric coefficient d33 of ~12.4 pC N−1 and an electromechanical coupling factor k33 of ~0.48 [3–6]. In the AuNI-ZNT nanohybrid, laser pulses induce structural alterations in AuNI by thermal effect, and this also affects the AuNI-interfaced ZNT mechanically, developing the piezoelectricity [7]. The piezoelectric potential developed in ZNT causes photo-excited electrons and holes to migrate in opposite directions. That is, positive piezoelectric polarization charges attract free electrons, while negative polarization attract holes, spatially separating the carriers, decreasing their likelihood of recombining, and thus increasing the analyte ionization efficiency [8, 9]. This is the first paper we’re aware of linking laser-induced surface reconstruction in LDI-MS to piezoelectricity. The enhanced LDI-MS is confirmed by analyzing the four kinds of fatty acids (behenic acid, oleic acid, palmitic acid, and stearic acid) This chapter is an edited version of the following published journal article: Moon-Ju Kim, Tae Gyeong Yun, Joo-Yoon Noh, Min-Jung Kang, and Jae-Chul Pyun, Small, 2021, 17, 2,103,745. Copyright 2021 Wiley–VCH GmbH. © The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 M.-J. Kim, Laser Desorption Ionization Mass Spectrometry Based on Nanophotonic Structure: From Material Design to Mechanistic Understanding, Springer Theses, https://doi.org/10.1007/978-981-99-6878-7_5

49

50

5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized …

and saccharides (glucose, sorbitol, galactose, and fucose). Non-polar fatty acids and low-acidic/basic saccharides are challenging to detect by MALDI-MS because they intrinsically have low ionization abilities. With our AuNI-ZNT nanohybrid, those analytes are effectively ionized, demonstrating that AuNI-ZNT can be employed as an efficient LDI-active nanostructure and can surmount the primary obstacles faced by traditional MALDI-MS.

5.2 Experimental Methods Synthesis of AuNI-ZNT Nanohybrid We first prepared the ZnO nanorod arrays (ZNR) by depositing a thin ZnO film (90 nm) on a highly p-doped Si wafer and hydrothermally growing the nanorods. After this hydrothermal reaction, the as-prepared ZNR was immersed in an aqueous NaOH solution to obtain the ZnO nanotube arrays (ZNT). The as-obtained ZNT was rinsed thoroughly with water. Eventually, to form Au nanoislands (AuNI) on the ZNT surface, a thin Au film was sputtered on the ZNT, followed by thermal dewetting. Nanostructure Characterization Surface morphology was characterized using a FE-SEM. The crystal structure was characterized using HR-XRD. The diffuse reflectance of the substrates was measured using a UV–vis spectrometer, and the optical absorption was calculated using the reflectance data. The bandgap energy was evaluated by extrapolation of the linear region in the Tauc plot. Micro-Raman spectroscopy was used to acquire the PL spectra, with 325 nm serving as the excitation wavelength. We examined the catalytic efficiencies by methylene blue (MB) degradation. UV and/or ultrasonic irradiation was applied to ZNT and AuNI-ZNT substrates submerged in an aqueous MB solution. We employed a continuous UV lamp in a UV-lithography system (284 nm) to investigate the effects of UV light on photocatalysis. We employed a 40 kHz ultrasonicator to investigate piezocatalysis by ultrasonic irradiation. The substrates were exposed to ultraviolet light and ultrasonic waves to facilitate piezo/photocatalysis. We analyzed the thermal behaviors of the ZNT and AuNI-ZNT substrates with the use of sucrose as a phase transition indicator. The sucrose melting reaction was tracked by heating the sample from 100 to 250 °C at a heating scan rate of 10 °C min−1 . In Situ Laser-Induced Surface Restructuring/Melting The Nitrogen laser was used to irradiate samples on a Microflex LRF 30 mass spectrometer (Bruker Daltonics). The laser intensity was modulated from 0 to 100%, for a total estimated energy output of 28 μJ. We observed the laser-induced surface reconstruction by FE-SEM. The ZnO lattice strain developed by Au structural changes was analyzed using HR-XRD and PL measurements.

5.3 Results and Discussion

51

LDI-MS The LDI-MS analysis was conducted with organic matrices (CHCA, DHB, and 9AA) and nanostructures (ZNT and AuNI-ZNT) using the Microflex LRF 30 mass spectrometer (Bruker Daltonics). Computational Analysis The ab initio approach was used to determine all of the geometries and electronic structures. Using the Gaussian 09 computational tool, we optimized the molecular geometry and calculated the vibrational frequencies using the HF theory and the 3–21 G + basis set. Gaussview 6.0 was used to create maps of the molecules’ electrostatic potentials (ESPs). All the molecules were calculated using the identical parameters to get their average dipole moment per unit of molecular volume. When a species A reacts with hydrogen in the gas phase, the reaction produces a negative enthalpy change (ΔH), which is its proton affinity [10]: H + (g) + A(g) ⇄ AH + (g) ( ) ΔH r xn = −( pr otona f f init yo f A) = H H + + H (A) − H (AH + ) Using the formula for an ideal gas, we were able to determine the proton’s enthalpy, H(H+ ): ) ( 5 H H + = U + P V = RT 2 where U is the internal energy; P and V are the pressure and volume, respectively; R is the universal gas constant; and T is the absolute temperature. It was determined that the H(H + ) in the gas phase, at standard temperature and pressure, had a value of 1.48 kcal mol−1 .

5.3 Results and Discussion 5.3.1 Fabrication and Characterization of AuNI-ZNT Nanohybrid Figure 5.1a illustrates a scheme of the synthesis procedure of the AuNI-ZNT nanohybrid [11–14]. The surface morphology and the side views of the pristine ZNT and the AuNI-ZNT were shown in Fig. 5.1b, where the ZNT was vertically grown with a hexagonal morphology and the Au was uniformly positioned on the ZNT surfaces in the AuNI-ZNT. As indicated in the HR-XRD patterns of Fig. 5.1c, ZNT and AuNI-ZNT had a wurtzite ZnO crystal structure, exhibiting the anisotropic property

52

5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized …

of piezoelectricity [6]. The noble metal Au in the AuNI-ZNT was identified by the (111) peak at 2θ = ≈38°. We also noticed the slight shifts of the XRD peaks for the (100), (002), and (101) ZnO peaks toward higher 2θ angles after Au functionalization, suggesting variations in the bonding characteristics. During the thermal dewetting of the Au film under Ar gas, the annealing at elevated temperature reorganized the Zn and O atoms, and thereby intrinsic lattice defects were formed [15]. There are some studies reporting that O2 -deficient conditions can introduce O vacancies into the metal oxides through the loss of O atoms at the oxide surface at high temperatures. Thus, the O vacancies repel the Zn2+ away from the vacant oxygen sites with a positive charge while attracting the Zn2+ toward the neighboring O2− , eventually shortening the Zn–O bond length. Consequently, the thermal dewetting process might be responsible for the shift of the ZnO peak toward higher angles in the AuNI-ZNT. Bandgap energy estimates for ZNT and AuNI-ZNT are shown in Fig. 5.1d; they are 3.30 and 3.24 eV, respectively, demonstrating that the dewetting procedure had only a little impact on the ZNT’s electrical structure. We confirmed the nanoisland formation by estimating the total Au surface area of the substrate (non-dewetted Au-ZNT and dewetted AuNI-ZNT). We used the 4-aminothiophenol (4-ATP) molecule, which can be chemisorbed on the Au surface [16]. The non-dewetted Au-ZNT was prepared by sputtering an Au film (thickness: 5 nm) on the ZNT without a thermal annealing process. The 4-ATP molecules interacted with the nanostructure substrates of the ZNT (control), non-dewetted Au-ZNT, and dewetted AuNI-ZNT (cross-sectional area: 0.20 cm2 ). After the Au-thiol formation, the amount of 4-ATP not adsorbed

Fig. 5.1 Characterization of the AuNI-ZNT nanohybrid. a The whole process of creating an AuNI-ZNT. b Surface morphology of both natural ZNT and AuNI-ZNT, with a magnified side view. c HR-XRD patterns and d Kubelka–Munk plots of the substrates. e Spectral absorbance measurements of substrates in the UV–vis range showing unbound 4-ATP. f Quantity of adsorbed 4-ATP molecules and substrate Au surface area covered by 4-ATP

5.3 Results and Discussion

53

onto the substrate was calculated by measuring the absorbance at λmax = 258 nm. The absorption spectra of the non-adsorbed 4-ATP are shown in Fig. 5.1e. The absorbance was the lowest for the ZNT surface, whereas the 4-ATP was immobilized onto the non-dewetted Au-ZNT and dewetted AuNI-ZNT. A relatively larger amount of 4-ATP molecules covered the surface of non-dewetted Au-ZNT than the dewetted AuNI-ZNT, demonstrating the reduction in total Au surface area by the thermal dewetting of the Au thin film and thus the nanoisland formation. By using the Beer-Lambert law, we calculated the number of substrate-adsorbed 4-ATP: 0.310 nmol (ZNT; non-specific interaction), 6.50 nmol (dewetted AuNI-ZNT), and 8.26 nmol (non-dewetted Au-ZNT) (Fig. 5.1f). Assuming that the 4-ATP has a molecular footprint of ≈0.20 nm2 per molecule [17, 18], nanostructured Au surface area was estimated to be 7.83 cm2 (dewetted AuNI-ZNT) and 9.95 cm2 (non-dewetted Au-ZNT), which were notably higher values than the substrate cross-sectional area (0.20 cm2 ). Although the non-dewetted Au-ZNT had an enlarged surface area of Au compared to the dewetted AuNI-ZNT, the Au thin film on the ZNT inevitably screened the photocatalytic-active ZNT surface, rendering the ZNT structure useless [19]. In contrast, the AuNI-ZNT nanohybrid provided a free ZNT surface as well as an enlarged Au surface area.

5.3.2 Strategies for Improving Desorption and Ionization Efficiencies 5.3.2.1

Piezoelectricity-Enhanced Photocatalytic Process in AuNI-ZNT Nanohybrid

We evaluated the ionization capability of the ZnO nanostructures in terms of their piezocatalytic and photocatalytic activities. Different irradiation settings, including ultraviolet (UV) irradiation, ultrasonic (US) irradiation, and simultaneous UV and US irradiation, were studied to determine their effects on the (1) photo-, (2) piezo-, and (3) piezo/photocatalytic processes. Figure 5.2a depicts the mechanism of the methylene blue (MB) degradation by the ZnO catalysts, where UV and ultrasonic irradiation modulate the electronic band structure of the ZnO (i.e., tilting the band edges). Considering the energy alignment between the ZnO bands and the redox energies of water and oxygen, water oxidation by holes and oxygen reduction by electrons are feasible, producing hydroxyl radicals (•OH). Accordingly, bleaching of the MB color occurs under the attack of •OH. Figure 5.2b shows the photocatalytic efficiency of the AuNI-ZNT nanohybrid by comparing it to that of the pristine ZNT under UV irradiation. In the absence of nanostructures, MB was stable under UV light, revealing that MB degradation was the result of photocatalysis. The AuNI-ZNT photocatalyst dramatically accelerated MB degradation in comparison to the ZNT, which was attributable to the Schottky barrier at the AuNI/ZNT heterojunction. As

54

5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized …

expected, the non-dewetted AuNI-ZNT was inefficient as a photocatalyst due to the screening effect (Fig. 5.3) [20]. We further examined the piezocatalytic performance of the ZnO-based catalysts under ultrasonic vibration (Fig. 5.2c). The extent of MB degradation by piezocatalysis was obviously lower than that by photocatalysis. Although piezoelectric polarization charges were produced in the ZNT and AuNI-ZNT upon mechanical agitation, these polarization charges were intrinsically confined inside the ZNT. Thus, polarization charges were ineffective in MB degradation [21]. Therefore, piezocatalytic MB degradation was hardly feasible in contrast to the photocatalytic process. Interestingly, MB deterioration was accelerated when piezoelectric polarization was coupled with photoexcitation [22, 23] (Fig. 5.2d). Within the piezopotential well by ultrasonic irradiation, the UV-excited electrons freely moved toward one ZNT surface with positive polarization charges, whereas holes were attracted to the other

Fig. 5.2 Degradation of methylene blue as a measure of ZNT and AuNI-ZNT’s catalytic activity. a Schematic representation of the fundamental piezo-photocatalytic mechanism in ZnO. b Reaction catalyzed by ultraviolet light (photocatalysis). c Piezocatalytic reaction activated by ultrasound. d Piezo/photocatalytic activity when exposed to ultrasonic and ultraviolet light

5.3 Results and Discussion

55

Fig. 5.3 The photocatalytic performance of non-dewetted Au-ZNT

with negative polarization, tilting the ZNT band edges. Moreover, the AuNI-ZNT nanohybrid presented a higher piezo/photocatalytic efficiency than the pristine ZNT. As depicted in the band diagram of Fig. 5.2d, the ultrasound-induced piezopotential changed the band edges at the AuNI/ZNT interface, modulating the Schottky barrier height. Positive piezoelectric polarization bent the band edges downward and reduced the height of the Schottky barrier, while negative polarization uplifted the band edges and raised the barrier height. Accordingly, an asymmetric band structure was obtained, which promoted electron migration from the ZNT to the AuNI side, whereas holes preferred to be accumulated at the opposite ZNT side. Thus, efficient carrier splitting considerably facilitated the MB degradation process. We also compared the catalytic efficiencies of substrates by analyzing the reaction kinetics, assuming that the MB degradation reaction followed pseudo-first-order kinetics (Fig. 5.4). AuNI-ZNT nanohybrid outperformed others in photo-, piezo-, and piezo/photocatalytic activities (Fig. 5.5). When comparing photo- and piezocatalysis, the combination of the two proved to be the most effective in destroying MB molecules. These results reveal that integrating piezocatalysis with photocatalysis in the AuNI-ZNT nanohybrid can markedly enhance the ionization by promoting carrier separation at the surface as well as in the bulk.

Fig. 5.4 Methylene blue degradation kinetics using ZNT and AuNI-ZNT. a Photocatalysts, b Piezocatalysts, and c Piezo-photocatalysts

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5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized …

Fig. 5.5 Methylene blue degradation kinetics with ZNT and AuNI-ZNT under various irradiation settings

5.3.2.2

Efficient Heat Transfer in AuNI-ZNT Nanohybrid

We assessed the desorption capability by estimating the heat transfer efficiency of nanostructures. We measured the heat flow vs. temperature with the use of a sucrose analyte. The DSC thermograms of sucrose, ZNT, and AuNI-ZNT mixtures, as well as pure sucrose, are shown in Fig. 5.6a. Sucrose melted at a peak temperature of 192.7 °C with a corresponding enthalpy of 304.4 mJ (Fig. 5.6b). In terms of melting temperature and enthalpy, the DSC findings for sucrose combined with ZNT were identical to those for pure sucrose. In contrast, sucrose combined with AuNI-ZNT had a much lower melting temperature and enthalpy, suggesting that metallic Au induced sucrose to melt easily. This was attributable to a higher thermal conductivity of Au than that of ZnO. Therefore, the AuNI-ZNT nanohybrid allowed for efficient heat transmission to analytes. We further compared the heat transfer efficiencies of the ZNT and AuNI-ZNT by examining the reaction kinetics of sucrose melting using the above DSC thermograms. Figure 5.6c presents the Arrhenius plot, where the activation energy (Ea ) was obtained from the straight-line slope. Compared to pristine ZNT, the AuNI-ZNT nanohybrid reduced the Ea of the sucrose melting reaction, further supporting efficient heat transfer. In addition, T50% was significantly lower with the AuNI-ZNT than with the ZNT (Fig. 5.6d). The thermal parameter values are available in Table 5.1. These DSC thermal results demonstrate that the outstanding thermal conduction of the AuNI facilitated heat transfer to the sucrose analyte, leading to the kinetically favored melting.

5.3.2.3

In Situ Photothermal Structural Changes in AuNI-ZNT Nanohybrid

A short laser pulse can trigger the surface restructuring/melting of the metal nanostructures (Fig. 5.7a). We estimated the laser-driven AuNI surface temperature to confirm its surface restructuring/melting. We theoretically calculated the laserinduced surface temperature rise of the AuNI with an average diameter of 15.2 nm.

5.3 Results and Discussion

57

Fig. 5.6 DSC thermal study of sucrose combined with ZNT and AuNI-ZNT nanostructures. a DSC thermograms. b Variations in sucrose melting start temperature and enthalpy. c Calculation of the activation energy for the sucrose melting process using Arrhenius plots. d Extent of the reaction

Table 5.1 Thermal parameters of the sucrose melting determined by DSC Sucrose

ZNT with sucrose

AuNI-ZNT with sucrose

Tonset (°C)

189.2

189.5

185.7

Fusion enthalpy (ΔH, mJ)

304.4

290.9

175.4

Activation energy (Ea , kJ/mol)

1806±27.61

1976±29.59

1532±68.95

T50% (°C)

188.9

189.0

185.1

The temperature rise on the metal surface in response to an incoming laser is dependent on three factors: (1) the photoabsorptivity of the metal, (2) the incident laser wavelength/energy, and (3) the specific heat capacity of the metal. Using the Rayleigh approximation, the photoabsorption efficiency (Q) of a metal nanostructure may be determined for a given laser wavelength. The Q value is also related to the radius of the metal nanostructure (R), wavelength of light (λ), and dielectric permittivity of the metal (ε = ε’ + iε”, where ε’ and ε” are the real and imaginary parts of ε, respectively), as shown in Eq. 5.1:

Q=

12ε'' R λ (ε' + 2)2 + (ε'' )2

(5.1)

58

5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized …

Fig. 5.7 Surface reorganization of the Au nanofurnace caused by laser. a Schematic depiction of the laser-induced surface restructuring/melting. b Surface temperature of Au after being heated by laser. c Laser-induced structural modifications in AuNI-ZNT. d Relative desorption performance of ZNT and AuNI-ZNT

Energy density absorbed into the metal (E, in J cm−3 ) is given in terms of laser fluence (Φ, in J cm−2 ) and maximum surface temperature of the metal under the assumption that all absorbed light is turned into heat and there is little to no heat loss (Eq. 5.2). E=

3Q Φ = cρT 4R

(5.2)

where c is the specific heat capacity and ρ is the density of the metal. As the laser fluence increased from 72.4, 89.8, to 101.8 mJ cm−2 , the estimated increase in Au temperature was 1362, 1689, and 1915 K, respectively, far over the bulk melting point of gold (Fig. 5.7b). Laser irradiation indeed resulted in the Au melting, also accompanying the Au morphological changes. As observed in Fig. 5.7c, higher laser fluence caused a more dramatic alteration of the AuNI shape by increasing the laser-induced heating temperature. Surface restructuring/melting occurred significantly more extensively in the AuNI than in the ZNT. ZNT virtually preserved its original shape, whereas morphological changes in the Au component of the AuNIZNT nanohybrid were clearly evident. Because Au has a lower specific heat than ZnO, a higher surface temperature was easily formed on the AuNI than on the ZNT. We also proved a correlation between surface restructuring/melting and the iondesorption efficiency of AuNI-ZNT by making use of methylthioninium chloride. As shown in Fig. 5.7d, the mass peak intensity of the methylthioninium cation was far higher with AuNI-ZNT compared with ZNT. These MB desorption results validate

5.3 Results and Discussion

59

that the Au functionalized on the ZNT surface acted as a photo-induced “nanofurnace” and played a key role in the ion-desorption through laser-induced heating and the resultant structural changes. Furthermore, we structurally investigated the laser-induced distortion of the AuNI-ZNT lattice based on the HR-XRD data (Fig. 5.8a). Structural parameters were determined using Bragg’s law and are summarized in Table 5.2. As shown in Fig. 5.8b, the wurtzite ZnO in the AuNI-ZNT was lengthened after being exposed to a laser with a fluence of 101.8 mJ cm−2 along the a- and c-axes, respectively, by 0.12% and 0.36%. Prominent ZNT expansion along the c-axis was ascribed to the anisotropic deformability of ZnO. Increases in c and the c/a ratio as a function of laser fluence are indicative of a relative displacement of the Zn2+ ion from the ZnO4 unit’s geometric center along the c-axis [24]. We also estimated the degree of the ZnO4 unit distortion with the applied laser by calculating the Zn–O bond length and the O–Zn–O bond angles. Variations in bond lengths and angles are summarized in Table 5.3. Changes in the ZnO lattice constants of the AuNI-ZNT predicted an increase in the closest Zn–O bond along the c-axis when laser fluence was applied (Fig. 5.8c). The O–Zn–O bond angle along the c-axis also increased, whereas the inplane O–Zn–O bond angle decreased with increasing laser fluence, demonstrating the ZnO anisotropic deformation. The ZnO lattice in the AuNI-ZNT after 101.8 cm−2 -laser irradiation is illustrated in Fig. 5.8d using the VESTA software.

Fig. 5.8 The laser-induced restructuring/melting of the Au nanofurnace and its effect on the strain evolution. a HR-XRD patterns. Variations in the b lattice constants and c/a ratio and the c Zn– O bond length and O–Zn–O bond angles for the wurtzite ZnO in the AuNI-ZNT with increasing laser fluence. d Differences in wurtzite ZnO lattice distortions before and after laser irradiation

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5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized …

Table 5.2 Variation in the lattice constants of (a) AuNI and (b) ZNT in the AuNI-ZNT as a function of laser fluence a Laser fluence (mJ cm−2 )

AuNI lattice constant (Å) AuNI lattice volume (Å3 )

0

4.073

a 67.57

72.4

4.075

67.67

89.8

4.079

67.87

101.8

4.080

67.92

b ZNT lattice constant (Å)

Laser fluence (mJ cm−2 )

a

ZNT lattice volume (Å3 )

c

0

3.249

5.192

47.46

15.4

3.250

5.202

47.58

21.0

3.253

5.208

47.72

26.6

3.253

5.214

47.78

Table 5.3 Estimated variations in ZNT structural characteristics as a function of laser fluence Parameters

Laser fluence (mJ cm−2 ) 0

72.4

89.8

l (Å)

1.976

1.978

1.979

l’ (Å)

3.216

3.225

3.229

108.8 1.980 3.234

α (deg.)

108.27

108.39

108.40

108.43

β (deg.)

110.64

110.53

110.52

110.49

Laser-induced out-of-plane expansion displaced the off-centered Zn ion and thereby increased the ionic displacement relative to the c value after the laser exposure. Without the Au nanofurnace (i.e., pristine ZNT), the laser-induced crystal distortion was not obvious (Fig. 5.9). These structural analysis results indicate that the laser-induced restructuring/melting of the Au nanofurnace distorted the adjacent ZNT and generated a piezoelectric field. We used PL spectroscopy to look at how the configurations of the energy bands changed in response to the laser pulses. Figure 5.10a shows the PL emission peaks of the AuNI-ZNT, presenting the two bands: relatively sharp UV (near-band-edge emission) and broad visible bands (defect states) [25]. UV peak intensity gradually reduced, and its position shifted toward a lower photon energy with increasing laser fluence (Fig. 5.10b). The UV peak intensity of the AuNI-ZNT decreased at a laser fluence of 101.8 mJ cm−2 , suggesting the inhibition of the photo-excited carrier recombination (Fig. 5.10c). The Schottky barrier at the Au/ZnO interface significantly decreased the intensity of the PL emission peak compared with that

5.3 Results and Discussion

61

Fig. 5.9 Structural analysis of the pristine ZNT based on the XRD data. a HR-XRD patterns. b Variations in the lattice parameter a, c, and c/a values with respect to the incoming laser fluence. c Variations in the Zn–O bond length along the c-axis and the O–Zn–O bond angles along the c-axis and in the ab-plane with respect to the laser fluence

of pristine ZnO [26] (Fig. 5.11). Since the wurtzite ZnO in the AuNI-ZNT was progressively subjected to tensile strain as a result of the laser-induced Au surface reconstruction, the Schottky barrier height decreased as the laser fluence increased. The migration of photo-excited electrons into the Au area was aided by a decrease in the Schottky barrier height. Thus, the decrease in the UV peak intensity with the applied laser fluence might arise from the modulated Schottky barrier height and thereby improved carrier separation. Moreover, the red-shift in the UV peak position was observable. Bandgap narrowing in piezoelectric ZnO has reportedly been achieved by c-axis tensile strain by the downward (upward) shift of the conduction band (valence band) [27, 28]. This red-shift in the UV peak location may have been caused by tensile strain along the c-axis as a consequence of laser-induced AuNI reconstruction, which caused an asymmetric band bending at the AuNI/ZNT interface. Some people may be concerned about the effect of laser exposure on PL measurements in the UV spectrum since laser irradiation may cause defect states in ZnO. However, the near-band-edge emission in the UV region is hardly affected by laserinduced defects because they typically exhibit deep donor and/or acceptor features [28]. In addition, the UV emission peak in the PL spectra of laser-irradiated ZNT hardly shifted. Therefore, we excluded the laser exposure effect on the defect state formation, and the tensile strain and corresponding piezoelectricity originated from the laser exposure. To further verify a correlation between the laser-induced surface reconstruction and the resultant piezoelectrity-induced changes in the energy bands, we analyzed the defect-related PL emission spectra of the AuNI-ZNT. Given the LDIMS measurement performed in high vacuum circumstances, e.g., an O2 -deficient condition, it is highly anticipated that most defects produced by laser pulses are oxygen vacancies. ZnO’s oxygen vacancies may be in one of three possible charge states: neutral VO X , singly positively ionized VO + , or doubly ionized VO ++ [29]. Thus, we investigated the defect-related PL emissions in terms of these three trap centers. As seen in Fig. 5.10d, we split the broad visible band into three peaks centered at 1.91, 2.19, and 2.48 eV. The 1.91 eV PL peak is reportedly related to native defects such as interstitial Zn (Zni ) and interstitial O (Oi ), while the latter two peaks originate from

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5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized …

Fig. 5.10 Changes in the energy band structures caused by the laser. a PL spectra of the AuNIZNT. b UV emission PL band. c Changes in the location and strength of the UV emission peak as a function of the incoming laser fluence. d Deconvoluted peaks in the PL band of the visible emission spectrum. e Variations in the defect ratios. f Schematics depicting the piezotronic effect caused by laser light in AuNI-ZNT nanohybrids

oxygen vacancies with different charge states. Because of the different Fermi levels between ZnO and Au, a depletion region at the AuNI/ZNT interface was present wherein the Fermi level of the ZNT reportedly passed below the VO + /VO ++ level, and thus most oxygen vacancies are in the VO ++ state. In the bulk region, however, most defects are in the VO + state [29, 30]. Considering the band bending-associated defect chemistry, the two peaks at 2.19 and 2.48 eV were assigned to have been attributed to the recombination of VO + and VO ++ centers, respectively. As a result of the electrons being swept through the depletion zone, most VO + was converted to a VO ++ state by trapping the holes at the valence band, and radiative recombination (i.e., PL emission) occurred between this center and the electrons at the conduction band, emitting a photon energy of 2.19 eV [31]. In the bulk region, the VO + state easily trapped the electrons in the conduction band and yielded the neutral VO x center. The

5.3 Results and Discussion

63

Fig. 5.11 PL emission spectra of pristine ZNT and AuNI-ZNT nanohybrid

radiative recombination of this neutral center with holes at the valence band emitted an energy of 2.48 eV. With increasing laser fluence, the relative defect-related PL band ratios gradually varied (Fig. 5.10e). Radiative recombination in the bulk area rose from 13 to 58%, while it reduced from 70 to 30% in the depletion zone, and the PL emission associated with the Zni -Oi transition remained almost constant. These PL results are in agreement with the interpretations of Fig. 5.10b, explaining that laser pulses reduced the depletion zone in comparison to the bulk one. As summarized in Fig. 5.10f, the laser-induced piezotronic effect in AuNI-ZNT nanohybrid was corroborated by the red-shift in the UV emission peak, the decrease in peak intensity, and the relative fluctuations in the visible PL emission fraction of the depletion and bulk areas.

5.3.3 LDI-MS Performance Enhancement Based on AuNI-ZNT Nanohybrid As an example of the improved LDI-MS with AuNI-ZNT nanohybrid, we analyzed tiny molecules of fatty acid (low polarity) and saccharide (low acidity/basicity) to prove our point. Fatty acids have a lengthy aliphatic chain and a small hydrophilic head. The electric dipole moments (μ) of the fatty acids were calculated in the gas phase, which allowed for an assessment of the asymmetry of the molecular charge distribution. We compared the μ value of the fatty acid to that of the four amino acids. Note that MALDI-MS allowed for the simple detection of these four amino acids; nevertheless, the matrix-related background noises were very disruptive to the MS results (Fig. 5.12). As can be seen in Fig. 5.13a, the minimal variation in electronegativity between C and H atoms results in a uniform distribution of electrons throughout the lengthy non-polar chain in all four fatty acids.

64

5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized …

Fig. 5.12 MALDI-MS analysis of four amino acids with a CHCA, b DHB, and c 9-AA

The estimated values of fatty acids were far lower than those of amino acids, as predicted given the lengthy hydrophobic chains (Fig. 5.14). Despite the basicity of the 9-AA, it hardly deprotonated the four fatty acids. The nanostructures of ZNT and AuNI-ZNT, on the other hand, clearly ionized the four fatty acids, and the AuNIZNT further resulted in notably high LDI-MS results (Fig. 5.13b). Four saccharides, including glucose, sorbitol, galactose, and fucose, were also tested. Due to their neutral nature (i.e., low acidity/basicity), saccharides present difficulties during ionization in LDI-MS. We measured the capacity of saccharides to be protonated by their gas-phase proton affinity (PA), which is the negative of the enthalpy change associated with the protonation process [10]. Here, glucose was used as a representative of the saccharides (Fig. 5.15a). Compared to the four amino acids, the average PA value of glucose in its standard condition was lower (Fig. 5.16a). Further, the PA of the deprotonated glucose was also computed to evaluate its stability (i.e., the reverse reaction is increasingly favorable as the PA value of the deprotonated product increases) (Fig. 5.15b). A greater PA for the deprotonated glucose than the deprotonated amino acids suggests that glucose deprotonation is less preferred than the deprotonation of amino acids (Fig. 5.16b). As can be seen in Fig. 5.16c, the ZNT and AuNI-ZNT indeed ionized the four saccharides, but the typical organic matrices

5.3 Results and Discussion

65

Fig. 5.13 LDI-MS of four types of low-polarity fatty acids. a The ESP maps of the four fatty acids. b Mass spectra of fatty acids with organic matrix (9-AA) and inorganic nanostructures (ZNT, AuNI-ZNT) compared

of CHCA, DHB, and 9-AA had no impact. In addition, the AuNI-ZNT nanohybrid allowed us to achieve significantly enhanced LDI-MS intensities. We further validated the practical usefulness of the AuNI-ZNT by detecting saccharides in the biological environment of human serum without any prior sample extraction or purification. We discovered a mass peak in the low mass range at m/z 202.8, which we believe was caused by glucose as a sodium adduct given the greatest concentration of glucose among other saccharides in normal human (Fig. 5.16d). With the use of tandem time-of-flight (TOF/TOF) MS [32], we were able to verify that these peaks were the result of glucose (Fig. 5.16e). Therefore, even in a complex biofluid, we discovered glucose with inherent poor ionization efficiency using the AuNI-ZNT nanohybrid, underscoring its practical usefulness in LDI-MS.

Fig. 5.14 The electron distribution in four different amino acids, as seen by the electrostatic potential (ESP) map, along with the overall dipole moment per molecule volume

66

5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized …

Fig. 5.15 Gas-phase proton affinity of a the four types of amino acids and b the deprotonated amino acids. The amino acids’ proton affinities were determined by protonating the amino group and deprotonating the carboxyl group of the backbone, respectively

5.4 Summary Inspired by the Au surface restructuring/melting by laser pulses, we designed the AuNI-ZNT nanohybrid to make use of the piezoelectricity of the ZNT interfaced with AuNI as an LDI-improving strategy. We achieved superior LDI-MS results with the AuNI-ZNT attributable to (1) piezoelectricity-assisted photocatalytic ionization, (2) improved heat transfer for thermal desorption, and (3) laser-induced surface reconstruction and improved LDI. Particularly, the AuNI played a significant role as a "nanofurnace” upon laser pulses exhibiting photodynamic behaviors, which generated the ZNT lattice distortion and thereby developed the piezoelectricity. During this, the energy band structure of the ZNT was modulated to spatially separate the photoexcited charge carriers, thus reducing their recombination and improving the efficiency of analyte ionization. Ultimately, we verified that our AuNI-ZNT nanohybrid was highly effective in ionizing tiny biomolecules such as fatty acids and saccharides, which posed a significant difficulty for conventional MALDI-MS.

5.4 Summary

67

Fig. 5.16 LDI-MS of the four low-acidity/-basicity monosaccharides. a Gas-phase proton affinity of the glucose. b Gas-phase proton affinity of the deprotonated glucose. c Comparison of the mass spectra of the four types of monosaccharides with organic matrix (CHCA, DHB, and 9-AA) and nanostructures (ZNT and AuNI-ZNT). d Glucose detection in human serum. (e) Tandem mass spectrum of the mass peak at m/z = 202.8

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5 Photothermal Structural Dynamics of Au Nanofurnace-Functionalized …

References 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30. 31. 32.

Gedda G, Wu H-F (2019) Sens actuators B: Chem 288:667–677 Du J, Zhu Q, Teng F, Wang Y, Lu N (2019) Talanta 192:79–85 Wang ZL, Song J (2006) Science 312:242–246 Kong XY, Wang ZL (2003) Nano lett 3:1625–1631 Wang M, Wang B, Huang F, Lin Z (2019) Angew Chem Int Ed 58:7526–7536 Jung YS, Choi HJ, Park JW, Cho YS (2021) Nano Energy 82:105690 Hanley L, Wickramasinghe R, Yung YP (2019) Ann Clin Lab Sci 12:225–245 Pan L, Sun S, Chen Y, Wang P, Wang J, Zhang X, Zou J-J, Wang ZL (2020) Adv Energy Mater 10:2000214 Wu J, Qin N, Bao D (2018) Nano Energy 45:44–51 East ALL, Smith BJ, Radom L (1997) J Am Chem Soc 119:9014–9020 Choi HJ, Jung YS, Han J, Cho YS (2020) Nano Energy 72:104735 Yang J, Lin Y, Meng Y, Liu Y (2012) Ceram Int 38:4555–4559 Thompson CV (2012) Annu Rev Mater Res 42:399–434 Kang M, Ahn M-S, Lee Y, Jeong K-H (2017) ACS Appl Mater Interfaces 9:37154–37159 Mamat MH, Khalin MIC, Mohammad NNHN, Khusaimi Z, Sin NDM, Shariffudin SS, Zahidi, MM, Mahmood MR (2012) J. Nanomater. 2012, Article 8 Xue Y, Li X, Li H, Zhang W (2014) Nat Commun 5:1–9 DeVetter BM, Mukherjee P, Murphy CJ, Bhargava R (2015) Nanoscale 7:8766–8775 Jiang C, Elliott JM, Cardin DJ, Tsang SC (2009) Langmuir 25:534–541 Ye Y, Wang K, Huang X, Lei R, Zhao Y, Liu P (2019) Catal. Sci Technol 9:3771–3778 Nguyen NT, Altomare M, Yoo J, Schmuki P (2015) Adv Mater 27:3208–3215 Feng Y, Ling L, Wang Y, Xu Z, Cao F, Li H, Bian Z (2017) Nano Energy 40:481–486 Wu M-H, Lee J-T, Chung YJ, Srinivaas M, Wu J-M (2017) Nano Energy 40:369–375 Liu L, Huang H, Chen Z, Yu H, Wang K, Huang J, Yu H, Zhang Y (2021) Angew Chem Int Ed 60:18303–18308 Mostafa NY, Heiba ZK, Ibrahim MM (2015) J Mol Struct 1079:480–485 Xu S, Guo W, Du S, Loy MMT, Wang N (2012) Nano Lett 12:5802–5807 Xiang D, Liu Z, Wu M, Liu H, Zhang X, Wang Z, Wang ZL, Li L (2020) Small 16:1907603 Choi HJ, Jang W, Mohanty BC, Jung YS, Soon A, Cho YS (2018) J Phys Chem Lett 9:5934– 5939 Han X, Kou L, Lang X, Xia J, Wang N, Qin R, Lu J, Xu J, Liao Z, Zhang X, Shan X, Song X, Gao J, Guo W, Yu D (2009) Adv Mater 21:4937–4941 Ye JD, Gu SL, Qin F, Zhu SM, Liu SM, Zhou X, Liu W, Hu LQ, Zhang R, Shi Y, Zheng YD (2005) Appl Phys A 81:759–762 Wojcik PM, Bastatas LD, Rajabi N, Bakharev PV, McIlroy DN (2020) Nanotechnol 32:035202 van Dijken A, Meulenkamp EA, Vanmaekelbergh D, Meijerink A (2000) J Lumin 87:454–456 Mandal SM (2011) Metallomics 3:1074–1078

Chapter 6

In Situ Evolution of Atomic Defect and Piezoelectricity in Graphitic Carbon Nitride Nanosheet (G-C3 N4 NS) by Short Laser Pulse for the Ionization in Mass Spectrometry

6.1 Introduction The purposes of increasing the detection sensitivity and repeatability of small biomolecules in LDI-MS have prompted extensive research on nanophotonic structures for high LDI-MS performance. However, little research has been conducted to investigate the effect of a laser-driven shock wave on the ionization in LDI-MS. In this chapter, for the first time, we comprehensively clarify the impact of the laser shock wave on the ionization using two-dimensional graphitic carbon nitride nanosheets (g-C3 N4 NS). We use the LDI-active g-C3 N4 NS, which has recently garnered a lot of interest as a possible option that may supplement carbon nanostructures like carbon nanotubes and graphene [1–4]. We focus on the effect of the laser shock wave on aspects of atomic defects and piezoelectricity in g-C3 N4 NS using laser pulses with nanosecond widths. By making estimates of the fluctuations in the crystallographic parameters, we structurally investigate the cause of the LDI improvement in terms of the laser shock pressure-driven anisotropic g-C3 N4 NS lattice distortions. Atomic defects, including N and C vacancies, that may operate as carrier trap sites and suppress carrier recombination are generated by the LDI laser shock pressure that is strong enough to break the bonds in the g-C3 N4 NS. Moreover, under the pressure of a laser shock, piezoelectricity is instantly developed in the g-C3 N4 NS, which reportedly has a piezoelectric coefficient e11 of 0.732 C m−2 because of the non-centrosymmetric triangular nanoholes aligned in the 2D NS layer [5, 6]. The in situ-developed piezoelectricity thus modulates the carrier motion and separates them, also suppressing their recombination [7, 8]. The role of atomic defect and piezoelectricity in efficient ionization has been confirmed by density functional theory (DFT) calculations using Mulliken This chapter is an edited version of the following published journal article: Moon-Ju Kim, Joo-Yoon Noh, Tae Gyeong Yun, Min-Jung Kang, Dong Hee Son, and Jae-Chul Pyun, ACS Nano 2022, 16, 18284–18297. Copyright 2022 American Chemical Society. © The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2023 M.-J. Kim, Laser Desorption Ionization Mass Spectrometry Based on Nanophotonic Structure: From Material Design to Mechanistic Understanding, Springer Theses, https://doi.org/10.1007/978-981-99-6878-7_6

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charges and dipole moments, together with experimental explorations. The findings of this study contribute to a greater mechanistic comprehension of the ionization procedures, which is essential for revealing the full power of LDI-MS’s laser shock waves.

6.2 Experimental Methods Preparation of the Few-Layered G-C 3 N 4 NS We first prepared bulk g-C3 N4 by thermal condensation of melamine. 7 g of melamine were subjected to a 520 °C, 2-h calcination in air at a heating rate of 5 °C min−1 . After being ground into a powder, the resultant yellow product was acid treated with concentrated H2 SO4 for 30 min while being stirred to create the NS dispersion. The sample was thoroughly washed with water, and the exfoliation process then began with a 5-h sonication session. The un-exfoliated g-C3 N4 particles were separated from the NS suspension by centrifuging the suspension at 7500 rpm for 10 min in water. Hummer’s Method We prepared the oxidized g-C3 N4 NS from the bulk using a modified Hummer’s method [9]. Bulk g-C3 N4 powder (1 g) was immersed in a concentrated H2 SO4 solution (30 mL) and then heated at 60 °C for 1 h under stirring, after which it was immediately cooled in an ice bath. Subsequently, the KMnO4 (1.2 g) was slowly added to the sample over 20 min. After that, we heated the mixture to 30 °C and stirred it constantly for 30 min. The water (200 mL) was progressively poured in over the course of 10 min, and the sample was placed in an ice bath. To avoid damaging the g-C3 N4 crystal structure, it was important to keep the sample of water-poured mixture below 30 °C during the first 5 min of preparation. The aforesaid solution was then treated with an H2 O2 (5%) aqueous solution. The g-C3 N4 NS product was filtered through clean water many times before being dried and used again. Sample Characterization FE-SEM, TEM, and AFM were all used to examine the g-C3 N4 samples and determine their shape and thickness. HR-XRD was used to determine the crystal structures utilizing Cu-Kα radiation in glancing-incidence mode. Micro-Raman spectroscopy with a 785-nm excitation wavelength was used to study surface states. Diffuse reflectance was measured using a UV–vis spectrometer, and optical absorption was calculated using the Kubelka–Munk function. Laser Pulse Irradiation Laser pulse irradiation during LDI-MS was responsible for the in situ development of the atomic defect and piezoelectricity in the g-C3 N4 NS. We performed a laser irradiation using an Autoflex Max mass spectrometer (Bruker Daltonics, Germany)

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71

equipped with a smartbeam II laser. The “medium” focus setting was used, which is known to produce a laser beam diameter of around 50 μm. The minimum and maximum laser energies were 12 and 24 μJ, respectively. Piezoelectric Energy Harvester We fabricated a g-C3 N4 NS-based flexible piezoelectric energy harvester. The polyethylene terephthalate (PET) substrates were cleaned with acetone and ethanol for 10 min using an ultrasonic bath and thoroughly dried at 70 °C for 15 min. A solution of poly(3,4-ethylenedioxythiophene): poly(styrenesulfonate) (PEDOT:PSS) was spin-coated on it at 3500 rpm for 30 s. The laser-irradiated g-C3 N4 NS was then deposited on the PEDOT:PSS-coated PET at 1000 rpm for 60 s and then dried at 70 °C for 15 min. At 5000 rpm for 90 s, insulating poly(methylmethacrylate) (PMMA) was spin-coated onto the g-C3 N4 to avoid electrical shorts. The PEDOT:PSS was deposited on the PMMA/g-C3 N4 /PEDOT:PSS/PET as a top electrode by spin coating, followed by annealing at 70 °C for 15 min. Cu wires were connected to the two PEDOT:PSS sides. The g-C3 N4 NS-based piezoelectric energy harvester was subjected to repeated bending-releasing testing at a bending frequency of 1 Hz utilizing a bending machine in which the circumstances, such as the bending frequency and radius of curvature, were under precise control. The output current was measured using a potentiostat (CompactStat, Ivium Technologies, Netherlands). LDI-MS The organic matrix solution (9-AA, DMAN, and THAP) was prepared as 10 mg mL−1 in ethanol. For the LDI-MS, we used a smartbeam II laser attached to an Autoflex Max mass spectrometer (Bruker Daltonics, Germany). Immunosuppressant Detection in Human Whole Blood Three immunosuppressants were added to human whole blood to examine the practical usefulness of the g-C3 N4 NS as an LDI-active nanostructure for drug detection in biological fluids. The drug (50 μL, 50 pmol μL−1 ) was added to the entire blood (50 μL), and the sample was incubated for 10 min at room temperature to bind the drugs and red blood cells. Following the addition of acetone (100 μL) and aqueous ZnSO4 (4%, 200 μL) for protein denaturation, the liquid was vortexed for 30 s to release the drugs from the red blood cell surfaces. Blood cells and denatured proteins were separated from the remainder of the liquid by centrifuging the immunosuppressantadded sample at 4000 rpm for 10 min. Liquid–liquid extraction was performed by mixing the supernatant with aqueous NaOH (100 mM, 20 μL) and 1-butanol (200 μL). After forcefully vortexing the mixture for 30 s, it was centrifuged at 4000 rpm for 10 min. Liquids were divided into two layers, with 1-butanol on top and water below, because of the different solvent densities (ρ) of water and 1-butanol. Since the immunosuppressant drugs were soluble exclusively in hydrophobic 1-butanol, they were found on the top layer. Therefore, we collected the top-layer solution by centrifugation. Finally, the extracted sample was resuspended in acetonitrile (100 μL) for LDI-MS analysis using the g-C3 N4 NS.

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6.3 Results and Discussion 6.3.1 Fabrication and Characterization of G-C3 N4 NS Figure 6.1a illustrates a conceptual depiction of this chapter with the instrumental setup. With the help of free carriers generated by a laser pulse in the g-C3 N4 NS, analytes adsorbed on the surface of the g-C3 N4 NS are ionized and subsequently accelerated to an electric field grid, where they then proceed to the mass analyzer and are detected. As a result of the shock waves created by the nanosecond laser pulses, the surface of a g-C3 N4 NS substrate that has been irradiated undergoes sudden and extreme fluctuations in pressure. This causes the substrate to undergo mechanical strain, which in turn distorts the lattice of the g-C3 N4 NS. Destructive shock waves from a powerful laser may break C-N bonds, releasing C and/or N atoms. Also, when subjected to in-situ laser shock pressure, g-C3 N4 NS possessing nanoholes with non-centrosymmetry may produce piezoelectricity. Consequently, atomic defects or piezoelectricity may trap photo-excited free carriers, resulting in less carrier recombination and higher ionization efficiency. We prepared the g-C3 N4 NS as a nanophotonic structure from bulk g-C3 N4 (Fig. 6.2). Our initial step was to create bulk g-C3 N4 by thermally condensing a melamine precursor that already had C-N core structures bound together. The fewlayered 2D g-C3 N4 NS was synthesized by liquid ultrasonic exfoliation; it has a graphite-like carbon network and weak interlayer connections through van der Waals

Fig. 6.1 Characterization of the 2D g-C3 N4 NS. a A schematic illustration of the LDI-MS procedure and experimental apparatus. b HR-TEM image. c HR-XRD patterns. d FTIR spectroscopic results of the 2D g-C3 N4 NS. e Bond length changes in a tri-s-triazine ring during moderate acidification and ultrasonication

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interactions. However, due to the hydrophobic and chemically inert surface of each individual g-C3 N4 NS layer, the layers tended to clump with one another, decreasing the exfoliation yield and restricting its potential uses [10]. The bulk g-C3 N4 was thus treated gently with H2 SO4 for 30 min, washed thoroughly with water, and then ultrasonically agitated to increase its hydrophilicity and exfoliation efficiency.

Fig. 6.2 Simplified flowchart of the steps to make 2D g-C3 N4 NS

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After 48 h, there was still no evidence of precipitation, proving that the colloidal NS dispersion remained stable (Fig. 6.3a). Figures 6.1b and 6.3b confirm the ≈1.71nm-thick ultrathin g-C3 N4 NS sheet, representing 4–5 atomic layers. Figure 6.1c presents the HR-XRD pattern of the bulk g-C3 N4 . In Fig. 6.4, the (100) and (200) peaks at 2θ ≈12.9° and ≈26.1° respectively, demonstrated the existence of an intraplanar tri-s-triazine structural motif [11, 12]. In addition, the (001) and (002) peaks at 2θ ≈14.0° and ≈27.4°, respectively, confirmed the interplanar stacking. The diffraction patterns of the g-C3 N4 NS were found to be consistent with those of bulk g-C3 N4 , indicating that the NS still retained the g-C3 N4 crystalline structure with a long-range order after being moderately acidified and exfoliated. Further, we noticed the XRD peak intensity of the g-C3 N4 NS was massively decreased compared to the bulk one, suggesting the reduced planar size and thickness of the NS. The bandgap energy of the g-C3 N4 NS was calculated to be 2.80 eV greater than that of bulk g-C3 N4 (2.64 eV) because of the quantum confinement effect (Fig. 6.5). Figure 6.1d shows the FTIR spectra of the bulk g-C3 N4 and g-C3 N4 NS, wherein the typical peaks at ≈806 cm−1 (tri-s-triazine ring out-of-plane bending) and ≈1200– 1700 cm−1 (aromatic CN heterocycle stretching) were obtained, revealing the preservation of the tri-s-triazine building blocks [13]. More specifically, the FTIR peaks at ≈1640, 1571, 1544, and 1459 cm−1 were assigned to the C = N stretching vibrations in the tri-s-triazine ring. Additionally, the peaks at ≈1318 and 1236 cm−1 originated from the sp3 -hybridized bridge N atom connecting the three different tri-s-triazine rings (N–(C)3 , Nα ) and corresponded to the stretching vibrations of the C–NH–C or N–(C)3 [14]. N–(C)3 unit asymmetry may have decreased as measured by the weakening of the peaks at 1318 and 1236 cm−1 , as seen in the magnified FTIR spectra. It is expected that the g-C3 N4 NS protonation during the H2 SO4 treatment attenuated the asymmetric vibration of the N–(C)3 , which partially altered the bonding properties

Fig. 6.3 Colloidal 2D g-C3 N4 NS. a Tyndall phenomenon. Pictures of the colloidal suspensions of g-C3 N4 NS before and after being let to stand for >48 h. b Ultrathin sheet shape of the 2D g-C3 N4 NS

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Fig. 6.4 Gaussian-fitted XRD patterns. a (100) and (001) peaks. b (200) and (002) peaks

Fig. 6.5 Optical properties of bulk g-C3 N4 and g-C3 N4 NS. a Absorbance spectra. b Optical bandgap energy

and impacted the vibration mode [14, 15]. Further, a slight blue-shift in the g-C3 N4 NS peaks from ≈1318 and 1236 cm−1 to 1322 and 1243 cm−1 was noticed, which is indicative of shortened Nα –C bond length. Individual g-C3 N4 layers exhibited modest corrugation due to the fact that the sp3 -hybridized N with a lone electron pair was not positioned on the tri-s-triazine plane [16]. As seen in Figs. 6.1e and 6.6, the molecule composed of nine tri-s-triazine rings exhibited a corrugated appearance. It has been observed that in g-C3 N4 , a proton is likely to connect with a lone electron pair of the sp2 -hybridized aromatic N atom (C–N = C, Nβ ) [17]. The tri-s-triazine ring unit becomes electron-deficient as a result of the protonated Nβ , attracting the neighboring long electron pair of the sp3 -hybridized bridge Nα atom to compensate

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Fig. 6.6 Corrugation of a single layer of g-C3 N4 . a before and b after structural optimization. Geometry optimization was performed using a DFT-based ab initio approach

the electron deficiency. This attraction pushes the Nα toward the ring plane, eventually changing the Nα –C bond length in the g-C3 N4 NS. Therefore, the Nα movement toward the ring plane during the H2 SO4 treatment accounts for the shortened Nα –C length as seen in the FTIR results. Although the protonation procedure altered the N–C bond lengths, the presence of FTIR peaks at ≈806 cm−1 and ≈1200–1700 cm−1 clearly demonstrates stable tri-s-triazine building blocks. Thus, we produced the g-C3 N4 NS, which displays a persistent colloidal dispersion in water while conserving its crystal structure based on the conjugated aromatic CN system. It should be noted that we primarily focused on the research of the in situ development of atomic defects and piezoelectricity in g-C3 N4 NS by the laser shock pressure rather than the synthesis procedure. Therefore, maintaining the crystal structure intact was crucial for further exploration. However, alternative g-C3 N4 NS synthesis methods, including Hummer’s approach [18], necessitated complicated experimental conditions, disrupted the crystallinity, and deteriorated the in-plane periodicity of the aromatic systems (Figs. 6.7 and 6.8).

6.3.2 Influence of Nanosecond Laser-Pulse-Driven Shock Pressure on LDI-MS Performance 6.3.2.1

Laser Shock Pressure to G-C3 N4 NS

After the film substrate is irradiated by the nanosecond laser pulse, it takes just a few nanoseconds to get from the first spark of plasma creation to the subsequent expansion of the plasma and the creation of the shock wave [19, 20]. Within the next few tens to a few hundred of nanoseconds, the plasma cools and shrinks. Thus, the laser shock wave exerts a substantial out-of-plane compression on the g-C3 N4 NS surface, which also accompanies an in-plane tension. The mechanical effect of the

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Fig. 6.7 Hummers’ method-based oxidation and exfoliation of g-C3 N4 without precise regulation of the reaction temperature. a HR-XRD pattern and b TEM image. Figure 6.7 shows the crystal structure and morphology of the g-C3 N4 NS product synthesized by adding water into a heated mixture composed of bulk g-C3 N4 , H2 SO4 , and KMnO4 under an ambient environment, and outside of an ice bath. Although the interlayer stacking was detected by the emergence of a (002) peak at 2θ = 27.6°, the disappearance of the diffraction peak at 2θ = 12.9° indicated that the inplanar tri-s-triazine structure was considerably disrupted. The melamine-melem adducts caused new peaks to appear in the XRD pattern: 2C3 N3 (NH2 )3 · C6 N7 (NH2 )3 , C3 N3 (NH2 )3 · C6 N7 (NH2 )3 , and C3 N3 (NH2 )3 · C6 N7 (NH2 )3 . The intra-planar morphology also seemed to be significantly damaged in the oxidized g-C3 N4 NS. After oxidation and exfoliation by the Hummers’ process, if the reaction temperature is not tightly controlled, the in-plane crystallinity and morphology of the g-C3 N4 NS are damaged, and impurities are formed

Fig. 6.8 Hummers’ method-based oxidation and exfoliation of g-C3 N4 under carefully monitored conditions. a HR-XRD pattern and b TEM image. The g-C3 N4 NS generated under strict experimental circumstances using the Hummers’ technique is shown in Fig. 6.8. The diffraction pattern of the as-prepared g-C3 N4 NS showed the presence of the (002) peak at 2θ = 27.6°. However, at 2θ = 12.9°, the tri-s-triazine-based planar structure vanished, indicating the insufficient longrange repetitive order of the in-plane arrangement. There were also detectable amounts of melaminemelem adduct contaminants. The ultrathin sheet shape of the g-C3 N4 NS was degraded, particularly at the edge, as shown in this TEM picture

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Fig. 6.9 Structural deformation in g-C3 N4 NS caused by laser shock pressure. a Illustration of the pressure and plasma generated by a short laser pulse. b Shock pressure P(t,r) as a function of time and distance at a laser power density of 401.2 MW cm−2 . c Pressure as a function of time and radial distance. The pressure as a function of time has a Gaussian form, and its tail is much longer during the damping phase. d Calculations of the peak pressure as a function of incoming laser power density using the Fabbro and Phipps models. e HR-XRD patterns of the g-C3 N4 NS exposed to lasers of varying powers. f Highlighted XRD peaks for the (100), (001), (200), and (002) planes, as determined by Gaussian deconvolution. g Variations in lattice constants a and c as a function of the power density of the incoming laser

laser shock wave on the g-C3 N4 NS was studied by estimating the pressure caused by the laser pulse using the Fabbro model, which characterizes the laser shocking process in a restricted ablation mode [21, 22]. Figure 6.9a presents the generated plasma close to the surface of the irradiated NS, which was placed between the film substrate and analytes. Figure 6.9b shows the shock pressure P(t,r) as a function of time t and radical distance r by laser irradiation at a power density of 401 MW cm−2 . As seen in Fig. 6.9c, the shock pressure varied with both time and radial distance [23]. Figure 6.10 shows P(t,r) distribution maps for varying laser power densities, with the shock pressure rising progressively with increasing laser power. Figure 6.9d shows the peak pressure values. Compared to the Phipps model, which explains direct laser shock in a vacuum [24], the Fabbro model predicted larger peak pressures. The Phipps model predicts that laser-produced plasma would adiabatically grow in a vacuum, rapidly cooling itself. As a result, we found that the Fabbro model yielded greater peak pressure values than the Phipps model. Perfect plasma confinement by the analytes was not possible due to their being desorbed/ionized by laser pulses and subsequently accelerated by an electric field grid. Therefore, we estimated the LDI shock pressure between the Fabbro and Phipps models respective values.

6.3 Results and Discussion Fig. 6.10 Laser shock pressure distribution maps as a function of time and radial distance for a variety of laser power densities. a 317.6, b 348.4, and c 382.3 MW cm−2

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6.3.2.2

6 In Situ Evolution of Atomic Defect and Piezoelectricity in Graphitic …

Atomic Defect Generation in G-C3 N4 NS

We explored the shock-induced g-C3 N4 NS distortion with the use of XRD data. Figure 6.9e shows the HR-XRD patterns of the g-C3 N4 NS after being bombarded with varying laser power densities. We determined the peak positions in the diffraction patterns by Gaussian fitting (Fig. 6.11). The gradual shift in (100) and (200) peaks toward the lower 2θ angle was evident with increasing laser power up to 348.4 MW cm−2 (Fig. 6.9f). In contrast, the (100) and (200) peaks shifted to the higher angle at a high laser power >348.4 MW cm−2 . The (001) and (002) peaks exhibited inverse variations in this deformation trend; the (001) and (002) peaks moved toward the higher 2θ angle with a laser power density ≤348.4 MW cm−2 , while they reversely shifted to the lower angle with a more intense laser irradiation. That is, the laser irradiation led to an anisotropic lattice strain in the g-C3 N4 NS depending on its power (Fig. 6.9g). The exact values of the structural parameters dependent on the laser power densities are shown in Table 6.1. The oppositely varying trends of the lattice deformation may arise from structural faults introduced by the laser shock wave [25]. After being exposed to laser light, the g-C3 N4 NS surface was instantly subjected to a significant out-of-plane compression parallel to the direction of the g-C3 N4 film thickness, also giving rise to the lateral extension [26]. As a result, the out-of-plane contraction and in-plane expansion in the g-C3 N4 NS film occurred at a laser power density ≤348.4 MW cm−2 . However, a considerably powerful laser shock created the atomic defects, e.g., C and N vacancies [27]. The overall CN bonds in the tri-s-triazine rings were reorganized as a consequence of the loss of C and N atoms, resulting in the observed in-plane contraction. In addition, the vacancies disrupted the π-conjugated systems

Fig. 6.11 HR-XRD patterns of g-C3 N4 NS, fitted with a Gaussian, after being subjected to varying laser power densities. a (100) and (001) peaks. b (200) and (002) peaks

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Table 6.1 Changes in the g-C3 N4 NS’s structural properties as a function of laser power Laser power density (MW cm−2 )

Lattice constant (Å)

Cell volume (Å3 )

a

c

0

3.436

6.411

65.548

317.6

3.438

6.381

65.318

348.4

3.442

6.366

65.316

382.3

3.433

6.374

65.056

401.2

3.417

6.394

64.654

by opening the tri-s-triazine aromatic rings, which reduced the strength of the interlayer van der Waals interactions. As a result of the powerful laser, the g-C3 N4 NS became less densely packed between the layers, allowing for the c-axis extension to occur. Thus, the laser power density determined the pattern of the anisotropic distortion in the g-C3 N4 NS, with vacancies being produced most noticeably at high laser power densities >348.4 MW cm−2 . To verify the formation of vacancies, we examined the Raman spectra of the laserirradiated g-C3 N4 NS for evidence of a reduction in delocalized π-electron density [28]. A Gaussian fit to the Raman spectrum of g-C3 N4 NS shows a peak at 705 cm−1 (in-plane tri-s-triazine ring breathing) and a peak at 1229 cm−1 (stretching vibration of the in-plane CN heterocycles) (Fig. 6.12a) [29]. Full-scale Raman spectra are shown in Fig. 6.13. As laser power density was raised, both peaks shifted toward a lower wavenumber, suggesting a weakening of the CN bonds in the tri-s-triazine ring planes (Fig. 6.12b). The in-plane extension at a laser power density of ≤348.4 MW cm−2 is explained by the red-shift in the g-C3 N4 NS Raman bands, as stated in the HR-XRD findings. On the contrary, with a laser power >348.4 MW cm−2 , there was a noticeable blue-shift in the Raman peaks, which may be read as a stiffening of the phonon mode. The shift in the Raman peaks to a high wavenumber (a phenomenon known as “blue-shift”) can be interpreted as the phonon mode hardening, resulting from the decreased aromatic property in the conjugated system [30]. Therefore, the decreased density of the delocalized π electron caused the Raman blue-shift of the g-C3 N4 NS after it was bombarded with a strong laser >348.4 MW cm−2 , which originated from the vacancy-induced breakage of the tri-s-triazine rings and thereby decreased aromaticity. These Raman spectroscopic results reveal that the laser shock structurally deformed the g-C3 N4 NS and the intense laser >348.4 MW cm−2 notably created the defects. Vacancies have the potential to act as electron donors or acceptors, which also have the effect of lowering the rate of charge carrier recombination [31–33]. We used a DFT-based ab initio approach to compute the Mulliken charge distributions to verify that the vacancy can work as a charge-trapping site. The charge-transfer amount in molecules can be evaluated by the Mulliken charges, which are the partial atomic charges [34]. Here, we employed a trimer consisting of three tri-s-triazine rings as a simplified approximation of the g-C3 N4 structure (Fig. 6.12c). We introduced a

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Fig. 6.12 Atomic defects in g-C3 N4 NS caused by the pressure of a laser shock. a Highlighted Raman bands at 705 and 1229 cm−1 with Gaussian fitting. b Variation in the location of the Raman peaks as a function of laser intensity. c Simplified depiction of the g-C3 N4 structure based on the three tri-s-triazine rings. Nedge 1 and Nedge 2 represent the two-coordinated N atoms. The three-coordinated N atom at the center of the tri-s-triazine ring is indicated as Ninner . The threecoordinated C atoms connected with/without Ninner are denoted as Cinner and Cbridge , respectively. Mulliken charges on the C and N atoms in d pristine g-C3 N4 , e N-deficient g-C3 N4 , and f C-deficient g-C3 N4 Fig. 6.13 The g-C3 N4 NS Raman spectra after being exposed to varying laser power densities

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Fig. 6.14 DFT-based optimized geometries of pristine g-C3 N4 , nitrogen-deficient g-C3 N4 , and carbon-deficient g-C3 N4

nitrogen vacancy (VN ) at the two-coordinated nitrogen atom site (Nedge 1 ) in the Ndeficient g-C3 N4 and a carbon vacancy (VC ) at the three-coordinated Cinner atom site in the C-deficient g-C3 N4 . Ab initio computing results in optimized molecular geometries are seen in Fig. 6.14. Figure 6.12d presents the Mulliken charges of the pristine g-C3 N4 on the Cbridge (0.675), Cinner (0.642), Ninner (−0.497), Nedge 1 (−0.452), and Nedge 2 (−0.452) atoms, which varied significantly after a vacancy formed. The Mulliken charges of the VN -adjacent Cbridge (0.398) and Cinner (0.363) atoms in N-deficient g-C3 N4 are shown in Fig. 6.12e; due to electron transfer from the VN site to its surrounds, the VN site was positively charged, making it less positive than the C atoms in pure g-C3 N4 . Contrary to this, the C-deficient g-C3 N4 shows evidence of electron capture at the VC site from its surroundings and a negative charge on the VC -adjacent Ninner (−0.383), Nedge1 (−0.322), and Nedge2 (−0.190) atoms (Fig. 6.12f). In addition, changes in Mulliken charges were less pronounced for atoms farther from the vacancy, suggesting that the electron transfer was most significant around the vacancy (Fig. 6.15). These Mulliken charge studies show that the VN and VC defects in laser-irradiated g-C3 N4 operated as electron and hole trapping sites, respectively, allowing for effective separation of photo-excited carriers. Repetitive multi-pulse laser irradiation of a material apparently leads to a lowering in the threshold of laser-induced damage, thereby producing flaws in the material, which is known as the fatigue effect [35]. The cumulative material changes caused by successive laser beams lead to fatigue. The influence of repeated laser shots on defect development was studied using Raman spectroscopy. We noticed a blue-shift in Raman bands at ≈705 and 1229 cm−1 for g-C3 N4 NS irradiated with 750 and 2000 laser pulses at a laser power density of 401.2 MW cm−2 (Fig. 6.16). The power density of a single laser pulse was constant, although we seldom saw any differences. More laser shots increased the defect formation, as shown by a smaller degree of blue-shift in Raman bands in the 750 laser pulse-irradiated g-C3 N4 NS as compared to the 2000 laser pulse-irradiated sample. During the course of 2000 laser shots, the g-C3 N4 NS was transformed by the initial few rounds (348.4 MW cm−2 by critically deforming the structure, proving the presence of vacancies. Considering that vacancy defects and crystallinity-dependent piezoelectricity are at odds with one another, an optimal laser power density is important for high LDI-MS. Although laser irradiation at 348.4 MW cm−2 power throughout the whole of numerous laser shots may induce piezoelectricity in g-C3 N4 NS, this laser power was inadequate to efficiently create the vacancies in g-C3 N4 NS. This indicates that a laser power density between 348.4 and 382.3 MW cm−2 may provide the best results for LDI-MS. Further, the dipole moment of the g-C3 N4 with both VN and VC defects was computed and it was found to be smaller than that of the pure g-C3 N4 as expected (Fig. 6.17h).

6.3 Results and Discussion

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6.3.3 G-C3 N4 NS as Efficient LDI-MS Substrate Mass analysis of three immunosuppressants (tacrolimus, sirolimus, and everolimus) validated the high LDI-MS results based on the g-C3 N4 NS. We first determined the most productive laser conditions for efficient LDI-MS. Figure 6.23 presents the g-C3 N4 NS-based LDI mass spectra of the three immunosuppressors at varying laser power densities. With a laser power of 360.6 MW cm−2 , the maximum mass peaks for the three immunosuppressants were attained. The laser power 360.6 MW cm−2 . However, it had a negative impact on the LDI-MS outcomes by substantially damaging the g-C3 N4 NS structure and lowering piezoelectricity. As a result, we determined that a laser power density of 360.6 MW cm−2 produced the best LDI-MS results, producing the most desirable lattice strain and creating effective defects as well as piezoelectricity in the g-C3 N4 NS. We analyzed the LDI-MS efficiency of the g-C3 N4 NS substrate in comparison to that of a standard organic 9-AA matrix and a non-piezoelectric graphene NS (Fig. 6.24a). It was only marginally effective to use 9-AA for drug ionization, and it produced matrix fragments, leading to a very low S/N ratio. On the contrary, the S/ N ratio based on the g-C3 N4 NS matrix increased 22.4-fold (44.8, –) for tacrolimus (sirolimus, everolimus), as compared to the 9-AA organic matrix. Alternative organic matrices (DMAN and THAP) rarely allowed for drug ionization (Fig. 6.25). And a 2D graphene sheet similar to the g-C3 N4 NS but without piezoelectricity failed to ionize the three immunosuppressors. Carboxylic acid- and amine-functionalized graphene (graphene-COOH and graphene-NH2 ) were also unsatisfactory. We assessed the analytical validity of the g-C3 N4 NS-based LDI-MS using the shot-to-shot and spot-to-spot reproducibility tests. Figure 6.24b presents the relative standard deviations of the shot-to-shot/spot-to-spot reproducibility for tacrolimus (sirolimus, everolimus), which were all 348.4 MW cm−2 ), atomic defects such N and C vacancies were created, which trapped the particular carriers and increased their lifetimes. Thus, ionization in LDI-MS was favorably impacted by both the laser shock-induced piezoelectricity and the atomic defects.