BLACK WRITING INK ANALYSIS BY DIRECT INFUSION ELECTROSPRAY MASS SPECTROSCOPY by CHRISTOPHER M. MOODY B.S.F.S., B.A. University of Central Florida, 2005 A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science in Forensic Science in the Department of Chemistry in the College of Sciences at the University of Central Florida Orlando, Florida Fall Term 2010 Major Professor: Michael E. Sigman ©2010 Christopher M. Moody ii ABSTRACT An optimized method of extraction, an instrumental analysis method and data analysis was proposed for black writing inks based on direct infusion electrospray-mass spectrometry (ESI-MS). The sampling and analysis method is both minimally destructive and able to assess differences in inks from a reference collection of thirty ballpoint, gel, and rollerball inks. The methanol extracts of ink on paper samples were analyzed with three direct infusion (ESI-MS) methods. Each method varied scan voltage negative and positive, ESI fragmentor applied voltage (+120V, +0V, and -120V), and mobile phase additive. Direct infusion ESI-MS analysis, followed by pair-wise comparisons of the observed ion data in binary form allowed inks to be distinguished from each other. The photobleaching of the dye Basic Violet 3 (BV3) in ink-on- paper samples was examined to determine the use of degradation products as a marker of the age of the writing sample. The extent of photobleaching of BV3 was determined using several illumination sources. Pair-wise comparison of observed ion data was able to distinguish 29 of 30 ink samples using the combined three instrumental methods. Out of 435 pair-wise comparisons 429 pairs could be discriminated from each other using the combined three methods. This is a 98.6% discrimination with the combined analysis scheme. iii To my mother and father: “Man’s mind, once stretched by a new idea, never regains its original dimensions.” — Oliver Wendell Holmes iv ACKNOWLEDGMENTS The completion of this work has been a long process, and I would like to thank anyone who has helped me along the way. I would especially like to thank Dr. Michael Sigman and Mary Williams at NCFS. To my family, real and extended thank you for your patience with me. I would like to thank Kimberly Painter for allowing me use of a quadruple mass analyzer, and for reminding me when it was time to eat. Anyone who has read a draft of this work and added any form of knowledge, love, guidance, constructive criticism or help, I am not sure where to begin to be able to thank everyone individually. However, the proper place to end is apparent. v TABLE OF CONTENTS LIST OF FIGURES ........................................................................................................................ x LIST OF TABLES ....................................................................................................................... xiii CHAPTER ONE: INTRODUCTION ............................................................................................. 1 Ink ............................................................................................................................................... 3 Forensic Document Examination .............................................................................................. 11 Ink Analysis ........................................................................................................................... 11 Non-Destructive Analysis Methods ...................................................................................... 12 Dichroic Filter Analysis .................................................................................................... 13 Digital Processing Methods .............................................................................................. 13 Gas Chromatography/Mass Spectrometry (GC-MS) Analysis of Inks............................. 14 Liquid Chromatography Mass Spectrometry (LC-MS) Analysis of Dyes ................................ 18 Electrospray Ionization (ESI) ................................................................................................ 18 Linear Quadrupole Mass Analyzers ...................................................................................... 22 LC-MS ESI Direct Infusion Analysis of Inks and Dyes ....................................................... 24 Challenges in Ink Analysis .................................................................................................... 25 Other Ink Analysis Methods .................................................................................................. 26 Thin Layer Chromatography (TLC) Analysis ....................................................................... 27 Chemical Spot Tests .............................................................................................................. 27 CHAPTER TWO: EXPERIMENTAL ......................................................................................... 29 vi Sample Preparation ................................................................................................................... 29 Instrumental Methods ................................................................................................................ 30 Direct Infusion Electrospray Ionization Mass Spectrometry (ESI-MS) Analysis ................. 30 Instrumental Parameters ........................................................................................................ 30 Extraction Optimization ............................................................................................................ 31 Direct Infusion ESI-MS Optimization ...................................................................................... 32 Spectral Subtraction to Yield “Pure Ink” Spectrum .............................................................. 32 Ink Volatiles Persistence Study ............................................................................................. 33 Effect of Paper on Extraction ................................................................................................ 34 Dye and Vehicle Standards ................................................................................................... 34 Photochemical Experiments ...................................................................................................... 36 Light Exposure Effects .......................................................................................................... 38 Potassium Ferrioxalate Actinometry ..................................................................................... 38 Method Validation of Direct Infusion ESI-MS Ink Analysis ................................................ 40 Analysis of Triplicate Run Data ....................................................................................... 40 Same Manufacturer Multipack Pen Analysis ................................................................... 40 Intersecting and Overlapping Ink Strokes (Deposition Order Determination) ......................... 41 Direct Infusion LC-MS .......................................................................................................... 41 CHAPTER THREE: RESULTS ................................................................................................... 44 Direct Infusion Electrospray Ionization Mass Spectrometry (ESI-MS) Analysis .................... 44 vii Extraction Optimization ........................................................................................................ 44 Direct Infusion ESI-MS Optimization ................................................................................... 44 Method 1 (+120V) ............................................................................................................ 44 Method 2 (+0 V) ............................................................................................................... 46 Method 3 (-120V) ............................................................................................................. 47 Volatile Persistence Study ................................................................................................ 47 Analysis of Standards ............................................................................................................ 48 Basic and Disperse Type Dye Analysis ............................................................................ 48 Acid and Solvent Type Dye Analysis ............................................................................... 48 Vehicle Analysis .................................................................................................................... 48 Photochemical Experiments ...................................................................................................... 51 Light Exposure Effects .......................................................................................................... 51 Potassium Ferrioxalate Actinometry................................................................................. 53 Method Validation .................................................................................................................... 54 Triplicate Analysis of Reference Pen Collection .................................................................. 54 Multipack Analysis ........................................................................................................... 60 Intersecting and Overlapping Ink Strokes ................................................................................