Database of proteinogenic amino acid Accession #: 01453, 01454, 01455, 01456, 01457, 01458, 01459, reference spectra for Bismuth-cluster 01460, 01461, 01462, 01463, 01464, 01465, 01466, 01467, ToF-SIMS. II. Positive polarity 01468, 01469, 01470, 01471, Maciej Kaweckia) 01472, 01473, 01474 Laboratory of Nanoscale Materials Science, Empa, CH-8600 Dubendorf,€ Switzerland and Technique: SIMS Department of Physics, University of Basel, CH-4056 Basel, Switzerland Host Material: Silicon (100) wafer Laetitia Bernard Instrument: IONTOF TOF-SIMS.5 Laboratory of Nanoscale Materials Science, Empa, CH-8600 Dubendorf,€ Switzerland Major Species in Spectra: C, H, N, O, S (Received 6 February 2018; accepted 1 May 2018; published 10 July 2018) Minor Species in Spectra: Na, K, Si The number of time-of-flight secondary ion mass spectrometry studies on biological tissues and Published Spectra: 22 cells strongly increased since the development of primary ion sources that allow not only elemental but also molecular analysis. Still, substantial fragmentation during ionic bombardment Spectra in Electronic Record: 66 results in a large amount of peaks, rendering data analysis complex. Complete and trustable sets of Published Figures: 44 reference spectra for the main biological building blocks such as proteins, sugars, and lipids are Spectral Category: Reference required. Here, the authors provide a database of reference spectra for the 21 proteinogenic amino acids þ glycine for bismuth instruments. Paper 1 of this two-part article series contains the nega- tive polarity spectra and Paper II the positive polarity spectra. These reference spectra were þ þ þþ obtained with the primary ion Bi3 , and spectra for the primary ions Bi1 and Bi3 are contained in the supplementary material as ASCII files. Published by the AVS. https://doi.org/10.1116/1.5024988 Keywords: ToF-SIMS, amino acid, mass spectrometry, fragmentation

INTRODUCTION made sure that each deposited droplet is spread over the entire wafer surface before leaving the wafers in the laminar Partial amino acid databases for time-of-flight secondary ion flow chamber for drying. mass spectrometry (ToF-SIMS) reference mass spectra have The spectra were obtained on a ToF-SIMS.5 instrument (ION- been already conducted based on the now largely replaced TOF) equipped with a 25 keV bismuth-manganese liquid metal monoatomic argon (Ref. 1) and caesium (Ref. 2) primary ion ion gun. The instrument was operated in high current bunched sources. The characteristic molecular fragmentation in ToF- mode. The primary ion dose was kept below the static limit SIMS measurements, however, depends on the primary ion (1012 ions/cm2) for each spectrum, and a low-energy electron impacting the surface. The current standard primary ion source flood gun was used to compensate charging effects. The raster is the bismuth/manganese liquid metal ion source. For bismuth size was 200 Â 200 lm2 with a raster resolution of 128 Â 128, and bismuth cluster primary ions, only limited lists attributing a cycle time of 100 ls and primary ion currents of 1.54 pA for prominent peaks to specific amino acids can be found in the lit- Bi þ, 0.37–0.4 pA for Bi þ, and 0.44 pA for Bi þþ. Fifty scans erature, e.g., Ref. 3. Here, we provide a database of original ref- 1 3 3 were acquired per single mass spectrum. All presented spectra erence spectra for the 21 proteinogenic amino acids þ glycine are normalized to their respective total ion count. Note that as well as the complete corresponding peak lists determining variations in mass resolution among the reference spectra (see the fragmentation products. Fragments which uniquely repre- Fig. 23) are due to crystallization geometry differences sent specific amino acids are highlighted in Table 24. This between the amino acids. Paper II of the database contains the positive polarity spectra. Negative polarity spectra are contained in Paper I (Ref. 4). Figures and tables presented contain only attributions for the most prominent peaks. Complete peak-lists attributing fragmenta- The amino acids were purchased through Sigma Aldrich tion products to 50–100 main peaks for each amino acid are (Article number: 09416, purity > 99.0%). Each amino acid included in the attachment. In the figures, the full molecule is was dissolved in H O (Sigma Aldrich, Article number: 2 referred to with “M.” Polymerization products are marked with 270733) and 0.1 M solutions were so prepared. Host silicon “p.p.”, cationization products with “c.p.,” and contaminants with wafers of 1 Â 1cmsizewerefirstwashedbysubmersionin “c.” Major peaks where a unique attribution was not possible, subsequent ultrasound baths of 2 Â 15mininacetoneand e.g., due to an overlap beyond mass resolution of two potential 1 Â 15 min in ethanol, and then plasma-treated for a duration fragmentation products, are marked with “n.a.” standing for not of 10 min to render the surface hydrophilic. The 0.1 M amino (uniquely) attributed. In the label of the supplementary material acid solutions were subsequently drop-deposited inside a files, the Spectrum ID# uniquely identifies each amino acid. Mass laminar flow chamber on separate silicon wafers. It was þ þ þþ spectra acquired using Bi3 ,Bi1 ,andBi3 primary ions are further identified by the file-suffix “01,” “02,” and “03,” respec- a)Electronic mail: [email protected] tively, and peak lists in ASCII format by the file-suffix “04.”

Surface Science Spectra, Vol. 25, 2018 1055-5269/2018/25(1)/015002/29/$30.00 Published by the AVS. 015002-1 SPECIMEN DESCRIPTION (ACCESSION # 01453, 01454, In Situ Preparation: None 01455, 01456, 01457, 01458, 01459, 01460, 01461, 01462, Charge Control: Low energy electrons 01463, 01464, 01465, 01466, 01467, 01468, 01469, 01470, Temp. During Analysis: 300 K 01471, 01472, 01473, 01474) Pressure During Analysis: 5.6 Â 10À8 Pa Host Material: Silicon (100) wafer Preanalysis Beam Exposure: None CAS Registry #: 7440-21-3 Host Material Characteristics: Homogeneous; solid; single crys- INSTRUMENT CONFIGURATION tal; semiconductor; semiconductor Manufacturer and Model: IONTOF TOF-SIMS.5 Chemical Name: L-alanine (01453), L-arginine monohydrochlor- Analyzer Type: Time-of-flight ide (01454), L-asparagine (01455), L-asparatic acid (01456), Detector Type: Microchannel plate L-cysteine (01457), L-cystine (01458), L-glutamic acid (01459), L-glutamine (01460), glycine (01461), L-histidine Experiment Type: Mass spectra monohydrochloride (01462), L–4-hydroxiproline (01463), Sample Rotation: None L-isoleucine (01464), L-leucine (01465), L-lysine monohydro- chloride (01466), L-methionine (01467), L-phenylalanine Oxygen Flood Source: None (01468), L-proline (01469), L-serine (01470), L-threonine Other Flood Source: Low energy electrons (01471), L-tryptophan (01472), L-tyrosine (01473), L-valine Energy Acceptance Window: 14.3 eV (01474) Postacceleration Voltage: 10 000 eV Source: Sigma Aldrich Sample Bias: 0eV Host Composition: Si and SiO2 Specimen Normal-to-analyzer (He): 0 Form: Crystalline powder dissolved in DI H2O and subsequently drop-deposited on the host wafer ᭿ Ion Sources

Lot Number: L-alanine: BCBS2461V, L-arginine monohydrochlor- Ion Source 1 of 1 ide: 1361811V, L-asparagine: 1371738V, L-asparatic acid: Purpose of this Ion Source: Analysis beam BCBR6815V, L-cysteine: BCBR4500V, L-cystine: BCBS1903V, L-glutamic acid: 1423805V, L-glutamine: BCBR7569V, glycine: Ion Source Manufacturer: IONTOF (Munster,€ Germany) 1119375V, L-histidine monohydrochloride: BCBS1876V, L-4- Ion Source Model: 25 keV Bi/Mn cluster source hydroxiproline: BCBL2666V, L-isoleucine: 1423806V, L-leu- cine: BCBBQ9986V, L-lysine monohydrochloride: Beam Mass Filter: Electrodynamic mass filter þ BCBN9886V, L-methionine: 1423807V, L-phenylalanine: Beam Species and Charge State: Bi3 BCBQ1493V, L-proline: BCBP4505V, L-serine: 1336081V, Beam Gating Used: None L-threonine: BCBD4901V, L-tryptophan: BCBR0168V, L-tyrosine: BCBR7417V, L-valine: BCBM0163V Beam Voltage: 25 000 eV Net Beam Voltage (impact voltage): 25 000 eV Structure: L-alanine: C3H7NO2, L-arginine monohydrochloride: C6H14N4O2ÁHCl, L-asparagine: C4H8N2O3, L-asparatic acid: Ion Pulse Width: 29.3 ns C H NO , L-cysteine: C H NO S, L-cystine: C H N O S , 4 7 4 3 7 2 6 12 2 4 2 Ion Pulse Rate: 10 kHz L-glutamic acid: C5H9NO4, L-glutamine: C5H10N2O3, glycine: C2H5NO2, L-histidine monohydrochloride: C6H9N3O2ÁHCl, L-4- Pulsed Beam Current: 0.00037–0.0004 nA hydroxiproline: C5H9NO3, L-isoleucine: C6H13NO2, L-leucine: Current Measurement Method: Faraday cup C H NO , L-lysine monohydrochloride: C H N O ÁHCl, 6 13 2 6 14 2 2 Beam Raster Size: 200 Â200 lm2 L-methionine: C5H11NO2S, L-phenylalanine: C9H11NO2, L-proline:  C5H9NO2, L-serine: C3H7NO3, L-threonine: C4H9NO3, L-trypto- Beam Incident Angle: 45 phan: C11H12N2O2, L-tyrosine: C9H11NO3, L-valine: C5H11NO2 Source-to-Analyzer Angle: 45 History and Significance: The proteinogenic amino acids are the building blocks of all proteins in the known life forms. ACKNOWLEDGMENTS Complete spectra databases of organic molecules facilitate the interpretation of ToF-SIMS data from biological specimen. The authors are thankful for the funding received from the Swiss However, it should be kept in mind that in biological specimens National Science Foundation through the Grant No. CR23I2-162828. amino acids are incorporated into larger molecules (proteins, peptidoglycans, etc.) which affects the fragmentation patterns. REFERENCES As Received Condition: Samples received in powder form 2 1. Preinstalled database on IONTOF GmbH instruments. Analyzed Region: Regions of 200 Â 200 lm on maximally 2. N. T. Samuel, M. S. Wagner, K. D. Dornfeld, and D. G. homogeneous specimen sites Castner, Surf. Sci. Spectra 8, 163 (2001). Ex Situ Preparation/Mounting: 0.1 M amino acid solutions were 3. M. Urbini, V. Piteto, F. de Notaristefani, F. Scaldaferri, A. prepared in H2O and drop-deposited on separate 1 Â 1 cm large Gasbarrini, and L. Tortora, Anal. Bioanal. Chem. 409, 6097 (2017). plasma-treated Si-Wafers in a laminar flow chamber. 4. M. Kawecki and L. Bernard, Surf. Sci. Spectra 25, 015001 (2018).

015002-2 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra ALANINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01453 44.0495 C2H6N Alanine þ … 55.0178 C3H3O … þ … 56.0495 C3H6N … þ … 70.0287 C3H4NO … þ … 74.0600 C3H8NO … þ … 90.0550 C3H8NO2 …

Accession # 01453-01, 01453-02, 01453-03, 01453-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH4N ,C3H7NO2 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-3 ARGININE (HYDROCHLORIDE) Spectrum ID # Mass (Da) Species Peak Assignment

þ 01454 60.0556 CH6N3 Arginine þ … 70.0777 C5H10 … þ … 73.0634 C2H7N3 … þ … 87.0791 C3H9N3 … þ … 100.0869 C4H10N3 … þ … 112.0869 C5H10N3 … þ … 130.1213 C5H14N4 … þ … 160.1081 C6H14N3O2 … þ … 175.1190 C6H15N4O2 …

Accession # 01454-01, 01454-02, 01454-03, 01454-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C6H14N4O2 Primary Ion Dose 4.73 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.00037 nA Beam Raster Size 200 Â 200 lm2

015002-4 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra ASPARAGINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01455 70.0287 C3H4NO Asparagine þ … 74.0237 C2H4NO2 … þ … 87.0553 C3H7N2O … þ … 89.0709 C3H9N2O … þ … 116.0342 C4H6NO3 … þ … 133.0608 C4H9N2O3 …

Accession # 01455-01, 01455-02, 01455-03, 01455-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,CH2N ,C4H10N2O3 Primary Ion Dose 4.73 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.00037 nA Beam Raster Size 200 Â 200 lm2

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-5 ASPARATIC ACID Spectrum ID # Mass (Da) Species Peak Assignment

þ 01456 70.0287 C3H4NO Asparatic acid þ … 74.0237 C2H4NO2 … þ … 88.0393 C3H6NO2 … þ … 90.0550 C3H8NO2 … þ … 134.0448 C4H8NO4 …

Accession # 01456-01, 01456-02, 01456-03, 01456-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,CH2N ,C4H9NO4 Primary Ion Dose 4.73 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.00037 nA Beam Raster Size 200 Â 200 lm2

015002-6 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra CYSTEINE Spectrum ID # Mass (Da) Species Peak Assignment 01457 31.9715 Sþ Cysteine þ … 34.9950 H3S … … 44.9793 CHSþ … þ … 46.9950 CH3S … þ … 58.9950 C2H3S … þ … 75.0137 C2H5NS … þ … 76.0215 C2H6NS … þ … 86.9899 C3H3OS … þ … 88.0393 C3H6NO2 … þ … 90.0550 C3H8NO2 … þ … 105.0005 C3H5O2S … þ … 122.0270 C3H8NO2S …

Accession # 01457-01, 01457-02, 01457-03, 01457-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,CH2N ,C3H7SNO2 Primary Ion Dose 4.73 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.00037 nA Beam Raster Size 200 Â 200 lm2

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-7 CYSTINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01458 63.9436 S2 Cystine þ … 58.9950 C2H3S … þ … 76.0215 C2H6NS … þ … 120.0114 C3H6NO2S … þ … 122.0270 C3H8NO2S … þ … 151.9834 C3H6NO2S2 … þ … 195.0256 C5H11N2O2S2 … þ … 207.0256 C6H11N2O2S2 … þ … 241.0311 C6H13N2O4S2 …

Accession # 01458-01, 01458-02, 01458-03, 01458-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C6H14N2O4S2 Primary Ion Dose 4.73 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.00037 nA Beam Raster Size 200 Â 200 lm2 þ Comment Region of interest, defined by a minimum threshold of the C2H4NO2 signal, was applied to eliminate the substrate signal.

015002-8 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra GLUTAMIC ACID Spectrum ID # Mass (Da) Species Peak Assignment

þ 01459 84.0444 C4H6NO Glutamic acid þ … 84.0808 C5H10N … þ … 91.0628 C3H9NO2 … þ … 102.0550 C4H8NO2 … þ … 148.0604 C5H10NO4 …

Accession # 01459-01, 01459-02, 01459-03, 01459-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C5H10NO4 Primary Ion Dose 4.73 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.00037 nA Beam Raster Size 200 Â 200 lm2

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-9 GLUTAMINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01460 84.0444 C4H6NO Glutamine þ … 86.0600 C4H8NO … þ … 101.0709 C4H9N2O … þ … 102.0550 C4H8NO2 … þ … 103.0866 C4H11N2O … þ … 114.0550 C5H8NO2 … þ … 130.0499 C5H8NO3 … þ … 147.0764 C5H11N2O3 …

Accession # 01460-01, 01460-02, 01460-03, 01460-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,CH2N ,C5H12N2O3 Primary Ion Dose 4.73 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.00037 nA Beam Raster Size 200 Â 200 lm2

015002-10 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra GLYCINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01461 43.0718 C2H3O Glycine þ … 60.0444 C2H6NO … þ … 76.0393 C2H6NO2 …

Accession # 01461-01, 01461-02, 01461-03, 01461-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C2H5NO2 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-11 HISTIDINE (HYDROCHLORIDE) Spectrum ID # Mass (Da) Species Peak Assignment

þ 01462 81.0447 C4H5N2 Histidine þ … 82.0526 C4H6N2 … þ … 83.0604 C4H7N2 … þ … 93.0447 C5H5N2 … þ … 94.0526 C5H6N2 … þ … 95.0604 C5H7N2 … þ … 110.0713 C5H8N3 … þ … 156.0768 C6H10N3O2 …

Accession # 01462-01, 01462-02, 01462-03, 01462-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C6H9N3O2 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

015002-12 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra HYDROXYPROLINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01463 68.0621 C5H8 Hydroxyproline þ … 74.0237 C2H4NO2 … þ … 84.0444 C4H6NO … þ … 86.0600 C4H8NO … þ … 87.0679 C4H9NO … þ … 88.0757 C4H10NO … þ … 114.0550 C5H8NO2 … þ … 116.0706 C5H10NO2 … þ … 132.0655 C5H10NO3 …

Accession # 01463-01, 01463-02, 01463-03, 01463-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,C5H9NO3 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-13 ISOLEUCINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01464 69.0699 C5H9 Isoleucine þ … 70.0651 C4H8N … þ … 75.0315 C2H5NO2 … þ … 84.0808 C5H10N … þ … 86.0964 C5H12N … þ … 87.1043 C5H13N … þ … 132.1019 C6H14NO2 …

Accession # 01464-01, 01464-02, 01464-03, 01464-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,CH2N ,C6H13NO2 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

015002-14 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra LEUCINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01465 70.0777 C5H10 Leucine þ … 74.0237 C2H4NO2 … þ … 84.0808 C5H10N … þ … 86.0964 C5H12N … þ … 132.1019 C6H14NO2 …

Accession # 01465-01, 01465-02, 01465-03, 01465-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,CH2N ,C6H13NO2 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-15 LYSINE (HYDROCHLORIDE) Spectrum ID # Mass (Da) Species Peak Assignment

þ 01466 84.0808 C5H10N Lysine þ … 86.0954 C5H12N … þ … 91.0628 C3H9NO2 … þ … 96.0808 C6H10N … þ … 98.0964 C6H12N … þ … 110.0839 C6H10N2 … þ … 128.0706 C6H10NO2 … þ … 132.1019 C6H14NO2 … þ … 147.1128 C6H15N2O2 …

Accession # 01466-01, 01466-02, 01466-03, 01466-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C6H14N2O2 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

015002-16 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra METHIONINE Spectrum ID # Mass (Da) Species Peak Assignment 01467 31.9715 Sþ Methionine þ … 46.9950 CH3S … þ … 61.0106 C2H5S … þ … 74.0185 C3H6S … þ … 75.0315 C2H5NO2 … þ … 88.0215 C3H6NS … þ … 88.0341 C4H8S … þ … 102.0550 C4H8NO2 … þ … 104.0528 C4H10NS … þ … 133.0318 C5H9O2S … þ … 150.0583 C5H12NO2S …

Accession # 01467-01, 01467-02, 01467-03, 01467-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C5H13SNO2 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2 þ Comment The assignments C2H5O2 and C3H6O2þ (in brackets) show a substantially lower deviation but are, in the author’s judgment, sufficiently inconsistent with the fragmentation patterns observed in other amino acids to warrant the choice of peak attribution made above.

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-17 PHENYLALANINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01468 77.0386 C6H5 Phenylalanine þ … 91.0628 C3H9NO2 … þ … 103.0542 C8H7 … þ … 105.0699 C8H9 … þ … 107.0491 C7H7O … þ … 115.0628 C5H9NO2 … þ … 120.0808 C8H10N … þ … 121.0886 C8H11N … þ … 166.0863 C9H12NO2 …

Accession # 01468-01, 01468-02, 01468-03, 01468-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C9H11NO2 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

015002-18 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra PROLINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01469 68.0621 C5H8 Proline þ … 70.0651 C4H8N … þ … 70.0777 C5H10 … þ … 71.0730 C4H9N … þ … 72.0808 C4H10N … þ … 116.0706 C5H10NO2 …

Accession # 01469-01, 01469-02, 01469-03, 01469-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,C5H9NO2 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-19 SERINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01470 60.0444 C2H6NO Serine þ … 70.0287 C3H4NO … þ … 73.0284 C3H5O2 … þ … 88.0393 C3H6NO2 … þ … 106.0499 C3H8NO3 …

Accession # 01470-01, 01470-02, 01470-03, 01470-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C3H9NO3 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

015002-20 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra THREONINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01471 56.0621 C4H8 Threonine þ … 74.0600 C3H8NO … þ … 84.0444 C4H6NO … þ … 102.0550 C4H8NO2 … þ … 120.0655 C4H10NO3 …

Accession # 01471-01, 01471-02, 01471-03, 01471-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C4H11NO3 Primary Ion Dose 5.11 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.0004 nA Beam Raster Size 200 Â 200 lm2

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-21 TRYPTOPHAN Spectrum ID # Mass (Da) Species Peak Assignment

þ 01472 77.0386 C6H5 Tryptophan þ … 115.0417 C8H5N … þ … 117.0573 C8H7N … þ … 118.0651 C8H8N … þ … 130.0651 C9H8N … þ … 132.0808 C9H10N … þ … 143.0730 C10H9N … þ … 144.0682 C9H8N2 … þ … 169.0917 C10H11N2 … þ … 188.0944 C11H12N2O … þ … 205.0972 C11H13N2O2 …

Accession # 01472-01, 01472-02, 01472-03, 01472-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C11H13N2O2 Primary Ion Dose 4.73 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.00037 nA Beam Raster Size 200 Â 200 lm2

015002-22 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra TYROSINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01473 77.0386 C6H5 Tyrosine þ … 91.0542 C7H7 … þ … 107.0491 C7H7O … þ … 115.0417 C8H5N … þ … 121.0648 C8H9O … þ … 136.0757 C8H10NO … þ … 149.0107 C7H3NO3 … þ … 165.0546 C9H9O3 … þ … 182.0812 C9H12NO3 …

Accession # 01473-01, 01473-02, 01473-03, 01473-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C9H12NO3 Primary Ion Dose 4.73 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.00037 nA Beam Raster Size 200 Â 200 lm2

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-23 VALINE Spectrum ID # Mass (Da) Species Peak Assignment

þ 01474 55.0542 C4H7 Valine þ … 68.0495 C4H6N … þ … 72.0808 C4H10N … þ … 73.0886 C4H11N … þ … 75.0315 C2H5NO2 … þ … 118.0862 C5H12NO2 …

Accession # 01474-01, 01474-02, 01474-03, 01474-04 ᭿ Host Material Silicon wafer ᭿ Technique SIMS Secondary Source Polarity Positive Mass Range Minimum 0Da Mass Range Maximum 250 Da þ þ þ þ þ Species Used for Mass Calibration CH3 ,NH3 ,NH4 ,CH2N ,C5H13NO2 Primary Ion Dose 4.73 Â 1011 cmÀ2 þ Primary Species Bi3 Primary Ion Pulse Width 29.3 ns Pulsed Beam Current 0.00037 nA Beam Raster Size 200 Â 200 lm2

015002-24 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra EXPANDED [M1H]1 PEAKS FOR EACH AMINO ACID SPECTRUM

01453-01, 01454-01, 01455-01, 01456-01, 01457-01, 01458-01, 01459-01, 01460-01, 01461-01, 01462-01, 01463-01, 01464-01, 01465-01, 01465-01, 01467-01, 01468-01, 01469-01, 01470-01, 01471-01, 01472-01, Accession # 01473-01, 01474-01

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-25 SPECTRAL FEATURES TABLE Mass (Da) Peak assignment Peak present in following amino acids: Peak most prominent in:

þ 17.0260 NH3 all amino acids — þ 18.0338 NH4 all amino acids — þ 28.0182 CH2N all amino acids — þ 30.0338 CH4N all amino acids — 31.9715 Sþ cysteine, cystine, methionine cysteine, cystine, methionine þ 34.9950 SH3 cysteine, cystine, methionine cysteine, cystine, methionine þ 39.0229 C3H3 all amino acids except for glycine — þ 41.0386 C3H5 all amino acids except for glycine — þ 42.0338 C2H4N all amino acids — þ 43.0417 C2H5N all amino acids — þ 43.0542 C3H7 most amino acids — þ 43.0718 C2H3O all amino acids — þ 44.0495 C2H6N all amino acids — 44.9793 CHSþ cysteine, cystine, methionine cysteine, cystine, methionine þ 45.0335 C2H5O all amino acids — þ 45.0573 C2H7N all amino acids — þ 45.9872 CH2S cysteine, cystine, methionine cysteine, cystine, methionine þ 46.0651 C2H8N all amino acids — þ 46.9950 CH3S cysteine, cystine, methionine cysteine, cystine, methionine þ 47.0491 C2H7O all amino acids — þ 51.0229 C4H3 multiple amino acids — þ 53.0386 C4H5 multiple amino acids — þ 55.0178 C3H3O all amino acids except for glycine — þ 55.0542 C4H7 multiple amino acids — þ 56.0495 C3H6N all amino acids except for glycine — þ 56.0621 C4H8 multiple amino acids — þ 57.0335 C3H5O all amino acids except for glycine — þ 57.0573 C3H7N multiple amino acids — þ 58.0287 C2H4NO all amino acids — þ 58.0651 C3H8N all amino acids except for glycine — þ 58.0777 C4H10 multiple amino acids — þ 58.9950 C2H3S cysteine, cystine, methionine cysteine, cystine, methionine þ 59.0491 C3H7O all amino acids except for glycine — þ 60.0444 C2H6NO all amino acids — þ 60.0556 CH6N3 arginine arginine þ 61.0106 C2H5S cysteine, cystine, methionine cysteine, cystine, methionine þ 61.0284 C2H5O2 all amino acids — þ 63.9436 S2 cystine cystine þ 67.0542 C5H7 multiple amino acids — þ 68.0495 C4H6N multiple amino acids — þ 68.0621 C5H8 multiple amino acids — þ 69.0335 C4H5O multiple amino acids — þ 69.0447 C3H5N2 arginine, asparagine, histidine asparagine, histidine þ 69.0573 C4H7N multiple amino acids — þ 69.0699 C5H9 multiple amino acids — þ 70.0287 C3H4NO all amino acids except for glycine —

015002-26 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra Mass (Da) Peak assignment Peak present in following amino acids: Peak most prominent in:

þ 70.0651 C4H8N multiple amino acids — þ 70.0777 C5H10 multiple amino acids — þ 71.0491 C4H7O multiple amino acids — þ 71.0730 C4H9N multiple amino acids proline þ 72.0444 C3H6NO all amino acids except for glycine — þ 72.0808 C4H10N multiple amino acids — þ 73.0284 C3H5O2 multiple amino acids — þ 73.0634 C2H7N3 arginine arginine þ 73.0648 C4H9O multiple amino acids — þ 73.0886 C4H11N valine valine þ 74.0237 C2H4NO2 all amino acids — þ 74.0362 C3H6O2 all amino acids except for glycine — þ 74.0600 C3H8NO all amino acids except for glycine — þ 75.0137 C2H5NS cysteine, cystine, methionine cysteine, cystine þ 75.0315 C2H5NO2 all amino acids — þ 76.0215 C2H6NS cysteine, cystine, methionine cysteine, cystine þ 76.0393 C2H6NO2 all amino acids — þ 77.0386 C6H5 multiple amino acids — þ 80.0495 C5H6N multiple amino acids — þ 80.0621 C6H8 multiple amino acids — þ 81.0447 C4H5N2 multiple amino acids histidine þ 82.0526 C4H6N2 multiple amino acids histidine þ 82.0651 C5H8N multiple amino acids — þ 82.0777 C6H10 multiple amino acids — þ 83.0604 C4H7N2 multiple amino acids histidine þ 84.0444 C4H6NO multiple amino acids — þ 84.0808 C5H10N multiple amino acids —

84.0933 C6H12þ multiple amino acids — þ 86.0600 C4H8NO multiple amino acids hydroxyproline þ 86.0964 C5H12N multiple amino acids isoleucine, leucine þ 86.9899 C3H3OS cysteine, cystine cysteine, cystine þ 87.0553 C3H7N2O asparagine asparagine þ 87.0679 C4H9NO multiple amino acids hydroxyproline þ 87.0791 C3H9N3 arginine arginine þ 87.0804 C5H11O multiple amino acids — þ 87.1043 C5H13N multiple amino acids isoleucine, leucine þ 88.0215 C3H6NS methionine methionine þ 88.0341 C4H8S methionine methionine þ 88.0393 C3H6NO2 multiple amino acids — þ 88.0757 C4H10NO multiple amino acids hydroxyproline þ 89.0386 C7H5 phenylalanine, tryptophan, tyrosine — þ 89.0709 C3H9N2O asparagine asparagine þ 90.0550 C3H8NO2 multiple amino acids — þ 91.0542 C7H7 phenylalanine, tryptophan, tyrosine — þ 93.0447 C5H5N2 arginine, histidine, lysine histidine þ 94.0526 C5H6N2 arginine, histidine, lysine histidine

95.0604 C5H7N2þ arginine, histidine, lysine histidine

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-27 Mass (Da) Peak assignment Peak present in following amino acids: Peak most prominent in:

þ 96.0808 C6H10N isoleucine, leucine, lysine — þ 98.0964 C6H12N isoleucine, leucine, lysine — þ 100.0393 C4H6NO2 multiple amino acids — þ 100.0869 C4H10N3 arginine arginine þ 101.0709 C4H9N2O asparagine, glutamine glutamine þ 102.0464 C8H6 phenylalanine, tryptophan, tyrosine — þ 102.0550 C4H8NO2 multiple amino acids — þ 103.0542 C8H7 phenylalanine, tryptophan, tyrosine phenylalanine þ 103.0866 C4H11N2O asparagine, glutamine glutamine þ 104.0528 C4H10NS methionine methionine þ 105.0005 C3H5O2S cysteine, dystine cysteine, cystine þ 105.0699 C8H9 phenylalanine, tyrosine phenylalanine þ 106.0499 C3H8NO3 serine, threonine serine þ 107.0491 C7H7O phenylalanine, tyrosine — þ 107.0577 C3H9NO3 hydroxyproline, serine, threonine serine þ 110.0713 C5H8N3 arginine, histidine histidine þ 110.0839 C6H10N2 lysine lysine þ 112.0869 C5H10N3 arginine arginine þ 114.0550 C5H8NO2 multiple amino acids — þ 114.1026 C5H12N3 arginine arginine þ 115.0417 C8H5N phenylalanine, tryptophan, tyrosine — þ 115.0628 C5H9NO2 multiple amino acids — þ 116.0342 C4H6NO3 asparagine asparagine þ 116.0706 C5H10NO2 multiple amino acids proline þ 117.0573 C8H7N phenylalanine, tryptophan, tyrosine — þ 118.0651 C8H8N phenylalanine, tryptophan, tyrosine phenylalanine, tryptophan þ 118.0862 C5H12NO2 multiple amino acids asparatic acid, glutamic acid, valine þ 119.0491 C8H7O phenylalanine, tryptophan, tyrosine — þ 120.0114 C3H6NO2S cysteine, cystine cysteine, cystine þ 120.0444 C7H6NO tryptophan, tyrosine tryptophan þ 120.0655 C4H10NO3 threonine threonine þ 120.0808 C8H10N phenylalanine phenylalanine þ 121.0648 C8H9O phenylalanine, tryptophan, tyrosine — þ 121.0886 C8H11N phenylalanine phenylalanine þ 122.0270 C3H8NO2S cysteine, cystine cysteine, cystine þ 128.0706 C6H10NO2 isoleucine, leucine, lysine isoleucine, leucine, lysine þ 129.0420 C5H7NO3 glutamic acid, glutamine, hydroxyproline glutamine þ 129.1135 C5H13N4 arginine arginine þ 130.0499 C5H8NO3 glutamic acid, glutamine, hydroxyproline glutamine þ 130.0651 C9H8N phenylalanine, tryptophan, tyrosine tryptophan þ 130.1213 C5H14N4 arginine arginine þ 131.0730 C9H9N phenylalanine, tryptophan, tyrosine tryptophan þ 131.1291 C5H15N4 arginine arginine þ 132.0655 C5H10NO3 glutamic acid, glutamine, hydroxyproline hydroxyproline þ 132.0808 C9H10N phenylalanine, tryptophan, tyrosine tryptophan þ 132.1019 C6H14NO2 isoleucine, leucine, lysine isoleucine, leucine þ 133.0318 C5H9O2S methionine methionine

015002-28 Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra Mass (Da) Peak assignment Peak present in following amino acids: Peak most prominent in:

þ 133.0608 C4H9N2O3 asparagine asparagine þ 134.0448 C4H8NO4 asparatic acid asparatic acid þ 136.0757 C8H10NO tyrosine tyrosine þ 143.0730 C10H9N tryptophan tryptophan þ 144.0682 C9H8N2 tryptophan tryptophan þ 147.0764 C5H11N2O3 glutamine glutamine þ 147.1128 C6H15N2O2 lysine lysine þ 148.0604 C5H10NO4 glutamic acid glutamic acid þ 149.0107 C7H3NO3 tyrosine tyrosine þ 150.0583 C5H12NO2S methionine methionine þ 151.9834 C3H6NO2S2 cystine cystine þ 156.0768 C6H10N3O2 histidine histidine þ 157.0760 C10H9N2 tryptophan tryptophan þ 158.0839 C10H10N2 tryptophan tryptophan þ 160.1081 C6H14N3O2 arginine arginine þ 165.0546 C9H9O3 tyrosine tyrosine þ 165.0764 C9H11NO2 tyrosine tyrosine þ 166.0863 C9H12NO2 phenylalanine phenylalanine þ 169.0917 C10H11N2 tryptophan tryptophan þ 175.1190 C6H15N4O2 arginine arginine þ 182.0812 C9H12NO3 tyrosine tyrosine þ 188.0944 C11H12N2O tryptophan tryptophan þ 195.0256 C5H11N2O2S2 cystine cystine

205.0972 C11H13N2O2þ tryptophan tryptophan þ 207.0256 C6H11N2O2S2 cystine cystine þ 241.0311 C6H13N2O4S2 cystine cystine

Surface Science Spectra, Vol. 25, 2018 Database of proteinogenic amino acid reference spectra 015002-29