Summary of Study ST003103

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001926. The data can be accessed directly via it's Project DOI: 10.21228/M8RT5W This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

Study ID	ST003103
Study Title	Reinforcing the Evidence of Mitochondrial Dysfunction in Long COVID Patients using a Multiplatform Mass Spectrometry-based Metabolomics Approach
Study Summary	Despite the recent and increasing knowledge surrounding COVID-19 infection, the underlying mechanisms of the persistence of symptoms long time after the acute infection are still not completely understood. Here, a multiplatform mass spectrometry-based approach was used for metabolomic and lipidomic profiling of human plasma samples from Long COVID patients (n=40) to reveal mitochondrial dysfunction when compared with individuals fully recovered from acute mild COVID-19 (n=40). Untargeted metabolomic analysis using CE-ESI(+/–)-TOF-MS and GC-Q-MS was performed. Additionally, a lipidomic analysis using LC-ESI(+/–)-QTOF-MS based on an in-house library revealed 471 lipid species identified with high confidence annotation level. The integration of complementary analytical platforms has allowed a comprehensive metabolic and lipidomic characterization of plasma alterations in Long COVID disease that found 46 relevant metabolites which allowed to discriminate between Long COVID and fully recovered patients. We report specific metabolites altered in Long COVID, mainly related to a decrease in the amino acid metabolism and ceramide plasma levels, and an increase in the tricarboxylic acid (TCA) cycle, reinforcing the evidence of an impaired mitochondrial function. The most relevant alterations shown in this study will help to better understand the insights of Long COVID syndrome by providing a deeper knowledge of the metabolomic basis of the pathology.
Institute	Universidad CEU San Pablo
Department	Chemistry and Biochemistry
Laboratory	CEMBIO
Last Name	Martinez
First Name	Sara
Address	Urbanización Montepríncipe, 28660, Boadilla del Monte, Madrid, Spain
Email	sara.martinezlopez@ceu.es
Phone	(+34)913724769
Submit Date	2024-02-15
Num Groups	2
Total Subjects	80
Num Males	14
Num Females	66
Raw Data Available	Yes
Raw Data File Type(s)	mzML
Analysis Type Detail	GC/LC-MS
Release Date	2024-03-25
Release Version	1

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Project:

Project ID:	PR001926
Project DOI:	doi: 10.21228/M8RT5W
Project Title:	Reinforcing the Evidence of Mitochondrial Dysfunction in Long COVID Patients using a Multiplatform Mass Spectrometry-based Metabolomics Approach
Project Summary:	Despite the recent and increasing knowledge surrounding COVID-19 infection, the underlying mechanisms of the persistence of symptoms long time after the acute infection are still not completely understood. Here, a multiplatform mass spectrometry-based approach was used for metabolomic and lipidomic profiling of human plasma samples from Long COVID patients (n=40) to reveal mitochondrial dysfunction when compared with individuals fully recovered from acute mild COVID-19 (n=40). Untargeted metabolomic analysis using CE-ESI(+/–)-TOF-MS and GC-Q-MS was performed. Additionally, a lipidomic analysis using LC-ESI(+/–)-QTOF-MS based on an in-house library revealed 471 lipid species identified with high confidence annotation level. The integration of complementary analytical platforms has allowed a comprehensive metabolic and lipidomic characterization of plasma alterations in Long COVID disease that found 46 relevant metabolites which allowed to discriminate between Long COVID and fully recovered patients. We report specific metabolites altered in Long COVID, mainly related to a decrease in the amino acid metabolism and ceramide plasma levels, and an increase in the tricarboxylic acid (TCA) cycle, reinforcing the evidence of an impaired mitochondrial function. The most relevant alterations shown in this study will help to better understand the insights of Long COVID syndrome by providing a deeper knowledge of the metabolomic basis of the pathology.
Institute:	Universidad CEU San Pablo
Department:	Chemistry and Biochemistry
Laboratory:	CEMBIO
Last Name:	Martinez
First Name:	Sara
Address:	Urbanización Montepríncipe, Boadilla del Monte, Madrid, 28660, Spain
Email:	sara.martinezlopez@ceu.es
Phone:	(+34)913724769

Subject:

Subject ID:	SU003218
Subject Type:	Human
Subject Species:	Homo sapiens
Taxonomy ID:	9606

Factors:

Subject type: Human; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id	local_sample_id	Sample source	Factor
SA333503	L52	Plasma	Long COVID
SA333504	L51	Plasma	Long COVID
SA333505	L50	Plasma	Long COVID
SA333506	L54	Plasma	Long COVID
SA333507	L56	Plasma	Long COVID
SA333508	L58	Plasma	Long COVID
SA333509	L57	Plasma	Long COVID
SA333510	L49	Plasma	Long COVID
SA333511	L55	Plasma	Long COVID
SA333512	L47	Plasma	Long COVID
SA333513	L103	Plasma	Long COVID
SA333514	L102	Plasma	Long COVID
SA333515	L101	Plasma	Long COVID
SA333516	L42	Plasma	Long COVID
SA333517	L43	Plasma	Long COVID
SA333518	L59	Plasma	Long COVID
SA333519	L46	Plasma	Long COVID
SA333520	L45	Plasma	Long COVID
SA333521	L48	Plasma	Long COVID
SA333522	L61	Plasma	Long COVID
SA333523	L74	Plasma	Long COVID
SA333524	L73	Plasma	Long COVID
SA333525	L72	Plasma	Long COVID
SA333526	L75	Plasma	Long COVID
SA333527	L76	Plasma	Long COVID
SA333528	L79	Plasma	Long COVID
SA333529	L78	Plasma	Long COVID
SA333530	L77	Plasma	Long COVID
SA333531	L70	Plasma	Long COVID
SA333532	L69	Plasma	Long COVID
SA333533	L63	Plasma	Long COVID
SA333534	L62	Plasma	Long COVID
SA333535	L100	Plasma	Long COVID
SA333536	L64	Plasma	Long COVID
SA333537	L65	Plasma	Long COVID
SA333538	L68	Plasma	Long COVID
SA333539	L67	Plasma	Long COVID
SA333540	L66	Plasma	Long COVID
SA333541	L60	Plasma	Long COVID
SA333542	L44	Plasma	Long COVID
SA333543	QC2	Plasma	Quality control
SA333544	QC3	Plasma	Quality control
SA333545	QC12	Plasma	Quality control
SA333546	QC11	Plasma	Quality control
SA333547	QC1	Plasma	Quality control
SA333548	QC4	Plasma	Quality control
SA333549	QC10	Plasma	Quality control
SA333550	QC5	Plasma	Quality control
SA333551	QC9	Plasma	Quality control
SA333552	QC8	Plasma	Quality control
SA333553	QC6	Plasma	Quality control
SA333554	QC7	Plasma	Quality control
SA333555	C18	Plasma	Recovered from acute COVID-19
SA333556	C17	Plasma	Recovered from acute COVID-19
SA333557	C19	Plasma	Recovered from acute COVID-19
SA333558	C21	Plasma	Recovered from acute COVID-19
SA333559	C16	Plasma	Recovered from acute COVID-19
SA333560	C20	Plasma	Recovered from acute COVID-19
SA333561	C2	Plasma	Recovered from acute COVID-19
SA333562	C13	Plasma	Recovered from acute COVID-19
SA333563	C10	Plasma	Recovered from acute COVID-19
SA333564	C22	Plasma	Recovered from acute COVID-19
SA333565	C11	Plasma	Recovered from acute COVID-19
SA333566	C12	Plasma	Recovered from acute COVID-19
SA333567	C14	Plasma	Recovered from acute COVID-19
SA333568	C15	Plasma	Recovered from acute COVID-19
SA333569	C23	Plasma	Recovered from acute COVID-19
SA333570	C52	Plasma	Recovered from acute COVID-19
SA333571	C55	Plasma	Recovered from acute COVID-19
SA333572	C49	Plasma	Recovered from acute COVID-19
SA333573	C43	Plasma	Recovered from acute COVID-19
SA333574	C42	Plasma	Recovered from acute COVID-19
SA333575	C59	Plasma	Recovered from acute COVID-19
SA333576	C1	Plasma	Recovered from acute COVID-19
SA333577	C8	Plasma	Recovered from acute COVID-19
SA333578	C7	Plasma	Recovered from acute COVID-19
SA333579	C68	Plasma	Recovered from acute COVID-19
SA333580	C61	Plasma	Recovered from acute COVID-19
SA333581	C41	Plasma	Recovered from acute COVID-19
SA333582	C4	Plasma	Recovered from acute COVID-19
SA333583	C27	Plasma	Recovered from acute COVID-19
SA333584	C29	Plasma	Recovered from acute COVID-19
SA333585	C26	Plasma	Recovered from acute COVID-19
SA333586	C25	Plasma	Recovered from acute COVID-19
SA333587	C24	Plasma	Recovered from acute COVID-19
SA333588	C31	Plasma	Recovered from acute COVID-19
SA333589	C32	Plasma	Recovered from acute COVID-19
SA333590	C37	Plasma	Recovered from acute COVID-19
SA333591	C35	Plasma	Recovered from acute COVID-19
SA333592	C34	Plasma	Recovered from acute COVID-19
SA333593	C33	Plasma	Recovered from acute COVID-19
SA333594	C9	Plasma	Recovered from acute COVID-19

Showing results 1 to 92 of 92

Showing results 1 to 92 of 92

Collection:

Collection ID:	CO003211
Collection Summary:	Samples were collected after fasting conditions. 1,500 μL of cold MeOH/EtOH (1:1, v/v) were added to 500 μL of plasma for virus inactivation. Afterward, samples were vigorously mixed using vortex for 1 min, followed by incubation on ice for 5 min, and subsequent centrifugation at 16,000 x g for 20 min at 4 °C to eliminate proteins by precipitation. The resulting extract, containing the metabolites of interest, was transferred to EppendorfTM tubes, and stored at –80 °C until analysis.
Sample Type:	Blood (plasma)
Storage Conditions:	-80℃

Treatment:

Treatment ID:	TR003227
Treatment Summary:	N/A

Sample Preparation:

Sampleprep ID:	SP003224
Sampleprep Summary:	For CE-MS analysis 200 µL of frozen plasma extract (MeOH/EtOH 1:1, v/v) were thawed on ice and dried completely using a SpeedVac Concentrator System. The resulting residue was then reconstituted in 100 µL of 0.2 mM MetS dissolved in 0.1 M FA. After vortex-mixing for 1 min, the samples were transferred to a Millipore filter with a 30 kDa protein cutoff and centrifuged at 2,000 x g for 40 min at 4 °C. Finally, the resulting ultrafiltrate was transferred to a CE-MS vial for further analysis. Prior to the analysis vials were centrifuged at 2,000 x g for 10 min at 4 °C to ensure that any possible sediment remained at the bottom of the vial. For GC-MS analysis 200 µL of each plasma extract was thawed on ice to room temperature and 30 µL of the internal standard (IS), palmitic acid-d31 in MeOH (80 mg/mL) was added to it. The mixture was vortexed for 5 min and 200 µL of the solution were transferred to a GC-MS vial. Then, samples were evaporated to dryness using a SpeedVac Concentrator System and maintained at 8 °C in the Gerstel Multiple Purpose Sample (MPS) Preparation Station. An automated two step derivatization process was performed before sample injection using a protocol adjusted from a previous reported method. First, each precipitate was redissolved in 20 µL of O-methoxyamine solution (15 mg/mL in pyridine) for the methoximation process, mixed 10 min at 1,000 rpm and incubated for 90 min at 60 °C at 750 rpm. Second, and after waiting 5 min, 40 µL of BSTFA with 1% TMCS were added for the silylation process. Then, samples were mixed for 10 min at 1,000 rpm and incubated for 60 min at 60 °C at 750 rpm. After waiting 30 min at 8 °C, 80 µL of heptane containing 20 mg/mL of tricosane (IS) were added and mixed for 5 min at 1,000 rpm. Finally, samples were maintained at 8 °C for 30 min before injection. For LC-MS 200 µL of each plasma extract was thawed on ice to room temperature and centrifuged for 10 min at 16,100 x g at 4 °C, transferred to a LC-MS vial and directly injected into the system.

Sampleprep ID:

SP003224

Sampleprep Summary:

For CE-MS analysis 200 µL of frozen plasma extract (MeOH/EtOH 1:1, v/v) were thawed on ice and dried completely using a SpeedVac Concentrator System. The resulting residue was then reconstituted in 100 µL of 0.2 mM MetS dissolved in 0.1 M FA. After vortex-mixing for 1 min, the samples were transferred to a Millipore filter with a 30 kDa protein cutoff and centrifuged at 2,000 x g for 40 min at 4 °C. Finally, the resulting ultrafiltrate was transferred to a CE-MS vial for further analysis. Prior to the analysis vials were centrifuged at 2,000 x g for 10 min at 4 °C to ensure that any possible sediment remained at the bottom of the vial. For GC-MS analysis 200 µL of each plasma extract was thawed on ice to room temperature and 30 µL of the internal standard (IS), palmitic acid-d31 in MeOH (80 mg/mL) was added to it. The mixture was vortexed for 5 min and 200 µL of the solution were transferred to a GC-MS vial. Then, samples were evaporated to dryness using a SpeedVac Concentrator System and maintained at 8 °C in the Gerstel Multiple Purpose Sample (MPS) Preparation Station. An automated two step derivatization process was performed before sample injection using a protocol adjusted from a previous reported method. First, each precipitate was redissolved in 20 µL of O-methoxyamine solution (15 mg/mL in pyridine) for the methoximation process, mixed 10 min at 1,000 rpm and incubated for 90 min at 60 °C at 750 rpm. Second, and after waiting 5 min, 40 µL of BSTFA with 1% TMCS were added for the silylation process. Then, samples were mixed for 10 min at 1,000 rpm and incubated for 60 min at 60 °C at 750 rpm. After waiting 30 min at 8 °C, 80 µL of heptane containing 20 mg/mL of tricosane (IS) were added and mixed for 5 min at 1,000 rpm. Finally, samples were maintained at 8 °C for 30 min before injection. For LC-MS 200 µL of each plasma extract was thawed on ice to room temperature and centrifuged for 10 min at 16,100 x g at 4 °C, transferred to a LC-MS vial and directly injected into the system.

Combined analysis:

Analysis ID	AN005077	AN005078	AN005079	AN005080	AN005081
Analysis type	MS	MS	MS	MS	MS
Chromatography type	Reversed phase	Reversed phase	GC	CE	CE
Chromatography system	Agilent 1290 Infinity II	Agilent 1290 Infinity II	Agilent 8890 GC System	Agilent 7100 CE	Agilent 7100 CE
Column	Agilent InfinityLab Poroshell 120 EC-C18 (100 x 3mm,2.7um)	Agilent InfinityLab Poroshell 120 EC-C18 (100 x 3mm,2.7um)	GC DB5-MS column (40 m length, 0.25 mm inner diameter, and 0.25 µm film of 95% dimethyl/5% diphenylpolysiloxane)	Agilent Technologies fused silica capillary (total length, 100 cm; internal diameter, 50 µm)	Agilent Technologies fused silica capillary (total length, 100 cm; internal diameter, 50 µm)
MS Type	ESI	ESI	EI	ESI	ESI
MS instrument type	QTOF	QTOF	Single quadrupole	TOF	TOF
MS instrument name	Agilent 6545 QTOF	Agilent 6545 QTOF	Agilent 5977B	Agilent 6230 TOF	Agilent 6230 TOF
Ion Mode	POSITIVE	NEGATIVE	UNSPECIFIED	POSITIVE	NEGATIVE
Units	Area	Corrected areas	Corrected areas	Corrected areas	Corrected areas

Chromatography:

Chromatography ID:	CH003834
Chromatography Summary:	RP-UHPLC-ESI(+)-QTOF-MS
Instrument Name:	Agilent 1290 Infinity II
Column Name:	Agilent InfinityLab Poroshell 120 EC-C18 (100 x 3mm,2.7um)
Column Temperature:	50°C
Flow Gradient:	The chromatographic gradient started at 70 % of B at 0 - 1 min, 86 % B at 3.5 - 10 min, 100% B at 11 - 17 min. The starting conditions were recovered by min 17, followed by a 2 min re-equilibration time, reaching a total running time of 19 min
Flow Rate:	0.6 mL/min
Solvent A:	10 mM CH3COONH4 and 0.2 mM NH4F in H2O/MeOH (9:1, v/v)
Solvent B:	10 mM CH3COONH4 and 0.2 mM NH4F in ACN/MeOH/IPA (2:3:5, v/v/v)
Analytical Time:	19 min
Chromatography Type:	Reversed phase

Chromatography ID:	CH003835
Chromatography Summary:	RP-UHPLC-ESI(-)-QTOF-MS
Instrument Name:	Agilent 1290 Infinity II
Column Name:	Agilent InfinityLab Poroshell 120 EC-C18 (100 x 3mm,2.7um)
Column Temperature:	50°C
Flow Gradient:	The chromatographic gradient started at 70 % of B at 0 - 1 min, 86 % B at 3.5 - 10 min, 100% B at 11 - 17 min. The starting conditions were recovered by min 17, followed by a 2 min re-equilibration time, reaching a total running time of 19 min
Flow Rate:	0.6 mL/min
Solvent A:	10 mM CH3COONH4 and 0.2 mM NH4F in H2O/MeOH (9:1, v/v)
Solvent B:	10 mM CH3COONH4 and 0.2 mM NH4F in ACN/MeOH/IPA (2:3:5, v/v/v)
Analytical Time:	19 min
Chromatography Type:	Reversed phase

Chromatography ID:	CH003836
Chromatography Summary:	GC-Q-MS
Instrument Name:	Agilent 8890 GC System
Column Name:	GC DB5-MS column (40 m length, 0.25 mm inner diameter, and 0.25 µm film of 95% dimethyl/5% diphenylpolysiloxane)
Column Temperature:	The column temperature gradient was programmed as follows: it started at 60 °C for 1 min, then increased by 10 °C/min until reaching 325 °C and held at this temperature for 10 min before cooling down
Flow Gradient:	Constant
Flow Rate:	0.5658 mL/min
Solvent A:	Helium
Solvent B:	N/A
Analytical Time:	37 min followed by 5 min of post-run period
Chromatography Type:	GC

Chromatography ID:	CH003837
Chromatography Summary:	CE-ESI(+)-TOF-MS
Instrument Name:	Agilent 7100 CE
Column Name:	Agilent Technologies fused silica capillary (total length, 100 cm; internal diameter, 50 µm)
Column Temperature:	20°C
Flow Gradient:	N/A
Flow Rate:	N/A
Solvent A:	BGE (1 M FA in 10% MeOH)
Solvent B:	N/A
Analytical Time:	26 min
Capillary Voltage:	30kV
Sheath Liquid:	MeOH: H2O (1:1, v/v) with 1mM FA containing two reference masses (purine: 121.0509 and HP-0922: 922.0098) at a flow rate of 0.6 mL/min (1:100 of split ratio)
Chromatography Type:	CE

Chromatography ID:	CH003838
Chromatography Summary:	CE-ESI(-)-TOF-MS
Instrument Name:	Agilent 7100 CE
Column Name:	Agilent Technologies fused silica capillary (total length, 100 cm; internal diameter, 50 µm)
Column Temperature:	20°C
Flow Gradient:	N/A
Flow Rate:	N/A
Solvent A:	BGE (1 M FA in 10% MeOH)
Solvent B:	N/A
Analytical Time:	40 min
Capillary Voltage:	-30kV
Sheath Liquid:	MeOH: H2O (1:1, v/v) with 1mM FA containing two reference masses (purine: 121.0509 and HP-0922: 922.0098) at a flow rate of 1.0 mL/min (1:100 of split ratio)
Chromatography Type:	CE

MS:

MS ID:	MS004815
Analysis ID:	AN005077
Instrument Name:	Agilent 6545 QTOF
Instrument Type:	QTOF
MS Type:	ESI
MS Comments:	The mass spectrometer operated in full scan mode, scanning from m/z 40 - 1700 at a scan rate of 3 spectra/s. During the analysis, a solution containing two reference mass compounds was continuously infused to the system at a flow rate of 1 mL/min to provide mass correction. The reference masses used were m/z 121.0509 (purine detected as [C5H4N4 + H]+) and m/z 922.0098 (HP-0921 detected as [C18H18O6N3P3F24 + H]+) for ESI(+) and m/z 119.0363 (purine detected as [C5H4N4 - H]-) and m/z 1033.9881 (HP-0921 detected as [C18H18O6N3P3F24 + CF3COOH-H]-) for ESI(–). At the end of the analysis, ten iterative-MS/MS runs were performed for both, positive and negative ionization modes using a QC sample. They were operated with a MS and MS/MS scan rates of 3 spectra/s, 3 precursors per cycle, a mass range of m/z 40 - 1700, a narrow (~ 1.3 amu) MS/MS isolation width, and 5000 counts and 0.001 % of MS/MS threshold. The collision energy for the first five iterative-MS/MS runs was set at 20 eV, and the subsequent five runs were performed at 40 eV. To ensure accuracy, reference masses and contaminants detected in blank samples were excluded from the analysis. This prevented thein inclusion in the iterative-MS/MS runs. Data was acquired using MassHunter Workstation Software LC-MS Data Acquisition v B.09.00 (Agilent Technologies, Waldbronn, Germany).
Ion Mode:	POSITIVE

MS ID:	MS004816
Analysis ID:	AN005078
Instrument Name:	Agilent 6545 QTOF
Instrument Type:	QTOF
MS Type:	ESI
MS Comments:	The mass spectrometer operated in full scan mode, scanning from m/z 40 - 1700 at a scan rate of 3 spectra/s. During the analysis, a solution containing two reference mass compounds was continuously infused to the system at a flow rate of 1 mL/min to provide mass correction. The reference masses used were m/z 121.0509 (purine detected as [C5H4N4 + H]+) and m/z 922.0098 (HP-0921 detected as [C18H18O6N3P3F24 + H]+) for ESI(+) and m/z 119.0363 (purine detected as [C5H4N4 - H]-) and m/z 1033.9881 (HP-0921 detected as [C18H18O6N3P3F24 + CF3COOH-H]-) for ESI(–). At the end of the analysis, ten iterative-MS/MS runs were performed for both, positive and negative ionization modes using a QC sample. They were operated with a MS and MS/MS scan rates of 3 spectra/s, 3 precursors per cycle, a mass range of m/z 40 - 1700, a narrow (~ 1.3 amu) MS/MS isolation width, and 5000 counts and 0.001 % of MS/MS threshold. The collision energy for the first five iterative-MS/MS runs was set at 20 eV, and the subsequent five runs were performed at 40 eV. To ensure accuracy, reference masses and contaminants detected in blank samples were excluded from the analysis. This prevented thein inclusion in the iterative-MS/MS runs. Data was acquired using MassHunter Workstation Software LC-MS Data Acquisition v B.09.00 (Agilent Technologies, Waldbronn, Germany).
Ion Mode:	NEGATIVE

MS ID:	MS004817
Analysis ID:	AN005079
Instrument Name:	Agilent 5977B
Instrument Type:	Single quadrupole
MS Type:	EI
MS Comments:	For ionization, the EI source was used with the following settings: filament source temperature at 230 °C and electron ionization energy at 70 eV. Mass spectra were collected in a mass range of m/z 50 - 600 at a scan rate of 2 spectra/s. Mass calibration was performed after every injection with internal reference masses m/z 68.9947, 263.9866 and 501.9706. Data acquisition was performed using the Agilent MassHunter Workstation GC-MS Data Acquisition v 10.0 (Agilent Technologies, Waldbronn, Germany).
Ion Mode:	UNSPECIFIED

MS ID:	MS004818
Analysis ID:	AN005080
Instrument Name:	Agilent 6230 TOF
Instrument Type:	TOF
MS Type:	ESI
MS Comments:	The mass spectra data were acquired in positive polarity mode with a full scan range of m/z 70 - 1000 at a rate of 1.36 spectra/s. The following MS parameters were employed: fragmentor set to 125 V, skimmer to 65 V, OCT RF Vpp to 750 V, drying gas temperature to 200 °C ESI(+), flow rate to 10 L/min, nebulizer to 10 psi, and capillary voltage to 3500 V ESI(+). The Agilent MassHunter Workstation v B.09.00 (Agilent Technologies, Waldbronn, Germany) was used for CE-MS data acquisition.
Ion Mode:	POSITIVE

MS ID:	MS004819
Analysis ID:	AN005081
Instrument Name:	Agilent 6230 TOF
Instrument Type:	TOF
MS Type:	ESI
MS Comments:	The mass spectra data were acquired in negative polarity mode with a full scan range m/z 60 - 1000 at a rate of 1 spectra/s. The following MS parameters were employed: fragmentor set to 125 V, skimmer to 65 V, OCT RF Vpp to 750 V, drying gas temperature to 275 °C ESI(–), flow rate to 10 L/min, nebulizer to 10 psi, and capillary voltage to 2000 V ESI(-). The Agilent MassHunter Workstation v B.09.00 (Agilent Technologies, Waldbronn, Germany) was used for CE-MS data acquisition.
Ion Mode:	NEGATIVE