#METABOLOMICS WORKBENCH amitch_20160908_112213 DATATRACK_ID:726 STUDY_ID:ST000466 ANALYSIS_ID:AN000728 PROJECT_ID:PR000358
VERSION             	1
CREATED_ON             	September 12, 2016, 1:31 pm
#PROJECT
PR:PROJECT_TITLE                 	Streptococcus mutans Amino Acid/Organic Acid Profile
PR:PROJECT_SUMMARY               	Determination of metabolite profiles of S. mutans strains UA159 (WT) and ΔspxA1
PR:PROJECT_SUMMARY               	in response to oxidative stress
PR:INSTITUTE                     	Univeristy of Florida
PR:DEPARTMENT                    	Oral Biology
PR:LABORATORY                    	Lemos-Abranches
PR:LAST_NAME                     	Lemos
PR:FIRST_NAME                    	Jose
PR:ADDRESS                       	1395 Center Drive, PO Box 100424 Gainesville, FL 32610
PR:EMAIL                         	jlemos@dental.ufl.edu
PR:PHONE                         	352-273-8843
PR:FUNDING_SOURCE                	UF start-up funds
#STUDY
ST:STUDY_TITLE                   	Streptococcus mutans organic acids/amino acid profiles, effect of oxidative
ST:STUDY_TITLE                   	stress; pilot
ST:STUDY_TYPE                    	Amino acid/organic acid profiles of wild-type and mutant strain in response to
ST:STUDY_TYPE                    	H2O2 treatment over time.
ST:STUDY_SUMMARY                 	S. mutans UA159 and ΔspxA1 will be grown in BHI to OD600 = 0.4. Each culture
ST:STUDY_SUMMARY                 	will be split in to 4 samples. For the first sample, 3E10 cells will immediately
ST:STUDY_SUMMARY                 	be harvested by fast filtration, scraped off of membrane into ice-cold PBS, and
ST:STUDY_SUMMARY                 	pelleted by centrifugation. The remaining three samples will be treated with
ST:STUDY_SUMMARY                 	H2O2 to 0.5 mM, and the harvesting process repeated after 15, 30 and 60 minutes.
ST:INSTITUTE                     	University of Florida
ST:DEPARTMENT                    	SECIM
ST:LABORATORY                    	Lemos-Abranches
ST:LAST_NAME                     	Hardin
ST:FIRST_NAME                    	Emily
ST:ADDRESS                       	1395 Center Drive, PO Box 100424, Gainesville, FL 32610
ST:EMAIL                         	ehardin@ufl.edu
ST:PHONE                         	352-273-8844
ST:NUM_GROUPS                    	6
ST:TOTAL_SUBJECTS                	6
#SUBJECT
SU:SUBJECT_TYPE                  	Bacterial cells
SU:SUBJECT_SPECIES               	Streptococcus mutans
SU:TAXONOMY_ID                   	1309
SU:GENOTYPE_STRAIN               	UA159
#SUBJECT_SAMPLE_FACTORS:         	SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data
SUBJECT_SAMPLE_FACTORS           	UA159_0	Lemos_010	Strain:WT | Exposure to H2O2:0.00	
SUBJECT_SAMPLE_FACTORS           	UA159_15min	Lemos_011	Strain:WT | Exposure to H2O2:15 min	
SUBJECT_SAMPLE_FACTORS           	UA159_30 min	Lemos_012	Strain:WT | Exposure to H2O2:30 min	
SUBJECT_SAMPLE_FACTORS           	UA159_60 min	Lemos_013	Strain:WT | Exposure to H2O2:60 min	
SUBJECT_SAMPLE_FACTORS           	spxA1_0	Lemos_014	Strain:ΔspxA1 | Exposure to H2O2:0.00	
SUBJECT_SAMPLE_FACTORS           	spxA1_60 min	Lemos_017	Strain:ΔspxA1 | Exposure to H2O2:60 min	
#COLLECTION
CO:COLLECTION_SUMMARY            	Upon reaching OD600 = 0.4, the culture is split into 4 samples. Three are
CO:COLLECTION_SUMMARY            	treated with H2O2 (0.5 mM). The other is harvested by filtration onto 0.45uM
CO:COLLECTION_SUMMARY            	nitrocellulose membrane. The cells will be scraped off of the membrane into
CO:COLLECTION_SUMMARY            	ice-cold PBS (~1 mL), centrifuged for 30 seconds at 12K RPM, liquid aspirated
CO:COLLECTION_SUMMARY            	and stored at -80C until sending for analysis. This process will be repeated
CO:COLLECTION_SUMMARY            	with the remaining three samples after 15, 30 and 60 minutes treatment with
CO:COLLECTION_SUMMARY            	H2O2.
CO:SAMPLE_TYPE                   	Bacterial cells
#TREATMENT
TR:TREATMENT_SUMMARY             	0.5 mM H2O2 for 15, 30 and 60 minutes. Control has 0 H2O2 added.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	Amino acids|Organic acids
SP:SAMPLEPREP_PROTOCOL_FILENAME  	SB_AA_Assay.pdf;SB_OA_Assay.pdf
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_SUMMARY        	Amino acids
CH:CHROMATOGRAPHY_TYPE           	Reversed phase
CH:INSTRUMENT_NAME               	Agilent 6490
CH:COLUMN_NAME                   	Waters AccQ Tag (100 x 2mm, 1.7um)
CH:METHODS_FILENAME              	SB_AA_Assay.pdf
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:MS_COMMENTS                   	-
MS:INSTRUMENT_NAME               	Agilent 6490 QQQ
MS:INSTRUMENT_TYPE               	Triple quadrupole
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	POSITIVE
#MS_METABOLITE_DATA
MS_METABOLITE_DATA:UNITS         	nmol/mg protein
MS_METABOLITE_DATA_START
Samples	Lemos_010	Lemos_011	Lemos_012	Lemos_013	Lemos_014	Lemos_017
Factors	Strain:WT | Exposure to H2O2:0.00	Strain:WT | Exposure to H2O2:15 min	Strain:WT | Exposure to H2O2:30 min	Strain:WT | Exposure to H2O2:60 min	Strain:ΔspxA1 | Exposure to H2O2:0.00	Strain:ΔspxA1 | Exposure to H2O2:60 min
Alanine	13.4968173	13.77484535	17.08474808	11.44786287	29.57921822	27.80150967
Arginine	0.727122952	0.93160068	1.487828796	1.178842015	13.91608989	16.93600365
Asparagine	1.279478236	1.628887762	2.069656943	2.224827051	3.26275759	7.785364418
Aspartic Acid	4.455220825	6.984646068	9.881306129	5.568899683	20.26276077	19.34355353
Citrulline	6.310064266	8.581588219	8.964833713	6.238985801	0.176973847	0.131098646
Glutamic Acid	17.10642159	24.37329968	21.11184161	15.2096646	92.45672307	49.25100429
Glutamine	2.531073596	7.180627198	5.92838685	3.511570294	6.258616833	32.17249124
Glycine	4.404880679	3.490639013	6.156741627	3.869487452	7.282333173	7.485359965
Histidine	0.586065066	0.637094911	0.833566217	0.640196052	1.007666776	1.454040575
Isoleucine	1.392103942	1.917462191	2.249132637	1.519424174	2.00070069	3.156461297
Leucine	4.119419121	7.778756768	6.827787797	4.803071231	10.34482415	12.07748425
Lysine	8.086046824	12.85291899	16.39849601	8.784116838	24.36911396	28.16432389
Methionine	0.964918644	1.249392478	1.421942208	0.976152554	1.442880753	2.198434199
Ornithine	3.072268435	3.612224813	4.246898133	2.245134481	0.368238481	0.451957404
Phenylalanine	1.52883581	1.889360051	2.492716603	1.422059683	2.041916121	2.798384215
Proline	1.450537447	1.050247783	1.117216511	0.671405546	2.036416928	1.908432913
Serine	1.201237763	2.257184921	2.551346905	1.775937567	2.378835817	3.716613361
Threonine	2.707296961	3.475857255	4.598548885	2.550898527	8.081660248	7.673625587
Tryptophan	0.378391222	0.390840699	0.464721524	0.353727692	0.471037662	0.60111904
Tyrosine	0.644789168	0.693982476	0.965012788	0.615679833	0.961151331	0.966145347
Valine	2.021256555	3.032133437	3.323429924	1.812314893	1.869078353	1.427014083
MS_METABOLITE_DATA_END
#METABOLITES
METABOLITES_START
metabolite_name	retention index	quantified m/z	PubChem ID	KEGG ID
Alanine	4.16	260.17	5950
Arginine	2.68	345.21	6322
Asparagine	2.25	303.13	6267
Aspartic Acid	2.99	304.11	5960
Citrulline	3.33	346.21	9750
Glutamic Acid	3.35	318.11	33032
Glutamine	2.65	317.21	5961
Glycine	2.81	246.08	750
Histidine	2.07	326.21	6274
Isoleucine	8.15	302.21	6306
Leucine	8.26	302.21	6106
Lysine	6.66	487.21	5962
Methionine	6.87	320.21	6137
Ornithine	5.93	473.22	6262
Phenylalanine	8.40	336.21	6140
Proline	4.84	286.16	145742
Serine	2.60	276.11	5951
Threonine	3.74	290.11	6288
Tryptophan	8.54	375.21	6305
Tyrosine	6.66	352.21	6057
Valine	7.09	288.21	6287
METABOLITES_END
#END