#METABOLOMICS WORKBENCH SIMR_Core_Facility_20221214_221037 DATATRACK_ID:3648 STUDY_ID:ST002415 ANALYSIS_ID:AN003936 PROJECT_ID:PR001555 VERSION 1 CREATED_ON December 20, 2022, 11:22 am #PROJECT PR:PROJECT_TITLE Proteomics and metabolomics of multiple sclerosis PR:PROJECT_SUMMARY Multiple sclerosis (MS) is a chronic autoimmune disease that affects the PR:PROJECT_SUMMARY myelination of the neurons present in the central nervous system (CNS). The PR:PROJECT_SUMMARY exact etiology of MS development is unclear, but various environmental and PR:PROJECT_SUMMARY genetic factors might play a role in initiating the disease. Current treatments PR:PROJECT_SUMMARY for MS enhance the quality of life and reduce the symptoms. One of these PR:PROJECT_SUMMARY treatments is dimethyl fumarate (DMF), commercially known as Tecfidera. PR:PROJECT_SUMMARY Experimental autoimmune encephalomyelitis (EAE) is a mouse model that is used to PR:PROJECT_SUMMARY study the pathophysiology of MS disease as well as the effects of possible PR:PROJECT_SUMMARY therapeutic agents. In this study, we investigated the effects of SIMR1707 which PR:PROJECT_SUMMARY is a novel compound designed at Sharjah Institute for Medical Research (SIMR). . PR:PROJECT_SUMMARY Single and multiple doses of SIMR1707 demonstrated high safety in mice studies. PR:PROJECT_SUMMARY Treatment of EAE mice with SIMR1707 was able to reduce the EAE clinical scores PR:PROJECT_SUMMARY and maintain their body weight similar to the MS FDA-approved (DMF, Tecfidera), PR:PROJECT_SUMMARY when they were used preventively, prophylactically, or therapeutically. The PR:PROJECT_SUMMARY histological and immunohistochemistry evaluations showed reduced clinical PR:PROJECT_SUMMARY features such as signs of inflammation, demyelination, and infiltration of PR:PROJECT_SUMMARY CD3-positive T cells into the brains of the EAE mice, as compared to PR:PROJECT_SUMMARY vehicle-treated, or untreated EAE mice. Moreover, multi-OMICS experiments PR:PROJECT_SUMMARY including Transcriptomics, Proteomics and Metabolomics were performed to gain PR:PROJECT_SUMMARY insights into the relevant mechanism of action of the SIMR1707 in EAE and thus PR:PROJECT_SUMMARY its therapeutic efficacy to treat MS. Same tissue samples extracted from the PR:PROJECT_SUMMARY cerebellum part of the brain of normal, EAE vehicle-treated, and therapeutic PR:PROJECT_SUMMARY SIMR1707 treated mice, were subjected for the whole RNA-sequencing for PR:PROJECT_SUMMARY transcriptomics, Nano MS for proteomics analysis and LC-MS metabolomics PR:PROJECT_SUMMARY analysis. The multi-OMICs integrative analysis showed that the treatment with PR:PROJECT_SUMMARY SIMR1707 downregulated key biomarkers functionally associated with top pathways PR:PROJECT_SUMMARY including calcium signaling, PI3K/AKT, and mTOR signaling pathways, which may PR:PROJECT_SUMMARY play important roles in EAE and MS pathophysiology. Additionally, the PR:PROJECT_SUMMARY metabolomics-based enriched-for-action pathway analysis showed that the top PR:PROJECT_SUMMARY significantly activated metabolites (FC > 2, p < 0.05) are cholic acid, PR:PROJECT_SUMMARY propionic acid, sphinganine, and nutriacholic acid. Consisting with the PR:PROJECT_SUMMARY functional enrichment pathway analysis, two potent markers, Snta1 and Fscn1, PR:PROJECT_SUMMARY involved in the actin-binding and cytoskeleton are commonly shared between PR:PROJECT_SUMMARY transcriptomics and proteomics and showed mRNA-protein expression correlation in PR:PROJECT_SUMMARY SIMR1707 treated compared to vehicle EAE mice. Importantly, these two markers PR:PROJECT_SUMMARY are involved in the MT2/AKT/GSK3 pathway and may potentially play role in MS and PR:PROJECT_SUMMARY EAE disease PR:INSTITUTE Sharjah Institute for Medical Research PR:LAST_NAME Facility PR:FIRST_NAME Core PR:ADDRESS M32, SIMR, College of Pharmacy, Health Sciences, University of Sharjah PR:EMAIL tims-tof@sharjah.ac.ae PR:PHONE 065057656 #STUDY ST:STUDY_TITLE Proteomics and metabolomics of multiple sclerosis ST:STUDY_SUMMARY Multiple sclerosis (MS) is a chronic autoimmune disease that affects the ST:STUDY_SUMMARY myelination of the neurons present in the central nervous system (CNS). The ST:STUDY_SUMMARY exact etiology of MS development is unclear, but various environmental and ST:STUDY_SUMMARY genetic factors might play a role in initiating the disease. Current treatments ST:STUDY_SUMMARY for MS enhance the quality of life and reduce the symptoms. One of these ST:STUDY_SUMMARY treatments is dimethyl fumarate (DMF), commercially known as Tecfidera. ST:STUDY_SUMMARY Experimental autoimmune encephalomyelitis (EAE) is a mouse model that is used to ST:STUDY_SUMMARY study the pathophysiology of MS disease as well as the effects of possible ST:STUDY_SUMMARY therapeutic agents. In this study, we investigated the effects of SIMR1707 which ST:STUDY_SUMMARY is a novel compound designed at Sharjah Institute for Medical Research (SIMR). . ST:STUDY_SUMMARY Single and multiple doses of SIMR1707 demonstrated high safety in mice studies. ST:STUDY_SUMMARY Treatment of EAE mice with SIMR1707 was able to reduce the EAE clinical scores ST:STUDY_SUMMARY and maintain their body weight similar to the MS FDA-approved (DMF, Tecfidera), ST:STUDY_SUMMARY when they were used preventively, prophylactically, or therapeutically. The ST:STUDY_SUMMARY histological and immunohistochemistry evaluations showed reduced clinical ST:STUDY_SUMMARY features such as signs of inflammation, demyelination, and infiltration of ST:STUDY_SUMMARY CD3-positive T cells into the brains of the EAE mice, as compared to ST:STUDY_SUMMARY vehicle-treated, or untreated EAE mice. Moreover, multi-OMICS experiments ST:STUDY_SUMMARY including Transcriptomics, Proteomics and Metabolomics were performed to gain ST:STUDY_SUMMARY insights into the relevant mechanism of action of the SIMR1707 in EAE and thus ST:STUDY_SUMMARY its therapeutic efficacy to treat MS. Same tissue samples extracted from the ST:STUDY_SUMMARY cerebellum part of the brain of normal, EAE vehicle-treated, and therapeutic ST:STUDY_SUMMARY SIMR1707 treated mice, were subjected for the whole RNA-sequencing for ST:STUDY_SUMMARY transcriptomics, Nano MS for proteomics analysis and LC-MS metabolomics ST:STUDY_SUMMARY analysis. The multi-OMICs integrative analysis showed that the treatment with ST:STUDY_SUMMARY SIMR1707 downregulated key biomarkers functionally associated with top pathways ST:STUDY_SUMMARY including calcium signaling, PI3K/AKT, and mTOR signaling pathways, which may ST:STUDY_SUMMARY play important roles in EAE and MS pathophysiology. Additionally, the ST:STUDY_SUMMARY metabolomics-based enriched-for-action pathway analysis showed that the top ST:STUDY_SUMMARY significantly activated metabolites (FC > 2, p < 0.05) are cholic acid, ST:STUDY_SUMMARY propionic acid, sphinganine, and nutriacholic acid. Consisting with the ST:STUDY_SUMMARY functional enrichment pathway analysis, two potent markers, Snta1 and Fscn1, ST:STUDY_SUMMARY involved in the actin-binding and cytoskeleton are commonly shared between ST:STUDY_SUMMARY transcriptomics and proteomics and showed mRNA-protein expression correlation in ST:STUDY_SUMMARY SIMR1707 treated compared to vehicle EAE mice. Importantly, these two markers ST:STUDY_SUMMARY are involved in the MT2/AKT/GSK3 pathway and may potentially play role in MS and ST:STUDY_SUMMARY EAE disease ST:INSTITUTE Sharjah Institute for Medical Research ST:LAST_NAME Facility ST:FIRST_NAME Core ST:ADDRESS M32, SIMR, College of Pharmacy, Health Sciences, University of Sharjah ST:EMAIL tims-tof@sharjah.ac.ae ST:PHONE 065057656 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS - 1TH1707 M1-01_10_1_3408 Group:Therapeutic RAW_FILE_NAME=TH1707 M1-01_10_1_3408.d SUBJECT_SAMPLE_FACTORS - TH1707 M1-02_10_1_3409 Group:Therapeutic RAW_FILE_NAME=TH1707 M1-02_10_1_3409.d SUBJECT_SAMPLE_FACTORS - TH1707 M2-01_11_1_3410 Group:Therapeutic RAW_FILE_NAME=TH1707 M2-01_11_1_3410.d SUBJECT_SAMPLE_FACTORS - TH1707 M2-02_11_1_3411 Group:Therapeutic RAW_FILE_NAME=TH1707 M2-02_11_1_3411.d SUBJECT_SAMPLE_FACTORS - TH1707 M3-01_12_1_3412 Group:Therapeutic RAW_FILE_NAME=TH1707 M3-01_12_1_3412.d SUBJECT_SAMPLE_FACTORS - TH1707 M3-02_12_1_3413 Group:Therapeutic RAW_FILE_NAME=TH1707 M3-02_12_1_3413.d SUBJECT_SAMPLE_FACTORS - TH1707 M4-01_13_1_3414 Group:Therapeutic RAW_FILE_NAME=TH1707 M4-01_13_1_3414.d SUBJECT_SAMPLE_FACTORS - TH1707 M4-02_13_1_3415 Group:Therapeutic RAW_FILE_NAME=TH1707 M4-02_13_1_3415.d SUBJECT_SAMPLE_FACTORS - VehicleEAE M1-01_7_1_3402 Group:Vehicle RAW_FILE_NAME=VehicleEAE M1-01_7_1_3402.d SUBJECT_SAMPLE_FACTORS - VehicleEAE M1-02_7_1_3403 Group:Vehicle RAW_FILE_NAME=VehicleEAE M1-02_7_1_3403.d SUBJECT_SAMPLE_FACTORS - VehicleEAE M2-01_8_1_3404 Group:Vehicle RAW_FILE_NAME=VehicleEAE M2-01_8_1_3404.d SUBJECT_SAMPLE_FACTORS - VehicleEAE M2-02_8_1_3405 Group:Vehicle RAW_FILE_NAME=VehicleEAE M2-02_8_1_3405.d SUBJECT_SAMPLE_FACTORS - VehicleEAE M3-01_9_1_3406 Group:Vehicle RAW_FILE_NAME=VehicleEAE M3-01_9_1_3406.d SUBJECT_SAMPLE_FACTORS - VehicleEAE M3-02_9_1_3407 Group:Vehicle RAW_FILE_NAME=VehicleEAE M3-02_9_1_3407.d #COLLECTION CO:COLLECTION_SUMMARY The experiments on the mice were approved by the University of Sharjah animal CO:COLLECTION_SUMMARY care and use committee (ACUC) (ACUC approval number: ACUC-16-003-EAE and CO:COLLECTION_SUMMARY ACUC-21-09-06-01). The mice were housed at the University of Sharjah animal CO:COLLECTION_SUMMARY facility, in plastic cages under a controlled environment of 24 ± 2 °C, 50% ± CO:COLLECTION_SUMMARY 5% humidity, 12 h/12 h light-dark cycle with unlimited food and water access. CO:SAMPLE_TYPE Brain #TREATMENT TR:TREATMENT_SUMMARY From Normal, EAE vehicle-treated, and therapeutic SIMR1707 treated mice, the TR:TREATMENT_SUMMARY cerebellum part of the brain was cut, and immediately snap frozen using liquid TR:TREATMENT_SUMMARY nitrogen. The samples then were weighed, made in powder form and homogenized TR:TREATMENT_SUMMARY using pistol method under liquid nitrogen. The cerebellum sample was lysed using TR:TREATMENT_SUMMARY 400 uL RIBA lysis buffer and let to set for 5-10 minutes to allow lysis. 50 ul TR:TREATMENT_SUMMARY of the homogenized sample was taken for RNA extraction using norgen biotek TR:TREATMENT_SUMMARY RNA/DNA purification kit (cat#:47700) according to the manufacturer’s TR:TREATMENT_SUMMARY instructions. The remaining 350 uL was processed for metabolites and proteins TR:TREATMENT_SUMMARY extraction. First, it was centrifuged for 5 minutes at 4 C, 13000 rpm. The TR:TREATMENT_SUMMARY supernatant was collected in a new 1.5 mL eppendorf tube. The supernatant was TR:TREATMENT_SUMMARY centrifuged again for 1 minute at 4 C, at maximum speed (>13000 rpm), and the TR:TREATMENT_SUMMARY supernatant was collected again in a new 1.5 mL tube and kept on ice as it was TR:TREATMENT_SUMMARY used in the following steps, while the pellet was discarded. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY 400 uL of methanol, and 300 uL of chloroform were added to the protein sample, SP:SAMPLEPREP_SUMMARY and centrifuged for 5 minutes, at 4 C, 13000 rpm. Three phases appeared: the SP:SAMPLEPREP_SUMMARY upper phase is the metabolites, the middle is the protein, and the lower phase SP:SAMPLEPREP_SUMMARY is other metabolites. The metabolites' upper phase was kept in a new eppendorf SP:SAMPLEPREP_SUMMARY tube and kept at -20 C. 300 uL methanol was added to the middle and the lower SP:SAMPLEPREP_SUMMARY phases then they were mixed and centrifuged for 1 minute at 13000 rpm. The SP:SAMPLEPREP_SUMMARY supernatant was added to the metabolite tube and then it was kept at - 80C for SP:SAMPLEPREP_SUMMARY longer storage. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Bruker Elute CH:COLUMN_NAME Hamilton Intensity Solo 2 C18(100 x 2.1 mm, 1.8 um) CH:SOLVENT_A 100% water; 0.1% formic acid CH:SOLVENT_B 100% acetonitrile; 0.1% formic acid CH:FLOW_GRADIENT 1%B to 99%B in 15 min CH:FLOW_RATE 250 uL/min CH:COLUMN_TEMPERATURE 35 #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Bruker timsTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS Metabolomics Data Processing. For metabolomic analysis, MetaboScape® 4.0 MS:MS_COMMENTS program (Bruker Daltonics, Billerica, MA, USA) was employed for data processing, MS:MS_COMMENTS feature extrac-tion and metabolite identification. The T-ReX 2D/3D workflow was MS:MS_COMMENTS used to identify the molecular features with the following settings: The minimum MS:MS_COMMENTS peak length was set to 7 spectra and the minimum intensity threshold was 1000 MS:MS_COMMENTS counts for peaks detection. The peak area was employed for quantification and MS:MS_COMMENTS the injected external calibrant in the in-terval of 0–0.3 min was used to MS:MS_COMMENTS recalibrate the mass spectra. The selected mass to charge ratio (m/z) and MS:MS_COMMENTS retention time for scanning were in the ranges of 20–1300 m/z and 0.3–30 MS:MS_COMMENTS min, respectively. MS/MS spectra for features were averaged on import and MS:MS_COMMENTS features found at least in 12 of the 40 injections were taken into further MS:MS_COMMENTS consideration. Metabolites were identified by matching to the human metabolome MS:MS_COMMENTS database (HMDB) by combined MS/MS, precursor m/z values, and isotopic pattern MS:MS_COMMENTS scores. Where multiple features matched a given database entry, the annotation MS:MS_COMMENTS quality score (AQ score) was used to select only the best matching feature. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS AU MS_METABOLITE_DATA_START Samples TH1707 M1-02_10_1_3409 TH1707 M2-01_11_1_3410 TH1707 M2-02_11_1_3411 TH1707 M3-01_12_1_3412 TH1707 M3-02_12_1_3413 TH1707 M4-01_13_1_3414 TH1707 M4-02_13_1_3415 VehicleEAE M1-01_7_1_3402 VehicleEAE M1-02_7_1_3403 VehicleEAE M2-01_8_1_3404 VehicleEAE M2-02_8_1_3405 VehicleEAE M3-01_9_1_3406 VehicleEAE M3-02_9_1_3407 Factors Group:Therapeutic Group:Therapeutic Group:Therapeutic Group:Therapeutic Group:Therapeutic Group:Therapeutic Group:Therapeutic Group:Vehicle Group:Vehicle Group:Vehicle Group:Vehicle Group:Vehicle Group:Vehicle 2,2-Dimethylsuccinic acid 9280 7100 7322 7486 7304 8512 8144 9486 9026 8682 7268 8886 9216 2-Hydroxycaproic acid 111688 87318 77618 81168 82904 89782 91432 80916 97558 93550 93408 60356 56478 2-Ketobutyric acid 39524 32112 35002 31520 34290 36650 40080 42678 38236 32538 31838 39992 38938 2-Ketohexanoic acid 8510 8146 8652 6092 6090 11126 0 5810 7304 7896 7656 0 0 2-Phenylbutyric acid 53310 54302 53826 42400 48072 50598 44770 52792 51782 51618 51368 1604 1550 2-Pyrrolidinone 2830 3898 3218 4340 3534 2566 2284 2860 3286 3872 4818 6388 6812 3,4-Dihydroxyphenylglycol 16948 16566 18202 15378 14930 16806 17010 13430 14232 16320 13618 13910 14322 3-Aminoisobutanoic acid 44826 39362 36368 31670 31038 46420 41526 53474 54532 45412 41196 34120 43346 3-Hexenedioic acid 3020 1670 2046 2790 2856 4194 3800 3128 2994 3524 3874 3448 3432 3b-Hydroxy-5-cholenoic acid 74716 72372 74956 75052 74972 72318 77058 71448 70746 69732 75760 16602 18510 5'-Methylthioadenosine 64062 52308 58022 77044 75194 73216 70898 53600 53128 41452 44564 70282 67364 Acrylamide 378462 692310 699450 737722 708144 738288 735246 733834 727676 661846 678372 762818 735206 Adenine 482192 201998 196652 597668 587756 0 0 577874 574206 367278 355148 770412 9272 Adenosine monophosphate 125264 80392 81030 138264 140928 149544 154802 0 142184 129648 132450 81270 79794 Adenylsuccinic acid 85180 12876 13476 71996 71464 89222 85466 65382 71040 68728 66064 16500 15058 ADP 9226 1792 1754 3418 2774 8070 8926 7744 9152 7050 7028 2164 1656 Benzocaine 63444 47098 42948 70492 71650 72168 75034 70936 69758 53860 48924 76240 72124 Cadaverine 2856 3426 3128 4172 3890 4674 4108 3654 3914 4208 3776 1910 2180 Caffeine 451178 343164 362312 358250 337984 357264 375800 316646 315902 316526 331694 177410 164052 Cholic acid 5902 2330 0 242 358 0 0 21166 20168 24052 1374 0 0 Cinnamic acid 4046 3626 3280 6896 5238 6876 5482 4992 5458 3958 3762 4830 6088 Creatine 93862 71728 71182 114928 115752 107644 107784 97496 98144 88314 88404 103030 105438 Cyclic AMP 2970 1296 1062 2328 2528 2718 2976 3106 2398 2510 2416 1710 1650 Cytidine 2060 1482 0 2304 2364 2140 2194 2564 2168 1696 1716 2936 2852 D-Alanine 40440 30124 29912 51268 49428 51020 48204 41024 46700 34902 34614 46068 44534 Diacetyl 37768 33482 32662 35052 30178 36712 46244 33926 35060 33368 31308 34364 34572 Glutathione 114786 30420 30758 90692 90754 99218 113800 103738 109102 86728 86962 83638 84738 Glycerol 23662 19880 22440 33026 22752 28662 19638 21368 20456 24246 27954 22478 18426 Glycerophosphocholine 22122 16024 15310 24104 22876 21380 22302 18008 17474 16812 16586 16930 16096 Guaifenesin 11424 7798 9912 10400 9910 8750 11038 5782 5252 5580 6872 10618 7088 Guanine 79466 77002 71648 110138 117944 98256 97046 125446 120512 88020 88662 173098 181246 Guanosine 60520 54358 52872 87090 87720 70446 68514 88212 81856 66090 62666 124920 131704 Guanosine 5'-diphosphate 12520 6106 6000 5000 4972 8000 8192 7628 8550 6344 6944 5486 4334 Guanosine monophosphate 19218 14782 15714 32506 31726 26116 27012 0 27230 23686 23024 27794 25682 Hyodeoxycholic acid 40544 20862 20834 10166 10190 20716 9662 22992 2374 25740 29904 2802 2562 Hypoxanthine 567314 644958 636882 725226 726094 699424 666380 738324 731124 634648 637646 739490 763420 Indole 6430 5456 4420 8120 8782 6928 7030 0 6570 4508 4646 8720 7184 Indoleacrylic acid 118188 100112 96894 156606 152826 139976 129226 109872 109374 91618 91528 147588 141986 Inosine 156374 200234 191136 272244 257928 220740 208310 271120 261938 206876 199744 298234 297036 Isobutyric acid 219072 188734 185696 174322 166516 181006 200248 230576 223228 194792 198904 141654 150942 Isovalerylcarnitine 9412 7048 8214 17648 17300 12956 13212 10110 9832 20640 20458 18524 14966 L(-)-Nicotine pestanal 2924 3390 2728 4862 3718 2784 3118 3338 3852 3008 3198 2524 2598 L-Acetylcarnitine 64086 38850 37004 49956 50350 82554 87436 66746 66258 75270 71664 67730 67356 L-Aspartic acid 18546 14192 14212 24512 23900 0 0 20666 21040 18390 17544 24172 23846 L-Carnitine 17924 17078 16718 26362 25724 23276 24722 19178 20584 23286 22386 25584 27730 L-Glutamic acid 33008 30618 31526 54756 47402 50216 51704 26434 0 32138 32942 32690 31454 L-Norleucine 3206 2504 1892 3104 2992 3378 2582 3300 3512 3294 2936 4868 4966 L-Tryptophan 29828 20220 18666 35334 34684 31204 30262 33670 27944 20006 23184 35442 37310 m-Coumaric acid 5332 3088 3674 5572 6778 4546 3824 5544 7216 2464 2830 7956 8376 Maltitol 10774 11456 11512 11632 11668 11628 12550 13202 11796 10998 11148 11308 10922 Medroxyprogesterone 22884 9914 11132 10146 11582 8298 5430 21584 24686 24884 20088 0 714 Methylhippuric acid 5420 6544 5200 5834 5004 5142 5784 5560 5998 4826 4854 5452 5450 N,N-Dimethylformamide 16370 31800 40030 83044 64572 0 85068 18406 16652 14294 10734 49194 90688 N-Acetyl-L-alanine 3112 2880 2982 3018 3390 2956 2938 2738 2598 2726 2852 3256 3136 N-Acetyl-L-aspartic acid 15820 11910 11558 18766 18560 17180 18860 15504 17030 15520 15076 0 20648 Niacinamide 13482 1832 1546 5098 4812 10840 12892 10316 12374 10740 10280 2712 3062 Normetanephrine 5302 4984 4978 6386 6458 5956 6200 6136 6976 4378 4408 6956 8110 Nutriacholic acid 35938 32514 34768 21236 19836 44984 33708 15988 16446 22554 21362 676 616 o-Tyrosine 5918 5222 4438 6786 8618 6782 6896 7026 7800 2912 3412 9920 10632 Pantothenic acid 93470 73722 71698 115524 104324 97210 96844 98356 107834 71872 73616 121604 123984 Phenylacetaldehyde 80258 92648 87680 87298 77218 83270 62748 90364 89934 91462 85668 0 1732 Phenylacetic acid 1886 2114 2646 1822 2076 1952 1600 2616 2442 2988 2638 2178 2480 Phosphocreatine 3582 3098 3102 0 0 0 0 3218 3138 0 0 0 0 Phosphoric acid 6274 3854 4430 5956 5892 5266 4856 5706 6030 4906 5826 7402 6508 Propionic acid 5638 4330 5176 2856 6078 7054 5948 484 1570 2424 2930 3402 4446 Propylene glycol 20648 19172 19170 20252 19594 20124 25912 20912 20568 17088 18884 20952 20668 Pyridine 19722 11872 11574 16958 22634 19892 16024 25032 24344 14562 22908 14016 24316 Pyroglutamic acid 145388 96222 93390 161960 157810 130444 164906 136462 160222 115092 111312 151680 157514 Rhamnose 4494 4972 5184 4476 4456 5524 5394 4480 4216 4234 3736 4948 3980 S-Adenosylhomocysteine 11808 13568 13098 18128 17186 17908 17504 19538 17344 11896 11866 22058 20970 Saccharopine 2426 5410 7270 4184 5994 4310 4682 2768 2870 2346 5446 5636 5356 Sorbitol 11768 12782 13154 11884 12296 12584 12788 11272 11450 11984 11844 12160 12154 Spermine 27558 16398 16602 17368 18542 24190 20172 24706 23826 16240 16720 7010 6368 Sphinganine 27664 20910 0 18334 13516 24890 0 16182 13622 13272 12426 306 328 Sphingosine 5448 9670 6654 8648 9222 5966 3786 5328 5800 5594 5578 1020 948 Succinic acid 14392 17662 17528 18870 19386 15010 14604 19552 20358 9848 9968 31156 30402 Uracil 5652 4322 4998 8252 7508 7790 8352 4718 5752 4628 4540 5970 4482 Urea 3298 2692 3644 3074 3460 2952 2816 2076 3040 3070 2944 2448 2798 Uridine 3682 3076 3152 5120 4026 4434 4912 2232 3132 2456 2462 3338 3304 Valeric acid 3300 2350 2622 3542 2974 3512 4682 3026 3366 2846 2794 3358 3756 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name RT [min] m/z meas. M meas. Ions 2,2-Dimethylsuccinic acid 8.68 147.06514 146.05787 [M+H]+ 2-Hydroxycaproic acid 10.36 133.08632 132.07904 [M+H]+ 2-Ketobutyric acid 8.9 103.03923 102.03195 [M+H]+ 2-Ketohexanoic acid 16.09 131.07092 130.06365 [M+H]+ 2-Phenylbutyric acid 17.22 165.09138 164.0841 [M+H]+ 2-Pyrrolidinone 19.79 86.06019 85.05291 [M+H]+ 3,4-Dihydroxyphenylglycol 7.39 171.06282 170.05555 [M+H]+ 3-Aminoisobutanoic acid 10.15 104.07095 103.06368 [M+H]+ 3-Hexenedioic acid 3.79 145.04948 144.04221 [M+H]+ 3b-Hydroxy-5-cholenoic acid 16.43 375.2898 374.28252 [M+H]+ 5'-Methylthioadenosine 8.55 298.09695 297.08968 [M+H]+ Acrylamide 9.22 72.04433 71.03706 [M+H]+ Adenine 7.67 136.06165 135.05438 [M+H]+ Adenosine monophosphate 4.26 348.07029 347.06301 [M+H]+ Adenylsuccinic acid 8.03 464.08134 463.07406 [M+H]+ ADP 7.4 428.03672 427.02944 [M+H]+ Benzocaine 8.16 166.08629 165.07901 [M+H]+ Cadaverine 10.73 103.12333 102.11605 [M+H]+ Caffeine 9.25 195.0876 194.08033 [M+H]+ Cholic acid 20.92 409.2928 408.28553 [M+H]+ Cinnamic acid 8.16 149.05978 148.0525 [M+H]+ Creatine 3.07 132.07671 131.06944 [M+H]+ Cyclic AMP 7.82 330.06 329.05273 [M+H]+ Cytidine 3.81 244.09285 243.08557 [M+H]+ D-Alanine 3.04 90.05504 89.04777 [M+H]+ Diacetyl 8.4 87.04427 86.037 [M+H]+ Glutathione 4.07 308.09108 307.08381 [M+H]+ Glycerol 3.32 93.0548 92.04753 [M+H]+ Glycerophosphocholine 3.46 258.11018 257.1029 [M+H]+ Guaifenesin 9.68 199.09434 198.08706 [M+H]+ Guanine 7.8 152.05668 151.04941 [M+H]+ Guanosine 7.8 284.099 283.09173 [M+H]+ Guanosine 5'-diphosphate 7.39 444.03169 443.02442 [M+H]+ Guanosine monophosphate 4.87 364.0653 363.05802 [M+H]+ Hyodeoxycholic acid 21.59 393.29808 392.29081 [M+H]+ Hypoxanthine 7.82 137.04567 136.03843 [M+H]+, [M-H2O+H]+ Indole 8.7 118.06523 117.05796 [M+H]+ Indoleacrylic acid 8.71 188.07068 187.0634 [M+H]+ Inosine 7.81 269.08799 268.08072 [M+H]+ Isobutyric acid 10.07 89.05968 88.0524 [M+H]+ Isovalerylcarnitine 8.94 246.17011 245.16283 [M+H]+ L(-)-Nicotine pestanal 7.38 163.12304 162.11577 [M+H]+ L-Acetylcarnitine 3.8 204.12307 203.1158 [M+H]+ L-Aspartic acid 4.02 134.04475 133.03747 [M+H]+ L-Carnitine 3.02 162.11253 161.10525 [M+H]+ L-Glutamic acid 7.41 148.06042 147.05314 [M+H]+ L-Norleucine 5.65 132.10194 131.09467 [M+H]+ L-Tryptophan 8.7 205.09729 204.09002 [M+H]+ m-Coumaric acid 6 165.05473 164.04746 [M+H]+ Maltitol 3.8 345.13921 344.13193 [M+H]+ Medroxyprogesterone 19.92 345.24047 344.23319 [M+H]+ Methylhippuric acid 8.21 194.07885 193.07157 [M+H]+ N,N-Dimethylformamide 7.3 74.06014 73.05286 [M+H]+ N-Acetyl-L-alanine 22.51 132.06557 131.05829 [M+H]+ N-Acetyl-L-aspartic acid 4.06 176.05543 175.04815 [M+H]+ Niacinamide 7.4 123.05537 122.04809 [M+H]+ Normetanephrine 8.41 184.09692 183.08964 [M+H]+ Nutriacholic acid 15.52 391.28466 390.27738 [M+H]+ o-Tyrosine 6.08 182.0813 181.07403 [M+H]+ Pantothenic acid 8.4 220.1181 219.11083 [M+H]+ Phenylacetaldehyde 17.23 121.06536 120.05809 [M+H]+ Phenylacetic acid 12.84 137.05978 136.05251 [M+H]+ Phosphocreatine 3.66 212.04314 211.03587 [M+H]+ Phosphoric acid 3.57 98.98443 97.97715 [M+H]+ Propionic acid 0.31 75.04424 74.03697 [M+H]+ Propylene glycol 8.4 77.05979 76.05251 [M+H]+ Pyridine 4.32 80.0496 79.04232 [M+H]+ Pyroglutamic acid 7.5 130.04987 129.04259 [M+H]+ Rhamnose 7.92 165.07585 164.06714 [M+H]+, [M+K]+ S-Adenosylhomocysteine 7.51 385.12898 384.12119 [M+H]+, [M+H+H]2+ Saccharopine 14.56 277.1413 276.13402 [M+H]+ Sorbitol 3.75 183.08641 182.07913 [M+H]+ Spermine 1.91 203.22315 202.21587 [M+H]+ Sphinganine 13.65 302.30557 301.29829 [M+H]+ Sphingosine 19.63 300.29019 299.2829 [M+H]+, [M+Na]+ Succinic acid 7.69 119.03529 118.02801 [M+H]+ Uracil 7.43 113.03449 112.02721 [M+H]+ Urea 22.81 61.03975 60.03248 [M+H]+ Uridine 7.37 245.07682 244.06955 [M+H]+ Valeric acid 8.39 103.07568 102.06841 [M+H]+ METABOLITES_END #END