#METABOLOMICS WORKBENCH Zhou999_20161213_161728 DATATRACK_ID:798 STUDY_ID:ST000522 ANALYSIS_ID:AN000796 PROJECT_ID:PR000385 VERSION 1 CREATED_ON December 19, 2016, 10:24 am #PROJECT PR:PROJECT_TITLE Metabolomics profiling studies for snout bream fish PR:PROJECT_TYPE LC-MS analysis PR:PROJECT_SUMMARY Metabolomics profiling studies on plasma samples from three different blunt PR:PROJECT_SUMMARY snout bream groups PR:INSTITUTE Huazhong Agricultural University, Wuhan PR:DEPARTMENT College of Fisheries PR:LABORATORY Key Lab of Freshwater Animal Breeding, Ministry of Agriculture PR:LAST_NAME Zhou PR:FIRST_NAME LaiFang PR:ADDRESS Hongshan, Wuhan, 430070 Hubei, China PR:EMAIL 156851836@qq.com PR:PHONE 15171617087 #STUDY ST:STUDY_TITLE Plasma metabolomics profiling for fish maturation in blunt snout bream ST:STUDY_TYPE Metabolomics experiment ST:STUDY_SUMMARY We investigated the comprehensive metabolic profiles of plasma among immature ST:STUDY_SUMMARY females, mature females ready to spawn, as well as already spawned breeders of ST:STUDY_SUMMARY blunt snout bream (Megalobrama amblycephala). The purpose of this study was to ST:STUDY_SUMMARY screen out potential biomarkers for sexual mature female M. amblycephala ST:STUDY_SUMMARY compared to immature female individuals and already spawned breeders. The three ST:STUDY_SUMMARY groups were set up in this study, including one year old immature females, 2 ST:STUDY_SUMMARY years old sexually mature females ready to spawning and successfully spawning ST:STUDY_SUMMARY females of M. amblycephala. The plasma samples were collected to investigate the ST:STUDY_SUMMARY comprehensive metabolic profiles through UPLC-MS/MS based metabolomics analysis ST:STUDY_SUMMARY method. According to multivariate and univariate statistical analysis, plasma ST:STUDY_SUMMARY metabolite profiles of three groups were obviously separated, and the plasma ST:STUDY_SUMMARY metabolite profiles of immature female M. amblycephala were much more different ST:STUDY_SUMMARY from mature females ready to spawn as well as already spawned breeders. The ST:STUDY_SUMMARY differential plasma metabolites from three hormone related pathways including ST:STUDY_SUMMARY GnRH signaling pathway, steroid hormone biosynthesis and steroid biosynthesis, ST:STUDY_SUMMARY were further analyzed. A total of 29 metabolites were identified as differential ST:STUDY_SUMMARY biomarkers associated with the female maturation status ST:INSTITUTE Huazhong Agricultural University, Wuhan ST:DEPARTMENT College of Fisheries ST:LABORATORY Key Lab of Freshwater Animal Breeding, Ministry of Agriculture ST:LAST_NAME Zhou ST:FIRST_NAME LaiFang ST:ADDRESS Hongshan, Wuhan, 430070 Hubei, China ST:EMAIL 156851836@qq.com ST:PHONE 15171617087 #SUBJECT SU:SUBJECT_TYPE Animal SU:SUBJECT_SPECIES Megalobrama amblycephala SU:TAXONOMY_ID 75352 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS P1503530001 20150724_YU_2_1_1 Phenotype:2 years old sexually mature without hormone treatment gender=female SUBJECT_SAMPLE_FACTORS P1503530002 20150724_YU_2_1_2 Phenotype:2 years old sexually mature without hormone treatment gender=female SUBJECT_SAMPLE_FACTORS P1503530003 20150724_YU_2_1_3 Phenotype:2 years old sexually mature without hormone treatment gender=female SUBJECT_SAMPLE_FACTORS P1503530004 20150724_YU_2_1_4 Phenotype:2 years old sexually mature without hormone treatment gender=female SUBJECT_SAMPLE_FACTORS P1503530005 20150724_YU_2_1_5 Phenotype:2 years old sexually mature without hormone treatment gender=female SUBJECT_SAMPLE_FACTORS P1503530006 20150724_YU_2_1_6 Phenotype:2 years old sexually mature without hormone treatment gender=female SUBJECT_SAMPLE_FACTORS P1503530007 20150724_YU_2_1_7 Phenotype:2 years old sexually mature without hormone treatment gender=female SUBJECT_SAMPLE_FACTORS P1503530008 20150724_YU_2_1_8 Phenotype:2 years old sexually mature without hormone treatment gender=female SUBJECT_SAMPLE_FACTORS P1503530009 20150724_YU_2_1_9 Phenotype:2 years old sexually mature without hormone treatment gender=female SUBJECT_SAMPLE_FACTORS P1503530010 20150724_YU_2_1_10 Phenotype:2 years old sexually mature without hormone treatment gender=female SUBJECT_SAMPLE_FACTORS P1503530011 20150724_YU_2_2_1 Phenotype:2 years old sexually mature after successfully spawning gender=female SUBJECT_SAMPLE_FACTORS P1503530012 20150724_YU_2_2_2 Phenotype:2 years old sexually mature after successfully spawning gender=female SUBJECT_SAMPLE_FACTORS P1503530013 20150724_YU_2_2_3 Phenotype:2 years old sexually mature after successfully spawning gender=female SUBJECT_SAMPLE_FACTORS P1503530014 20150724_YU_2_2_4 Phenotype:2 years old sexually mature after successfully spawning gender=female SUBJECT_SAMPLE_FACTORS P1503530015 20150724_YU_2_2_5 Phenotype:2 years old sexually mature after successfully spawning gender=female SUBJECT_SAMPLE_FACTORS P1503530016 20150724_YU_2_2_6 Phenotype:2 years old sexually mature after successfully spawning gender=female SUBJECT_SAMPLE_FACTORS P1503530017 20150724_YU_2_2_11 Phenotype:2 years old sexually mature after successfully spawning gender=female SUBJECT_SAMPLE_FACTORS P1503530018 20150724_YU_2_2_8 Phenotype:2 years old sexually mature after successfully spawning gender=female SUBJECT_SAMPLE_FACTORS P1503530019 20150724_YU_2_2_9 Phenotype:2 years old sexually mature after successfully spawning gender=female SUBJECT_SAMPLE_FACTORS P1503530020 20150724_YU_2_2_10 Phenotype:2 years old sexually mature after successfully spawning gender=female SUBJECT_SAMPLE_FACTORS P1503530021 20150724_YU_1_1_1 Phenotype:1 year old immature gender=female SUBJECT_SAMPLE_FACTORS P1503530022 20150724_YU_1_1_3 Phenotype:1 year old immature gender=female SUBJECT_SAMPLE_FACTORS P1503530023 20150724_YU_1_1_5 Phenotype:1 year old immature gender=female SUBJECT_SAMPLE_FACTORS P1503530024 20150724_YU_1_1_6 Phenotype:1 year old immature gender=female SUBJECT_SAMPLE_FACTORS P1503530025 20150724_YU_1_1_9 Phenotype:1 year old immature gender=female SUBJECT_SAMPLE_FACTORS P1503530026 20150724_YU_1_1_10 Phenotype:1 year old immature gender=female SUBJECT_SAMPLE_FACTORS P1503530027 20150724_YU_1_1_11 Phenotype:1 year old immature gender=female SUBJECT_SAMPLE_FACTORS P1503530028 20150724_YU_1_1_12 Phenotype:1 year old immature gender=female SUBJECT_SAMPLE_FACTORS P1503530029 20150724_YU_1_1_14 Phenotype:1 year old immature gender=female SUBJECT_SAMPLE_FACTORS P1503530030 20150724_YU_1_1_22 Phenotype:1 year old immature gender=female #COLLECTION CO:COLLECTION_SUMMARY About 2 ml of blood was collected via caudal vein by syringe for each female CO:COLLECTION_SUMMARY M.amblycephala and then directly put into K3 EDTA centrifuge tube. CO:COLLECTION_PROTOCOL_FILENAME Zhou999_20161213_161728_PR_CO_collection_files.docx CO:SAMPLE_TYPE Plasma #TREATMENT TR:TREATMENT_SUMMARY The blood was centrifuged at 4000 rpm for 5 min at 4°C as soon as it was TR:TREATMENT_SUMMARY extracted from fish and about 1 ml supernatant was pipetted into another TR:TREATMENT_SUMMARY tube.The extracted plasma samples were then frozen in liquid nitrogen TR:TREATMENT_SUMMARY immediately and stored in the -80°C. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY To initiate the experiment, each plasma sample was mixed with the same volume SP:SAMPLEPREP_SUMMARY and labeled as Quality control (QC) sample. Plasma sample (100 µl, including QC SP:SAMPLEPREP_SUMMARY sample) was added into new Eppendorf tube with ice-cold methanol (300 µl), SP:SAMPLEPREP_SUMMARY vortex mixing for 1 min, standing for 1 min, and make sure the solution volume SP:SAMPLEPREP_SUMMARY had no obvious observable difference. Then samples were precipitated for 20 min SP:SAMPLEPREP_SUMMARY at -20°C, and centrifuged at 14000 rpm for 20 min at 4°C to precipitate the SP:SAMPLEPREP_SUMMARY proteins. A total of 250 µl of protein free supernatant was collected and dried SP:SAMPLEPREP_SUMMARY with nitrogen at 37°C. The concentration of extractant was diluted depending on SP:SAMPLEPREP_SUMMARY the mass spectrum response signals. SP:SAMPLEPREP_PROTOCOL_FILENAME Zhou999_20161213_161728_PR_SP_sampleprep.docx #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Chromatographic separation was performed on a ACQUITY UPLC BEH C18 analytical CH:CHROMATOGRAPHY_SUMMARY column (100×2.1 mm, 1.7 µm, Waters, Milford, USA) using a 2777C UPLC system CH:CHROMATOGRAPHY_SUMMARY (Waters). The flow rate was 0.40 mL/min and the mobile phase was composed of CH:CHROMATOGRAPHY_SUMMARY solvents A (95% H2O/5% acetonitrile + 0.1% formic acid) and B (95% CH:CHROMATOGRAPHY_SUMMARY acetonitrile/5% H2O＋0.1% formic acid).The gradient program was optimized as CH:CHROMATOGRAPHY_SUMMARY follows: 0–0.1 min, 100% A; 0.1–0.6 min, 100% A-50% A; 0.6–5 min, 50%–0% CH:CHROMATOGRAPHY_SUMMARY A; 5–8 min, 0% A; 8–10 min, 0% A-100% A. Injection volume was 10 µl. The CH:CHROMATOGRAPHY_SUMMARY eluent from the column was directly added in to the mass spectrometer without CH:CHROMATOGRAPHY_SUMMARY split. CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Waters 2777C CH:COLUMN_NAME Waters Acquity BEH C18 (150 x 2mm, 1.7um) CH:FLOW_GRADIENT 100% acetonitrile CH:FLOW_RATE 0.40 mL/min CH:SOLVENT_A 95% H2O/5% acetonitrile + 0.1% formic acid CH:SOLVENT_B 95% acetonitrile/5% H2O＋0.1% formic acid #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:MS_COMMENTS - MS:INSTRUMENT_NAME Waters Synapt G2 XS QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE POSITIVE #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS peak intensity MS_METABOLITE_DATA_START Samples 20150724_YU_2_1_1 20150724_YU_2_1_2 20150724_YU_1_1_1 20150724_YU_2_1_3 20150724_YU_2_1_4 20150724_YU_1_1_3 20150724_YU_2_1_5 20150724_YU_2_1_6 20150724_YU_1_1_5 20150724_YU_2_1_7 20150724_YU_2_1_8 20150724_YU_1_1_6 20150724_YU_2_1_9 20150724_YU_2_1_10 20150724_YU_1_1_9 20150724_YU_2_2_1 20150724_YU_2_2_2 20150724_YU_1_1_10 20150724_YU_2_2_3 20150724_YU_2_2_4 20150724_YU_1_1_11 20150724_YU_2_2_5 20150724_YU_2_2_6 20150724_YU_1_1_12 20150724_YU_2_2_11 20150724_YU_2_2_8 20150724_YU_1_1_14 20150724_YU_2_2_9 20150724_YU_2_2_10 20150724_YU_1_1_22 20150724_YU_2_1_1 20150724_YU_2_1_2 20150724_YU_1_1_1 20150724_YU_2_1_3 20150724_YU_2_1_4 20150724_YU_1_1_3 20150724_YU_2_1_5 20150724_YU_2_1_6 20150724_YU_1_1_5 20150724_YU_2_1_7 20150724_YU_2_1_8 20150724_YU_1_1_6 20150724_YU_2_1_9 20150724_YU_2_1_10 20150724_YU_1_1_9 20150724_YU_2_2_1 20150724_YU_2_2_2 20150724_YU_1_1_10 20150724_YU_2_2_3 20150724_YU_2_2_4 20150724_YU_1_1_11 20150724_YU_2_2_5 20150724_YU_2_2_6 20150724_YU_1_1_12 20150724_YU_2_2_11 20150724_YU_2_2_8 20150724_YU_1_1_14 20150724_YU_2_2_9 20150724_YU_2_2_10 20150724_YU_1_1_22 Factors Phenotype:2 years old sexually mature without hormone treatment Phenotype:2 years old sexually mature without hormone treatment Phenotype:1 year old immature Phenotype:2 years old sexually mature without hormone treatment Phenotype:2 years old sexually mature without hormone treatment Phenotype:1 year old immature Phenotype:2 years old sexually mature without hormone treatment Phenotype:2 years old sexually mature without hormone treatment Phenotype:1 year old immature Phenotype:2 years old sexually mature without hormone treatment Phenotype:2 years old sexually mature without hormone treatment Phenotype:1 year old immature Phenotype:2 years old sexually mature without hormone treatment Phenotype:2 years old sexually mature without hormone treatment Phenotype:1 year old immature Phenotype:2 years old sexually mature after successfully spawning Phenotype:2 years old sexually mature after successfully spawning Phenotype:1 year old immature Phenotype:2 years old sexually mature after successfully spawning Phenotype:2 years old sexually mature after successfully spawning Phenotype:1 year old immature Phenotype:2 years old sexually mature after successfully spawning Phenotype:2 years old sexually mature after successfully spawning Phenotype:1 year old immature Phenotype:2 years old sexually mature after successfully spawning Phenotype:2 years old sexually mature after successfully spawning Phenotype:1 year old immature Phenotype:2 years old sexually mature after successfully spawning Phenotype:2 years old sexually mature after successfully spawning Phenotype:1 year old immature Phenotype:2 years old sexually mature without hormone treatment Phenotype:2 years old sexually mature without hormone treatment Phenotype:1 year old immature Phenotype:2 years old sexually mature without hormone treatment Phenotype:2 years old sexually mature without hormone treatment Phenotype:1 year old immature Phenotype:2 years old sexually mature without hormone treatment Phenotype:2 years old sexually mature without hormone treatment Phenotype:1 year old immature Phenotype:2 years old sexually mature without hormone treatment Phenotype:2 years old sexually mature without hormone treatment Phenotype:1 year old immature Phenotype:2 years old sexually mature without hormone treatment Phenotype:2 years old sexually mature without hormone treatment Phenotype:1 year old immature Phenotype:2 years old sexually mature after successfully spawning Phenotype:2 years old sexually mature after successfully spawning Phenotype:1 year old immature Phenotype:2 years old sexually mature after successfully spawning Phenotype:2 years old sexually mature after successfully spawning Phenotype:1 year old immature Phenotype:2 years old sexually mature after successfully spawning Phenotype:2 years old sexually mature after successfully spawning Phenotype:1 year old immature Phenotype:2 years old sexually mature after successfully spawning Phenotype:2 years old sexually mature after successfully spawning Phenotype:1 year old immature Phenotype:2 years old sexually mature after successfully spawning Phenotype:2 years old sexually mature after successfully spawning Phenotype:1 year old immature 16a/16b-Hydroxyestrone 105087.2972 88938.85216 135947.0284 72664.83197 78592.33569 136983.7912 87602.83566 90660.79711 152198.3772 122120.2283 69975.96797 153124.1744 54523.71478 79941.76492 273899.5475 135601.6603 95940.03038 159631.3067 86270.31074 106418.8968 145941.0435 72138.69979 97960.77836 181175.331 88245.90714 94056.75268 134826.7919 88839.68151 141752.6624 159494.7541 130164.2597 118574.6527 128725.4467 82109.3556 79423.39059 113131.4717 93218.70774 80384.86953 139149.6455 124370.9985 71572.8385 139376.2551 53436.31971 85258.16133 237910.2393 138239.9949 106804.8837 151410.8486 86270.31074 103738.3429 141653.137 71492.25817 88066.08851 164818.7773 88249.93373 95873.24131 122291.6475 89989.91843 140692.2328 164741.725 2-Methoxyestrone 44679.00074 81890.08677 92901.64541 102729.7511 59895.66268 30291.65145 26188.94366 82305.76284 311749.8805 84321.47148 23458.9788 81846.88796 28193.95811 96472.19683 285900.8469 40422.43624 60707.17243 498136.5472 61390.02098 42049.15159 322842.0102 29852.4348 56315.44134 323113.9809 41631.71472 27662.79628 237739.7818 27246.77768 59115.15701 349306.1755 55340.74252 109177.1297 87966.6584 116081.9262 60529.01432 25017.11392 27867.81349 72976.83473 285021.9967 85875.58139 23994.31904 74498.44403 27631.6712 102887.9226 248334.6158 41208.91564 67582.035 472484.2445 61390.02098 40989.98805 313356.5609 29584.92435 50627.20738 293943.1707 41633.61434 28197.03921 215636.5896 27599.55078 58672.92573 360797.4583 21-Hydroxy-5b-pregnane-3,11,20-trione 12035.06908 15378.07806 20875.05668 18054.73541 12051.99871 16047.73048 11224.1903 9743.422938 51819.83929 6686.111506 6680.841212 26077.5772 8154.461799 16634.55464 40658.91981 5498.223902 7722.363955 66451.90201 8390.001368 9242.313295 46413.35402 3794.49406 8203.24853 35241.5473 9884.353323 4860.27099 33008.77582 7898.386784 5804.786696 62712.36765 14906.99542 20502.29142 19766.16208 20401.3778 12179.43954 13253.41744 11943.72885 8639.056865 47377.06408 6809.34171 6833.299817 23736.25894 7991.833086 17740.80851 35316.4999 5605.200131 8596.893089 63029.85977 8390.001368 9009.51142 45049.67921 3760.491244 7374.665894 32059.93169 9884.804338 4954.135883 29939.87708 8000.649826 5761.361991 64775.44469 21-Deoxycortisol 12035.06908 15378.07806 20875.05668 18054.73541 12051.99871 16047.73048 11224.1903 9743.422938 51819.83929 6686.111506 6680.841212 26077.5772 8154.461799 16634.55464 40658.91981 5498.223902 7722.363955 66451.90201 8390.001368 9242.313295 46413.35402 3794.49406 8203.24853 35241.5473 9884.353323 4860.27099 33008.77582 7898.386784 5804.786696 62712.36765 14906.99542 20502.29142 19766.16208 20401.3778 12179.43954 13253.41744 11943.72885 8639.056865 47377.06408 6809.34171 6833.299817 23736.25894 7991.833086 17740.80851 35316.4999 5605.200131 8596.893089 63029.85977 8390.001368 9009.51142 45049.67921 3760.491244 7374.665894 32059.93169 9884.804338 4954.135883 29939.87708 8000.649826 5761.361991 64775.44469 Cortexolone 12035.06908 15378.07806 20875.05668 18054.73541 12051.99871 16047.73048 11224.1903 9743.422938 51819.83929 6686.111506 6680.841212 26077.5772 8154.461799 16634.55464 40658.91981 5498.223902 7722.363955 66451.90201 8390.001368 9242.313295 46413.35402 3794.49406 8203.24853 35241.5473 9884.353323 4860.27099 33008.77582 7898.386784 5804.786696 62712.36765 14906.99542 20502.29142 19766.16208 20401.3778 12179.43954 13253.41744 11943.72885 8639.056865 47377.06408 6809.34171 6833.299817 23736.25894 7991.833086 17740.80851 35316.4999 5605.200131 8596.893089 63029.85977 8390.001368 9009.51142 45049.67921 3760.491244 7374.665894 32059.93169 9884.804338 4954.135883 29939.87708 8000.649826 5761.361991 64775.44469 Corticosterone 12035.06908 15378.07806 20875.05668 18054.73541 12051.99871 16047.73048 11224.1903 9743.422938 51819.83929 6686.111506 6680.841212 26077.5772 8154.461799 16634.55464 40658.91981 5498.223902 7722.363955 66451.90201 8390.001368 9242.313295 46413.35402 3794.49406 8203.24853 35241.5473 9884.353323 4860.27099 33008.77582 7898.386784 5804.786696 62712.36765 14906.99542 20502.29142 19766.16208 20401.3778 12179.43954 13253.41744 11943.72885 8639.056865 47377.06408 6809.34171 6833.299817 23736.25894 7991.833086 17740.80851 35316.4999 5605.200131 8596.893089 63029.85977 8390.001368 9009.51142 45049.67921 3760.491244 7374.665894 32059.93169 9884.804338 4954.135883 29939.87708 8000.649826 5761.361991 64775.44469 Pregnenolone 410361.5488 891357.3776 136502.9879 628639.2418 476537.8031 18596.89142 881581.9385 367063.3002 148767.4259 598064.7136 823693.8339 258672.9238 2000944.008 594181.1583 43936.28116 318808.8549 901864.2263 55478.77188 645250.8763 538365.4403 67659.6789 480581.4364 218220.2766 64871.84054 709120.368 971699.2392 75077.23617 2066073.314 351139.841 351277.0253 508286.0503 1188371.436 129251.8734 710345.8668 481576.8323 15358.70542 938096.6776 325458.5933 136012.8469 609087.5086 842490.7502 235448.5408 1961038.131 633696.2051 38163.22904 325011.7615 1003997.012 52621.80775 645250.8763 524804.7137 65671.76397 476274.9013 196178.578 59015.19472 709152.7246 990465.3625 68097.13983 2092823.453 348513.0185 362833.144 5alpha-Dihydrodeoxycorticosterone 18314.88181 16731.85029 46354.81241 10067.23787 11926.0573 50988.11599 17412.75804 22675.27039 17205.99285 9762.371641 12931.81925 47079.96074 12734.89571 12878.38343 11291.64353 14278.10211 9219.024265 19360.2656 6357.780876 10388.30284 20399.16301 10427.60506 24697.92065 11672.70761 16168.37123 8611.789012 44172.21258 13016.12244 11921.82692 13463.45251 22685.3587 22307.16147 43892.41902 11375.71493 12052.1664 42109.80404 18529.02124 20105.14699 15730.83662 9942.299697 13226.92687 42852.98937 12480.917 13734.83927 9807.966602 14555.9041 10263.04464 18363.27914 6357.780876 10126.63498 19799.81342 10334.16232 22203.26647 10618.89268 16169.10898 8778.105799 40065.42449 13184.64651 11832.64159 13906.36578 3a-Hydroxy-5b-pregnane-20-one 43740.66755 59795.43309 46354.81241 28644.94951 8163.602845 0 104026.7033 9534.592263 0 43058.00404 70984.95889 2.35009396 146858.7054 42708.3142 0 8955.928039 27724.81004 0 14597.32758 1573.059158 0 19999.78854 2334.796981 0 42641.0213 57265.46412 0 99194.75716 7955.310805 7.534588025 54178.49505 79720.19583 0 32368.04217 8249.926812 0 110695.4447 8453.896056 0 43851.59635 72604.85487 2.139095911 143929.8252 45548.56083 0 9130.179112 30864.54215 0 14597.32758 1533.43584 0 19820.56857 2098.966965 0 42642.96697 58371.41411 0 100479.0647 7895.798364 7.782456766 5alpha-Pregnan-20alpha-ol-3-one 43740.66755 59795.43309 46354.81241 28644.94951 8163.602845 0 104026.7033 9534.592263 0 43058.00404 70984.95889 2.35009396 146858.7054 42708.3142 0 8955.928039 27724.81004 0 14597.32758 1573.059158 0 19999.78854 2334.796981 0 42641.0213 57265.46412 0 99194.75716 7955.310805 7.534588025 54178.49505 79720.19583 0 32368.04217 8249.926812 0 110695.4447 8453.896056 0 43851.59635 72604.85487 2.139095911 143929.8252 45548.56083 0 9130.179112 30864.54215 0 14597.32758 1533.43584 0 19820.56857 2098.966965 0 42642.96697 58371.41411 0 100479.0647 7895.798364 7.782456766 Allopregnanolone 43740.66755 59795.43309 46354.81241 28644.94951 8163.602845 0 104026.7033 9534.592263 0 43058.00404 70984.95889 2.35009396 146858.7054 42708.3142 0 8955.928039 27724.81004 0 14597.32758 1573.059158 0 19999.78854 2334.796981 0 42641.0213 57265.46412 0 99194.75716 7955.310805 7.534588025 54178.49505 79720.19583 0 32368.04217 8249.926812 0 110695.4447 8453.896056 0 43851.59635 72604.85487 2.139095911 143929.8252 45548.56083 0 9130.179112 30864.54215 0 14597.32758 1533.43584 0 19820.56857 2098.966965 0 42642.96697 58371.41411 0 100479.0647 7895.798364 7.782456766 17-Hydroxyprogesterone 52919.82688 46343.73799 46354.81241 42351.87045 37142.61908 46253.5497 40658.2511 43706.88555 81388.73903 43900.79951 34747.77717 38656.45325 24286.97206 33823.16701 119591.2645 33682.22094 34819.30791 110625.9339 30584.18991 26422.47248 99692.38606 36978.30279 37432.91466 73203.22994 26623.70225 35154.21503 83618.75864 24893.83732 29037.67633 70768.77267 65548.0755 61786.18796 66008.71519 47856.50358 37535.37438 38199.64468 43264.69113 38752.93847 74410.87347 44709.92518 35540.73085 35185.7681 23802.60423 36072.52145 103877.4493 34337.5593 38762.46565 104929.0824 30584.18991 25756.9247 96763.31537 36646.93676 33651.94143 66594.42423 26624.91706 35833.13739 75844.5381 25216.14611 28820.45 73096.8849 20a,22b-Dihydroxycholesterol 49387.9512 57414.58277 46354.81241 25913.2646 63922.03016 5071.898954 59617.26469 7827.244418 8522.021412 25608.42122 57708.22611 8170.925392 79635.58825 70647.92753 2594.420974 42559.99015 81569.89484 7887.613979 22119.37887 18679.96795 28.4210877 31656.0712 14484.15733 4784.231148 104618.633 82835.92671 13566.86453 34716.95343 85554.34158 30804.54689 61173.38897 76546.00937 5250.706794 29281.30982 64597.95761 4188.753926 63439.09226 6940.067167 7791.385694 26080.40422 59025.14346 7437.316716 78047.37391 75346.25248 2253.524405 43388.05888 90807.38349 7481.429245 22119.37887 18209.44381 27.58604524 31372.39819 13021.16115 4352.309576 104623.4067 84435.71104 12305.52319 35166.44538 84914.32288 31817.93796 Estrone 135455.5604 83360.15129 46354.81241 220146.0049 197202.2678 106458.3411 285652.439 222737.8197 88016.11865 178893.0948 143975.7422 109926.6052 103619.3096 75310.14861 67690.20703 154815.7743 67672.1091 69102.5396 258764.5817 194377.4686 68289.02269 234449.17 176630.1677 66135.02302 264950.1118 174992.6219 60083.66011 117342.9964 55188.41957 77929.56849 167779.296 111137.0424 98057.56287 248759.2155 199287.5336 87921.26941 303964.4895 197491.6518 80470.05453 182190.2328 147261.3077 100057.0852 101552.7753 80318.52695 58795.98378 157827.9484 75335.72497 65543.99873 258764.5817 189481.3525 66282.61695 232348.2491 158789.3465 60164.33678 264962.2012 178372.1997 54497.54962 118862.2752 54775.5636 80493.25266 Tetrahydrodeoxycorticosterone 35100.62001 39834.80354 46354.81241 19789.60772 65513.11381 11366.22269 27007.04631 37852.48187 21196.09916 43782.96477 32797.21983 29609.1732 37128.56345 66822.55432 5152.185244 15154.69525 11726.50946 20608.29049 15773.03374 32501.82183 8680.071976 20978.80293 33364.26602 4040.229841 14339.10533 26182.27342 15406.14984 21262.36946 35942.54989 17554.3687 43476.67455 53108.37592 14829.06616 22361.73804 66205.86577 9387.077766 28738.36147 33562.10086 19378.85107 44589.91866 33545.66127 26950.7783 36388.08902 71266.47907 4475.208651 15449.55268 13054.49325 19547.03508 15773.03374 31683.14312 8425.042024 20790.80992 29994.25335 3675.476891 14339.75962 26687.92348 13973.80609 21537.66043 35673.66928 18131.86269 Calcitriol 3352276.502 7496443.565 46354.81241 9001253.07 1920324.722 240699.7425 10624990.49 1058604.785 972253.1499 5588228.9 13860632.08 2218502.908 16383840.65 3093955.446 252356.2417 1463585.736 8098092.184 310960.7282 6552594.691 1242863.428 319444.1339 5607427.999 737848.5919 213629.3003 6041982.48 13367209.36 299503.6076 7551205.388 2626958.152 3226617.278 4152229.632 9994374.455 339290.715 10171180.05 1940630.714 198787.8703 11306116.7 938617.4649 888897.0019 5691224.279 14176935.45 2019319.474 16057089.12 3299713.896 219197.6379 1492062.001 9015171.149 294947.3303 6552594.691 1211557.312 310058.5179 5557179.314 663320.979 194342.7942 6042258.171 13625366.09 271658.0961 7648973.357 2607306.287 3332764.762 4a-Methylzymosterol-4-carboxylic acid 11954323.37 12158898.3 46354.81241 6863354.789 6478517.402 1174284.402 9852236.489 2347362.534 736486.1477 6191928.185 11401772.73 1900996.942 10382188 9935253.802 683640.8874 7342434.388 11864995.23 575513.3942 4833997.01 2798499.922 718202.8667 7247852.113 2025707.904 533721.7413 15036836.44 11547913.14 619332.8403 6194595.655 10119691.6 2106810.559 14806981.38 16210431.19 920960.5556 7755411.024 6547022.856 969811.9862 10483824.49 2081301.258 673343.4895 6306050.209 11661964.27 1730320.087 10175130.57 10595981.62 593813.2012 7485292.511 13208662.02 545876.4526 4833997.01 2728009.341 697101.2857 7182903.434 1821097.939 485537.2104 15037522.56 11770934.38 561752.0993 6274799.146 10043987.77 2176119.256 4a-Carboxy-4b-methyl-5a-cholesta-8,24-dien-3b-ol 11954323.37 12158898.3 46354.81241 6863354.789 6478517.402 1174284.402 9852236.489 2347362.534 736486.1477 6191928.185 11401772.73 1900996.942 10382188 9935253.802 683640.8874 7342434.388 11864995.23 575513.3942 4833997.01 2798499.922 718202.8667 7247852.113 2025707.904 533721.7413 15036836.44 11547913.14 619332.8403 6194595.655 10119691.6 2106810.559 14806981.38 16210431.19 920960.5556 7755411.024 6547022.856 969811.9862 10483824.49 2081301.258 673343.4895 6306050.209 11661964.27 1730320.087 10175130.57 10595981.62 593813.2012 7485292.511 13208662.02 545876.4526 4833997.01 2728009.341 697101.2857 7182903.434 1821097.939 485537.2104 15037522.56 11770934.38 561752.0993 6274799.146 10043987.77 2176119.256 Calcidiol 83463.37723 99005.038 46354.81241 76843.25872 53816.84216 6136.16542 125458.3971 70437.50953 5713.394286 97676.95149 94455.42622 7932.328236 126940.1297 85915.31253 1867.088812 65447.95072 81681.74173 3290.595734 78474.86881 39521.59149 6647.727465 72506.69827 38822.36735 12061.09463 79232.05258 90398.71424 10107.81934 150471.9589 56704.29626 18270.75215 103380.2277 131995.0473 6977.821913 86830.8683 54385.91483 5067.704864 133501.0398 62453.78591 5223.556285 99477.21321 96610.92461 7220.141484 124408.496 91628.96996 1621.760789 66721.3392 90931.89664 3121.141479 78474.86881 38526.0939 6452.410004 71856.95899 34901.05016 10972.21603 79235.66787 92144.55632 9168.073064 152420.1694 56280.10025 18871.81332 (S)-2,3-Epoxysqualene 38649.14302 32454.73361 46354.81241 28935.93779 22430.18973 4202.87947 26829.31819 8215.827262 5581.912282 20401.13888 22831.4319 12983.57957 20259.05865 30767.17343 916.5690107 27757.05887 23093.80119 415.2404767 21830.62802 10458.61251 0 23048.98659 5905.346001 0 26800.13919 23583.78662 352.863311 21319.66165 27892.20743 15192.08701 47871.98096 43269.15259 3847.924438 32696.85129 22667.37213 3471.052567 28549.2399 7284.606176 5103.346893 20777.14764 23352.45135 11817.87726 19855.02161 32813.29401 796.135498 28297.11427 25709.0887 393.8570338 21830.62802 10195.17364 0 22842.44247 5308.866796 0 26801.36206 24039.25291 320.0568302 21595.69441 27683.55017 15691.86795 Obtusifoliol 138621.1511 268172.2738 46354.81241 507978.3037 432050.0332 502114.3949 364161.8582 408734.2108 173104.2623 281604.3277 293258.3636 375077.652 260768.183 708334.6816 44118.71146 82266.84768 328850.9281 55541.98668 128127.9355 202719.9083 171585.9222 320681.6851 195943.396 52354.52656 176497.1286 320750.3697 198627.3395 403309.8987 454100.1872 123887.2918 171700.291 357531.4213 89370.35522 574002.1692 436618.6377 414683.6643 387506.8378 362406.324 158263.1641 286794.5132 299950.5988 341402.1249 255567.5463 755441.3218 38321.68872 83867.47314 366092.0785 52681.76721 128127.9355 197613.657 166544.5412 317808.0266 176151.8103 47627.94695 176505.182 326944.9212 180160.5173 408531.6862 450703.1344 127962.868 7-Dehydrodesmosterol 60693.52153 90702.5814 46354.81241 1867164.713 2871613.294 22364.43066 678646.1488 496759.8444 2764942.325 96930.85908 67580.2066 2453334.351 310929.0959 5952771.96 239217.7348 108323.6044 131810.5309 1038464.395 57636.1438 93682.0458 185836.7617 151316.3139 162826.4658 255866.4357 145329.369 293254.0216 278634.2339 187766.5165 63274.10178 2623949.023 75176.80548 120926.0838 1157853.62 2109847.188 2901978.448 18470.22143 722151.4753 440454.712 2527889.926 98717.36974 69122.40521 2233067.089 304728.0737 6348651.329 207785.4784 110431.2033 146737.5857 984987.084 57636.1438 91322.31667 180376.6756 149960.3543 146379.9103 232766.7506 145336.0003 298917.5447 252728.9942 190197.592 62800.75808 2710270.258 Arachidonic acid 9225.799876 12758.97163 46354.81241 7272.919636 10199.43766 13963.38074 6332.217167 7980.423755 26693.09001 5301.674323 9771.879792 31148.26007 13830.28271 20319.08287 11322.58072 14149.26767 43561.51844 15002.65566 127268.6122 121475.2668 4009.816268 66146.87509 196339.1353 8685.354258 4189.065379 85124.67966 31954.55335 81644.96408 163109.5617 9239.224851 11427.35082 17010.45822 3629.314905 8218.208567 10307.28906 11532.00535 6738.150621 7075.884426 24404.55727 5399.388279 9994.876734 28351.68162 13554.45813 21670.37026 9834.838772 14424.56298 48494.69916 14230.07098 127268.6122 118415.4625 3892.003506 65554.12677 176507.5774 7901.238327 4189.256523 86768.66596 28983.668 82702.0486 161889.3645 9543.171798 Androsterone glucuronide 140445.7491 105268.4131 46354.81241 112552.2551 122859.3764 125891.8114 95123.70313 109351.9877 113017.0303 100458.0513 104058.8113 172217.2413 105910.7816 98176.64638 108763.4288 93114.32976 70682.43799 74437.7768 75848.06997 80465.40139 86928.67716 78270.27705 119471.548 113064.1146 107102.3434 72456.97967 75678.85405 55216.28065 61554.14086 55065.06014 173960.2925 140345.4758 108996.2868 127181.0983 124158.522 103970.8843 101221.7083 96957.5113 103327.5124 102309.5708 106433.4616 156755.0928 103798.5473 104705.7238 94472.34805 94926.00933 78686.96246 70604.48974 75848.06997 78438.58237 84374.61811 77568.8898 107404.1274 102856.6584 107107.2304 73856.31864 68642.82396 55931.18421 61093.66392 56876.56026 Etiocholanolone glucuronide 140445.7491 105268.4131 46354.81241 112552.2551 122859.3764 125891.8114 95123.70313 109351.9877 113017.0303 100458.0513 104058.8113 172217.2413 105910.7816 98176.64638 108763.4288 93114.32976 70682.43799 74437.7768 75848.06997 80465.40139 86928.67716 78270.27705 119471.548 113064.1146 107102.3434 72456.97967 75678.85405 55216.28065 61554.14086 55065.06014 173960.2925 140345.4758 108996.2868 127181.0983 124158.522 103970.8843 101221.7083 96957.5113 103327.5124 102309.5708 106433.4616 156755.0928 103798.5473 104705.7238 94472.34805 94926.00933 78686.96246 70604.48974 75848.06997 78438.58237 84374.61811 77568.8898 107404.1274 102856.6584 107107.2304 73856.31864 68642.82396 55931.18421 61093.66392 56876.56026 Cortolone 42895.65431 30540.84184 46354.81241 72490.51918 64383.60849 93009.69802 105980.8895 56130.95762 34811.27438 44480.3016 57818.51114 116505.5389 202795.7866 72273.04257 47627.05348 45474.88997 32446.15828 53373.23866 60669.58727 41744.17419 58931.78299 70426.52841 63781.80975 33570.34182 35311.70136 42384.24301 30684.06452 23493.88296 59785.36273 114630.4133 53131.83646 40717.52249 85670.06446 81912.38671 65064.41678 76814.37294 112774.9061 49768.8069 31826.72891 45300.10795 59137.94521 106045.3438 198751.3237 77079.44313 41369.04859 46359.67247 36120.56561 50624.70218 60669.58727 40692.69263 57200.30313 69795.42972 57339.42295 30539.60305 35313.3126 43202.79663 27831.29932 23798.06608 59338.11775 118401.4617 Tetrahydrocortisol 42895.65431 30540.84184 46354.81241 72490.51918 64383.60849 93009.69802 105980.8895 56130.95762 34811.27438 44480.3016 57818.51114 116505.5389 202795.7866 72273.04257 47627.05348 45474.88997 32446.15828 53373.23866 60669.58727 41744.17419 58931.78299 70426.52841 63781.80975 33570.34182 35311.70136 42384.24301 30684.06452 23493.88296 59785.36273 114630.4133 53131.83646 40717.52249 85670.06446 81912.38671 65064.41678 76814.37294 112774.9061 49768.8069 31826.72891 45300.10795 59137.94521 106045.3438 198751.3237 77079.44313 41369.04859 46359.67247 36120.56561 50624.70218 60669.58727 40692.69263 57200.30313 69795.42972 57339.42295 30539.60305 35313.3126 43202.79663 27831.29932 23798.06608 59338.11775 118401.4617 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name retention index quantified m/z PubChem ID KEGG ID 16a/16b-Hydroxyestrone 1 614.3514 115116 C05300 2-Methoxyestrone 1.43 342.2053 440624 C05299 21-Hydroxy-5b-pregnane-3,11,20-trione 1.75 410.2299 44263345 C05477 21-Deoxycortisol 1.75 410.2299 222803 C05497 Cortexolone 1.75 410.2299 53477676 C05488 Corticosterone 1.75 410.2299 5753 C02140 Pregnenolone 2.65 671.4385 53477678 C01953 5alpha-Dihydrodeoxycorticosterone 2.69 365.2688 6451493 C18040 3a-Hydroxy-5b-pregnane-20-one 3.23 675.4687 24779614 C05480 5alpha-Pregnan-20alpha-ol-3-one 3.23 675.4687 3080558 C18041 Allopregnanolone 3.23 675.4687 92786 C13712 17-Hydroxyprogesterone 4.52 372.2529 6238 C01176 20a,22b-Dihydroxycholesterol 3.44 436.378 53477897 C05501 Estrone 2.94 558.3542 5870 C00468 Tetrahydrodeoxycorticosterone 2.21 367.2833 101771 C13713 Calcitriol 3.18 480.3452 5280453 C01673 4a-Methylzymosterol-4-carboxylic acid 3.8 506.3611 53477909 C15808 4a-Carboxy-4b-methyl-5a-cholesta-8,24-dien-3b-ol 3.8 506.3611 23724596 C15808 Calcidiol 3.25 418.3668 5283731 C01561 (S)-2,3-Epoxysqualene 2.97 459.4205 53477723 C01054 Obtusifoliol 1.16 460.4145 656449 C01943 7-Dehydrodesmosterol 1.46 383.3304 C05107 Arachidonic acid 1.87 327.2312 444899 C00219 Androsterone glucuronide 1.99 489.2468 114833 C11135 Etiocholanolone glucuronide 1.99 489.2468 443078 C11136 Cortolone 4.28 796.5018 160499 C05481 Tetrahydrocortisol 4.28 796.5018 44725717 C05472 METABOLITES_END #END