#METABOLOMICS WORKBENCH michaelsa93_20160428_121518 DATATRACK_ID:611 STUDY_ID:ST000395 ANALYSIS_ID:AN000632 PROJECT_ID:PR000308 VERSION 1 CREATED_ON May 10, 2016, 12:37 pm #PROJECT PR:PROJECT_TITLE The circadian oscillator in Synechococcus elongatus controls metabolite PR:PROJECT_TITLE partitioning during diurnal growth PR:PROJECT_SUMMARY Cyanobacteria are increasingly being considered for use in large-scale outdoor PR:PROJECT_SUMMARY production of fuels and industrial chemicals. Cyanobacteria can anticipate daily PR:PROJECT_SUMMARY changes in light availability using an internal circadian clock and rapidly PR:PROJECT_SUMMARY alter their metabolic processes in response to changes light availability. PR:PROJECT_SUMMARY Understanding how signals from the internal circadian clock and external light PR:PROJECT_SUMMARY availability are integrated to control metabolic shifts will be important for PR:PROJECT_SUMMARY engineering cyanobacteria for production in natural outdoor environments. This PR:PROJECT_SUMMARY study has assessed how “knowing” the correct time of day, via the circadian PR:PROJECT_SUMMARY clock, affects metabolic changes when a cyanobacterium goes through a PR:PROJECT_SUMMARY dark-to-light transition. Our data show that the circadian clock plays an PR:PROJECT_SUMMARY important role in inhibiting activation of the oxidative pentose phosphate PR:PROJECT_SUMMARY pathway in the morning. Synechococcus elongatus PCC 7942 is a genetically PR:PROJECT_SUMMARY tractable model cyanobacterium that has been engineered to produce industrially PR:PROJECT_SUMMARY relevant biomolecules and is the best-studied model for a prokaryotic circadian PR:PROJECT_SUMMARY clock. However, the organism is commonly grown in continuous light in the PR:PROJECT_SUMMARY laboratory, and data on metabolic processes under diurnal conditions are PR:PROJECT_SUMMARY lacking. Moreover, the influence of the circadian clock on diurnal metabolism PR:PROJECT_SUMMARY has been investigated only briefly. Here, we demonstrate that the circadian PR:PROJECT_SUMMARY oscillator influences rhythms of metabolism during diurnal growth, even though PR:PROJECT_SUMMARY light–dark cycles can drive metabolic rhythms independently. Moreover, the PR:PROJECT_SUMMARY phenotype associated with loss of the core oscillator protein, KaiC, is distinct PR:PROJECT_SUMMARY from that caused by absence of the circadian output transcriptional regulator, PR:PROJECT_SUMMARY RpaA (regulator of phycobilisome-associated A). Although RpaA activity is PR:PROJECT_SUMMARY important for carbon degradation at night, KaiC is dispensable for those PR:PROJECT_SUMMARY processes. Untargeted metabolomics analysis and glycogen kinetics suggest that PR:PROJECT_SUMMARY functional KaiC is important for metabolite partitioning in the morning. PR:PROJECT_SUMMARY Additionally, output from the oscillator functions to inhibit RpaA activity in PR:PROJECT_SUMMARY the morning, and kaiC-null strains expressing a mutant KaiC phosphomimetic, PR:PROJECT_SUMMARY KaiC-pST, in which the oscillator is locked in the most active output state, PR:PROJECT_SUMMARY phenocopies a ΔrpaA strain. Inhibition of RpaA by the oscillator in the morning PR:PROJECT_SUMMARY suppresses metabolic processes that normally are active at night, and kaiC-null PR:PROJECT_SUMMARY strains show indications of oxidative pentose phosphate pathway activation as PR:PROJECT_SUMMARY well as increased abundance of primary metabolites. Inhibitory clock output may PR:PROJECT_SUMMARY serve to allow secondary metabolite biosynthesis in the morning, and some PR:PROJECT_SUMMARY metabolites resulting from these processes may feed back to reinforce clock PR:PROJECT_SUMMARY timing. PR:INSTITUTE UC Davis PR:DEPARTMENT Genome and Biomedical Sciences Facility PR:LABORATORY WCMC Metabolomics Core PR:LAST_NAME Fiehn PR:FIRST_NAME Oliver PR:ADDRESS Health Sciences Drive, Davis, California, 95616, USA PR:EMAIL ofiehn@ucdavis.edu PR:PHONE (530) 754-8258 PR:FUNDING_SOURCE NIH U24DK097154 PR:PUBLICATIONS doi: 10.1073/pnas.1504576112 #STUDY ST:STUDY_TITLE The circadian oscillator in Synechococcus elongatus controls metabolite ST:STUDY_TITLE partitioning during diurnal growth (part II) ST:STUDY_SUMMARY Cyanobacteria are increasingly being considered for use in large-scale outdoor ST:STUDY_SUMMARY production of fuels and industrial chemicals. Cyanobacteria can anticipate daily ST:STUDY_SUMMARY changes in light availability using an internal circadian clock and rapidly ST:STUDY_SUMMARY alter their metabolic processes in response to changes light availability. ST:STUDY_SUMMARY Understanding how signals from the internal circadian clock and external light ST:STUDY_SUMMARY availability are integrated to control metabolic shifts will be important for ST:STUDY_SUMMARY engineering cyanobacteria for production in natural outdoor environments. This ST:STUDY_SUMMARY study has assessed how “knowing” the correct time of day, via the circadian ST:STUDY_SUMMARY clock, affects metabolic changes when a cyanobacterium goes through a ST:STUDY_SUMMARY dark-to-light transition. Our data show that the circadian clock plays an ST:STUDY_SUMMARY important role in inhibiting activation of the oxidative pentose phosphate ST:STUDY_SUMMARY pathway in the morning. Synechococcus elongatus PCC 7942 is a genetically ST:STUDY_SUMMARY tractable model cyanobacterium that has been engineered to produce industrially ST:STUDY_SUMMARY relevant biomolecules and is the best-studied model for a prokaryotic circadian ST:STUDY_SUMMARY clock. However, the organism is commonly grown in continuous light in the ST:STUDY_SUMMARY laboratory, and data on metabolic processes under diurnal conditions are ST:STUDY_SUMMARY lacking. Moreover, the influence of the circadian clock on diurnal metabolism ST:STUDY_SUMMARY has been investigated only briefly. Here, we demonstrate that the circadian ST:STUDY_SUMMARY oscillator influences rhythms of metabolism during diurnal growth, even though ST:STUDY_SUMMARY light–dark cycles can drive metabolic rhythms independently. Moreover, the ST:STUDY_SUMMARY phenotype associated with loss of the core oscillator protein, KaiC, is distinct ST:STUDY_SUMMARY from that caused by absence of the circadian output transcriptional regulator, ST:STUDY_SUMMARY RpaA (regulator of phycobilisome-associated A). Although RpaA activity is ST:STUDY_SUMMARY important for carbon degradation at night, KaiC is dispensable for those ST:STUDY_SUMMARY processes. Untargeted metabolomics analysis and glycogen kinetics suggest that ST:STUDY_SUMMARY functional KaiC is important for metabolite partitioning in the morning. ST:STUDY_SUMMARY Additionally, output from the oscillator functions to inhibit RpaA activity in ST:STUDY_SUMMARY the morning, and kaiC-null strains expressing a mutant KaiC phosphomimetic, ST:STUDY_SUMMARY KaiC-pST, in which the oscillator is locked in the most active output state, ST:STUDY_SUMMARY phenocopies a ΔrpaA strain. Inhibition of RpaA by the oscillator in the morning ST:STUDY_SUMMARY suppresses metabolic processes that normally are active at night, and kaiC-null ST:STUDY_SUMMARY strains show indications of oxidative pentose phosphate pathway activation as ST:STUDY_SUMMARY well as increased abundance of primary metabolites. Inhibitory clock output may ST:STUDY_SUMMARY serve to allow secondary metabolite biosynthesis in the morning, and some ST:STUDY_SUMMARY metabolites resulting from these processes may feed back to reinforce clock ST:STUDY_SUMMARY timing. ST:INSTITUTE UC Davis ST:DEPARTMENT Genome and Biomedical Sciences Facility ST:LABORATORY WCMC Metabolomics Core ST:LAST_NAME Fiehn ST:FIRST_NAME Oliver ST:ADDRESS Health Sciences Drive, Davis, California, 95616, USA ST:EMAIL ofiehn@ucdavis.edu ST:PHONE (530) 754-8258 ST:STUDY_COMMENTS The first 4 samples were a test run to see how efficient the analysis was and ST:STUDY_COMMENTS were run on a lipidomics platform. The next 12 samples were the used in the ST:STUDY_COMMENTS paper and were the same as the original 4 samples, but they were split into 3 ST:STUDY_COMMENTS biological replicates and run on the GC platform. ST:PUBLICATIONS doi: 10.1073/pnas.1504576112 #SUBJECT SU:SUBJECT_TYPE Cells SU:SUBJECT_SPECIES Synechococcus elongatus PCC 7942 SU:TAXONOMY_ID 1140 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS WT_C_T0_005 140619dlvsa11_1 Genotype:WT | Time Point:- SUBJECT_SAMPLE_FACTORS WT_B_T4_004 140619dlvsa12_2 Genotype:WT | Time Point:4 SUBJECT_SAMPLE_FACTORS WT_B_T0_003 140619dlvsa03_1 Genotype:WT | Time Point:- SUBJECT_SAMPLE_FACTORS WT_C_T4_006 140619dlvsa02_1 Genotype:WT | Time Point:4 SUBJECT_SAMPLE_FACTORS WT_A_T0_001 140619dlvsa05_1 Genotype:WT | Time Point:- SUBJECT_SAMPLE_FACTORS WT_A_T4_002 140619dlvsa04_1 Genotype:WT | Time Point:4 SUBJECT_SAMPLE_FACTORS KaiC_B T0_009 140619dlvsa01_1 Genotype:KaiC mutant | Time Point:- SUBJECT_SAMPLE_FACTORS KaiC_C T0_011 140619dlvsa06_2 Genotype:KaiC mutant | Time Point:- SUBJECT_SAMPLE_FACTORS KaiC_A T0_007 140619dlvsa07_2 Genotype:KaiC mutant | Time Point:- SUBJECT_SAMPLE_FACTORS KaiC_A T4_008 140619dlvsa08_3 Genotype:KaiC mutant | Time Point:4 SUBJECT_SAMPLE_FACTORS KaiC_B T4_010 140619dlvsa09_2 Genotype:KaiC mutant | Time Point:4 SUBJECT_SAMPLE_FACTORS KaiC_C T4_012 140619dlvsa10_2 Genotype:KaiC mutant | Time Point:4 #COLLECTION CO:COLLECTION_SUMMARY Bacteria were grown in a turbidostat/bioreactor at equal cell density (measured CO:COLLECTION_SUMMARY by optical density at 750nm), under a 12:12h Light/Dark cycle. After collection CO:COLLECTION_SUMMARY samples were immediately placed on ice and then centrifuged at 5000RPM for 10min CO:COLLECTION_SUMMARY at 4 degrees Celsius. After centrifugation supernatant was decanted and cell CO:COLLECTION_SUMMARY pellets were immediately frozen in liquid N2. CO:COLLECTION_PROTOCOL_FILENAME StudyDesign-SpencerDiamond-10814.pdF CO:COLLECTION_TIME Samples were collected at T0 (beginning of day) and T4 (4h into day). CO:VOLUMEORAMOUNT_COLLECTED 40ml of sample was collected at each time point CO:STORAGE_CONDITIONS Samples were put into a 50mL conical tube containing ice up to the 30ml mark. #TREATMENT TR:TREATMENT_SUMMARY 2: WT bacteria and KaiC mutant The phenotype associated with loss of the core TR:TREATMENT_SUMMARY oscillator protein, KaiC, is distinct from that caused by absence of the TR:TREATMENT_SUMMARY circadian output transcriptional regulator, RpaA (regulator of TR:TREATMENT_SUMMARY phycobilisome-associated A). Untargeted metabolomics analysis and glycogen TR:TREATMENT_SUMMARY kinetics suggest that functional KaiC is important for metabolite partitioning TR:TREATMENT_SUMMARY in the morning. Additionally, output from the oscillator functions to inhibit TR:TREATMENT_SUMMARY RpaA activity in the morning, and kaiC-null strains expressing a mutant KaiC TR:TREATMENT_SUMMARY phosphomimetic, KaiC-pST, in which the oscillator is locked in the most active TR:TREATMENT_SUMMARY output state, phenocopies a ΔrpaA strain. KaiC-null strains show indications of TR:TREATMENT_SUMMARY oxidative pentose phosphate pathway activation as well as increased abundance of TR:TREATMENT_SUMMARY primary metabolites. Inhibitory clock output may serve to allow secondary TR:TREATMENT_SUMMARY metabolite biosynthesis in the morning, and some metabolites resulting from TR:TREATMENT_SUMMARY these processes may feed back to reinforce clock timing. TR:TREATMENT_PROTOCOL_FILENAME StudyDesign-SpencerDiamond-10814.pdf #SAMPLEPREP SP:SAMPLEPREP_SUMMARY 1. Add 0.5mL of extraction solvent to tube, gently pipet to remove all cells, SP:SAMPLEPREP_SUMMARY transfer cells to 2mL eppendorf tube. Repeat for a total of 1mL extraction SP:SAMPLEPREP_SUMMARY solvent + cells in 2mL eppendorf tube. 2. Add 2 small stainless steel grinding SP:SAMPLEPREP_SUMMARY beads to eppendorf tube 3. Use the GenoGrinder to grind for 3 minutes at 1,250 SP:SAMPLEPREP_SUMMARY rpm. 4. Centrifuge at 14,000xg for 5 minutes. 5. Transfer supernatant to a fresh SP:SAMPLEPREP_SUMMARY 2mL eppendorf tube. 6. Add 1mL of extraction solvent to tube containing cell SP:SAMPLEPREP_SUMMARY pellet + beads, and repeat steps 3 and 4. 7. Collect supernatant, and combine SP:SAMPLEPREP_SUMMARY with supernatant collected in step 5. Total volume of extracted sample will be SP:SAMPLEPREP_SUMMARY approximately 2mL. 8. Dry down 50uL of extracted sample in 1.5mL eppendorf tube SP:SAMPLEPREP_SUMMARY for GC-TOF analysis. 9. Store backups in -20 or -80C. SP:SAMPLEPREP_PROTOCOL_FILENAME SOP_Extraction_of_Yeast_Cells.pdf #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Leco Pegasus III GC CH:COLUMN_NAME Restek Rtx-5Sil MS (30 x 0.25mm, 0.25um) CH:COLUMN_NAME 0.25 μm film made of 95% dimethyl/5%diphenylpolysiloxane) CH:FLOW_RATE 1 ml/min CH:COLUMN_TEMPERATURE 50-330C CH:METHODS_FILENAME SOP_Extraction_of_Yeast_Cells.pdf CH:COLUMN_PRESSURE 7.7 PSI CH:INJECTION_TEMPERATURE 50 C ramped to 250 C by 12 C/s CH:SAMPLE_INJECTION 0.5 uL CH:OVEN_TEMPERATURE 50°C for 1 min, then ramped at 20°C/min to 330°C, held constant for 5 min CH:TRANSFERLINE_TEMPERATURE 230C CH:WASHING_BUFFER Ethyl Acetate CH:SAMPLE_LOOP_SIZE 30 m length x 0.25 mm internal diameter CH:RANDOMIZATION_ORDER Excel generated #ANALYSIS AN:ANALYSIS_TYPE MS AN:LABORATORY_NAME WCMC Metabolomics Core AN:DETECTOR_TYPE TOF AN:SOFTWARE_VERSION ChromaTOF AN:DATA_FORMAT .peg, .txt, .cdf #MS MS:MS_COMMENTS - MS:INSTRUMENT_NAME Leco Pegasus III GC TOF MS:INSTRUMENT_TYPE GC-TOF MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:ION_SOURCE_TEMPERATURE 250 C MS:IONIZATION_ENERGY 70 eV MS:SOURCE_TEMPERATURE 250 C MS:SCAN_RANGE_MOVERZ 85-500 Da MS:SCANNING_CYCLE 17 Hz MS:SCANNING_RANGE 85-500 Da MS:SKIMMER_VOLTAGE 1850 V #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS counts MS_METABOLITE_DATA_START 140619dlvsa11_1 140619dlvsa12_2 140619dlvsa03_1 140619dlvsa02_1 140619dlvsa05_1 140619dlvsa04_1 140619dlvsa01_1 140619dlvsa06_2 140619dlvsa07_2 140619dlvsa08_3 140619dlvsa09_2 140619dlvsa10_2 xylulose NIST 600 529 610 255 436 557 248 576 789 506 518 304 xanthosine 728 983 1090 992 723 1061 2208 1007 2048 1700 1135 1227 valine 2702 10267 4963 12700 4824 8305 9451 4708 7627 11747 17091 18313 uridine 14445 2383 14979 8100 8397 9238 18688 8382 8759 12112 7864 6355 urea 831 8730 6135 18187 7405 5890 2612 1147 3953 3108 5590 4213 uracil 3286 3239 2787 1895 3477 1943 2419 2591 2075 3743 2158 2293 UDP-N-acetylglucosamine 183 273 357 244 292 271 209 149 214 574 392 260 tyrosine 5302 7129 3337 9678 3702 7642 7982 6601 4697 10808 11169 11778 tryptophan 5004 4224 1880 5589 1982 6936 17133 26374 13675 18752 20812 35579 trans-4-hydroxy-L-proline 1087 1794 3100 1515 1936 1311 1182 1222 993 1328 927 563 thymine 6932 6863 6637 7893 6342 5279 10041 6598 6224 8504 4977 5226 thymidine-5-phosphate 2033 690 1121 435 711 427 2175 1496 653 1863 870 510 thymidine 7077 3279 4823 5598 4149 3864 5080 3771 3658 4830 2745 2400 threonine 1947 3489 2039 4239 2378 2956 4049 3035 2673 5111 5050 4875 threitol 1530 2659 2276 2425 2878 2213 1392 1083 1703 1995 1622 1375 sulfuric acid 10797 206 3833 167 921 1289 4268 8479 405 6753 325 279 sucrose 560 6345 448 6104 482 8370 2578 1323 2274 52732 46288 34140 stearic acid 269226 272593 206216 137203 287227 221562 171345 234654 225601 128578 148689 161548 spermidine 361 323 339 448 307 644 350 635 1541 709 1282 431 sorbitol 837 997 1147 515 619 363 777 532 666 763 1249 1569 serine 1899 4959 2732 11261 2233 3267 18106 2650 3578 6696 6211 5869 salicylaldehyde 1878 3241 2536 2396 1312 1755 1601 2899 2207 2177 3116 3289 ribose-5-phosphate 1454 1675 927 1660 1096 1769 876 1102 1343 1999 2212 1586 ribose 1527 3151 183 3212 1675 2919 2051 2280 2391 2577 1552 2957 ribonic acid 302 267 456 266 523 371 308 211 432 349 338 192 pyrophosphate 5172 5810 5154 3060 6758 11852 3980 2383 2219 14410 6445 3906 putrescine 1175 1003 1669 1955 3304 1104 828 736 2492 1101 2425 1456 propane-1,3-diol NIST 921 1648 2002 1568 1249 2045 1045 754 1562 1060 942 820 proline 357 768 1249 1348 2002 1094 1235 644 589 508 940 645 pimelic acid 1011 916 1064 1477 1799 1471 544 1174 982 912 708 860 phytol 59536 38022 73314 48760 59282 37398 73959 61141 64258 55688 42874 41752 phosphate 20719 56096 22398 52082 40430 27431 46372 70315 77209 28668 30762 22037 phenylalanine 657 706 593 1104 1221 764 798 181 627 217 804 757 pentadecanoic acid 2526 4094 3174 1917 3685 636 4317 3438 2667 4971 4617 4379 pelargonic acid 11584 13503 17783 10600 19162 13311 11141 8774 11703 14504 11106 10300 p-cresol 121793 117460 179180 103911 154086 117275 115565 76650 93378 104627 80404 65090 parabanic acid NIST 606 2504 4158 2027 3581 2585 2449 992 2623 2107 2513 3967 palmitoleic acid 162613 109167 212559 111336 165247 115270 134638 147522 115478 83128 95164 108998 palmitic acid 69867 69685 88856 50660 81753 62737 54306 55043 50285 43365 47490 50949 oxoproline 30097 224799 154997 332432 163547 180197 228608 91560 182136 249007 430621 454503 oxalic acid 285 320 499 326 1015 510 866 1181 125 1023 734 444 ornithine 632 1270 816 2562 1020 796 1223 1242 884 1363 791 569 oleic acid 9704 20202 28239 8995 8877 15184 20362 23994 20160 18891 24639 17277 oleamide NIST 854 1496 1457 464 2078 613 2665 1358 509 1976 1004 876 octanol NIST 655 824 1027 778 1203 818 1055 585 784 430 495 499 octadecanol 1255 643 1067 631 442 468 832 709 562 666 917 664 nicotinic acid 1122 829 727 948 718 773 771 471 418 697 853 507 nicotinamide 3495 4901 4840 5552 7412 4160 3172 2606 3804 2312 3438 3257 N-acetylglutamate 5883 5189 2818 6532 4979 1875 8512 6762 2717 15681 1630 1414 N-acetyl-D-mannosamine 319 269 334 360 254 282 230 179 334 556 394 303 myristic acid 17265 16175 21092 13423 21443 14713 18025 18356 14456 13430 14555 15027 methionine 548 499 653 1014 711 590 860 159 836 414 1328 1088 mannose-6-phosphate NIST 218 85 325 143 327 481 85 163 139 258 72 199 maltotriose 8583 3651 7638 3617 2611 3219 5444 8037 6416 3987 4517 6450 maltotriitol 392 428 394 334 343 145 536 285 393 226 306 215 maltose 1928 1633 2809 1201 2071 2066 1992 1138 1860 1421 1540 1831 maltitol 371 423 197 343 551 381 542 576 777 578 314 420 maleimide 1989 5579 9560 12183 8377 6611 12549 3696 5095 2319 3250 3858 lyxitol 287 560 625 671 556 522 461 193 386 808 499 449 lysopalmitoyl monogalactosylglycerol 172130 67682 140348 51913 49543 59804 71945 95024 69761 38142 43850 57936 lysine 3101 1089 1460 2550 3728 893 11105 14012 13102 2228 1548 2243 levoglucosan 390 706 562 981 980 779 812 404 673 951 744 653 leucine 894 4253 1934 5628 2071 2517 3309 1208 2676 2370 5516 5858 lauric acid 5707 7384 8491 6183 6913 6704 6093 6433 5470 6080 7339 7291 lactic acid 4356 9183 7068 22937 5545 8312 5156 2943 3674 3331 4947 3713 isothreonic acid 382 441 1084 657 980 895 528 594 452 590 595 690 isoleucine 468 2627 1897 4123 1840 1881 2274 764 1937 2107 4733 5006 inulotriose 245 296 422 411 513 408 965 1090 1376 1123 1043 979 hydroxylamine 24462 27246 39602 24253 34690 27141 25049 17648 22509 22727 17874 14798 homoserine 229 443 508 574 543 920 518 172 578 236 598 645 hexose-6-phosphate 311 607 277 607 409 638 359 320 446 726 579 862 heptadecanoic acid 3972 7165 3990 3269 5385 4732 4498 4389 4472 4328 4447 4910 guanosine 4631 7622 4304 4886 4926 5946 12780 11398 10644 11733 9812 7871 guanine 801 869 1041 868 1172 793 679 873 1376 662 667 905 glycolic acid 3153 3458 6195 3595 5464 3495 3094 2750 2690 2743 2624 2991 glycine 168 956 627 2087 736 719 1793 231 414 738 979 929 glycerol-alpha-phosphate 74147 41016 84178 45579 65878 53711 74442 64780 50526 55358 29973 44197 glycerol-3-galactoside 173238 72576 119191 90669 59871 67625 72265 120430 59618 33289 40307 70391 glycerol 49916 26839 20302 79922 106729 77020 16904 54056 85860 87034 17521 59295 glyceric acid 1418 1794 2245 1930 1941 2190 1797 1680 1518 2007 1695 2024 glutaric acid 344 361 593 459 188 416 260 318 212 355 216 344 glutamine 415 1362 804 934 1591 1175 1442 2258 2064 3270 2938 3108 glutamic acid 5864 5660 10707 5947 14825 15722 6385 5224 6254 23258 12228 10720 glucose-6-phosphate 833 1987 913 1372 637 1610 1096 1167 1130 5320 4290 4089 glucose-1-phosphate 2681 5725 3779 4200 2444 8548 5831 5184 4237 10681 6885 3394 glucose 63663 156855 99320 120410 71742 152854 145613 121555 131305 198026 217635 151712 gluconic acid 384 1001 662 714 835 2066 487 957 184 545 834 714 galactinol 279244 208492 129385 253529 148816 298446 129330 163934 117937 125726 134059 132784 fumaric acid 699 1010 1192 367 480 307 296 459 395 586 549 538 fucose 2434 2988 2667 3061 3687 4603 3501 2383 2942 3501 4105 2655 fructose-6-phosphate 443 1019 388 774 675 748 878 456 503 2751 2027 2036 fructose 1945 3223 3212 3034 1345 5011 5237 3808 2714 3387 3654 3965 erythrose 336 482 556 440 391 938 417 473 662 666 798 414 erythronic acid 436 1543 582 853 855 1264 900 822 1135 1162 938 691 enolpyruvate NIST 329 847 585 380 941 375 413 251 304 570 412 392 cytosin 191 806 545 1005 1084 891 284 218 644 385 644 737 cytidine 772 697 559 495 713 795 1710 1756 2198 1659 1557 990 citric acid 3965 18787 6064 8432 50809 16851 5749 5551 12676 14104 5598 3655 capric acid 1536 2683 2661 2201 3644 1943 1716 1514 1587 2175 1712 1980 butyrolactam NIST 531 1391 1295 1016 934 686 1132 578 698 597 494 701 beta-glycerolphosphate 726 1261 1209 679 1472 997 963 892 575 496 417 849 beta-alanine 250 927 194 253 457 273 173 193 359 275 229 565 benzoic acid 17706 17741 23499 16303 24095 20354 14335 12444 14347 15105 12995 11713 behenic acid 8632 21674 7259 18952 16244 22876 38877 19151 26415 42307 33262 27771 azelaic acid 1015 1281 670 1051 1289 1525 1315 1459 289 2046 656 1429 aspartic acid 1055 4356 4290 3481 5837 2070 5523 3365 4574 2565 4333 5081 asparagine 338 213 431 262 355 85 211 152 95 203 255 204 arachidic acid 37590 3543 4809 3241 3905 6656 7171 70912 84134 9199 6134 5833 alpha-ketoglutarate 950 2302 1489 2588 1200 1241 987 485 707 1736 1355 1557 alanine-alanine 248 475 382 253 398 274 191 145 362 252 258 421 alanine 8548 11958 11768 17066 10382 10680 24151 12244 13738 27389 21028 21827 adipic acid 2125 3189 3753 3316 3690 2894 2171 1666 2367 2739 1942 1837 adenosine 111755 106015 36060 95007 81017 111389 189643 200223 213457 212043 164373 137716 adenine 12060 24250 20393 25507 27785 19963 14974 14605 19351 16462 16172 14691 acetophenone NIST 3760 2981 4589 3322 6367 3760 4862 2306 1910 2222 2147 1681 6-deoxyglucose 4337 5322 5028 6112 6613 5391 3640 4994 3731 5656 5222 4284 5-hydroxymethyl-2-furoic acid NIST 359 17156 16882 240 525 309 161 240 257 566 214 299 5'-deoxy-5'-methylthioadenosine 218 461 299 350 393 253 2004 1657 1785 1868 899 501 4-hydroxybenzoate 638 938 710 1058 1583 651 363 533 852 445 652 283 3-phosphoglycerate 2333 3283 4318 3056 11633 6429 3718 4312 4153 9115 2984 2052 3-hydroxypropionic acid 908 974 690 1044 708 986 532 645 698 797 361 603 3-hydroxypalmitic acid 678 585 1124 446 421 495 1916 1905 1800 1374 1421 1504 2-ketoadipic acid 1582 1787 2687 1864 2187 1757 1182 1021 1525 1302 1148 1092 2-deoxytetronic acid 172 383 297 321 188 553 421 319 443 488 288 321 2'-deoxyguanosine 1064 741 368 462 678 590 743 4546 634 679 6785 589 2,3-dihydroxypyridine 394 614 482 422 272 597 828 486 543 324 463 349 1-monopalmitin 871 860 779 585 500 597 636 435 470 572 317 528 1-hexadecanol 602 446 847 627 574 448 383 342 596 478 304 325 1,2,4-benzenetriol 254 1019 3557 1095 3636 2459 520 462 530 340 647 596 2,5-dihydroxypyrazine NIST 724 647 727 499 990 1494 1271 1297 711 2409 494 794 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name retention index quantified mz KEGG ID PubChem ID xylulose NIST 553450 173 C00312 439205 xanthosine 926133 325 C01762 64959 valine 313502 144 C00183 6287 uridine 861508 217 C00299 6029 urea 326574 189 C00086 1176 uracil 385816 241 C00106 1174 UDP-N-acetylglucosamine 623789 226 C00043 445675 tyrosine 671252 218 C00082 6057 tryptophan 780482 202 C00078 6305 trans-4-hydroxy-L-proline 457910 158 C01157 5810 thymine 420133 255 C00178 1135 thymidine-5-phosphate 608111 170 C00364 9700 thymidine 349402 170 C00214 5789 threonine 409568 218 C00188 6288 threitol 467595 217 C16884 169019 sulfuric acid 282411 227 C00059 1118 sucrose 915139 271 C00089 5988 stearic acid 787622 117 C01530 5281 spermidine 792924 144 C00315 1102 sorbitol 667922 217 C00794 5780 serine 395020 218 C00065 5951 salicylaldehyde 406586 119 C06202 6998 ribose-5-phosphate 731096 315 C00117 439167 ribose 553135 217 C00121 5779 ribonic acid 599680 292 C01685 5460677 pyrophosphate 326948 110 C00013 1023 putrescine 588119 174 C00138 1049 propane-1,3-diol NIST 214380 177 C02457 10442 proline 364716 142 C00148 145742 pimelic acid 523205 155 C02656 385 phytol 761712 143 C01389 5280435 phosphate 345365 314 C00009 1004 phenylalanine 537804 218 C00079 6140 pentadecanoic acid 674647 117 C16537 13849 pelargonic acid 399229 117 C01601 8158 p-cresol 280360 165 C01468 2879 parabanic acid NIST 464991 100 C00802 67126 palmitoleic acid 706508 117 C08362 445638 palmitic acid 713809 313 C00249 985 oxoproline 485935 156 C01879 7405 oxalic acid 260513 190 C00209 971 ornithine 619548 142 C00077 6262 oleic acid 779120 339 C00712 445639 oleamide NIST 849710 144 C19670 5283387 octanol NIST 247010 187 octadecanol 755409 327 8221 nicotinic acid 366992 180 C00253 938 nicotinamide 471602 179 C00153 936 N-acetylglutamate 604748 216 C00624 70914 N-acetyl-D-mannosamine 726375 319 myristic acid 634414 117 C06424 11005 methionine 483560 176 C00073 6137 mannose-6-phosphate NIST 822643 387 C00275 65127 maltotriose 1179224 361 C01835 439586 maltotriitol 1214242 361 3625615 maltose 946601 204 C00208 439186 maltitol 976828 361 D04845 493591 maleimide 245118 154 C07272 10935 lyxitol 573587 217 C00532 439255 lysopalmitoyl monogalactosylglycerol 1183271 204 53483072 lysine 663483 156 C00047 5962 levoglucosan 569637 204 2724705 leucine 346101 158 C00123 6106 lauric acid 547906 117 C02679 3893 lactic acid 217657 191 C01432 612 isothreonic acid 489385 292 C00639 151152 isoleucine 359251 158 C00407 6306 inulotriose 1131821 361 C01355 22833608 hydroxylamine 255241 146 C00192 787 homoserine 443878 218 C00263 12647 hexose-6-phosphate 806282 387 C02965 208 heptadecanoic acid 751309 117 10465 guanosine 954962 324 C00387 6802 guanine 744307 352 C00242 764 glycolic acid 227636 177 C00160 757 glycine 368707 248 C00037 750 glycerol-alpha-phosphate 590747 357 C03189 754 glycerol-3-galactoside 805227 204 C05401 16048618 glycerol 344466 205 C00116 753 glyceric acid 377495 189 C00258 439194 glutaric acid 421596 261 C00489 743 glutamine 600315 156 C00064 5961 glutamic acid 529100 246 C00025 33032 glucose-6-phosphate 810287 387 C01172 439427 glucose-1-phosphate 594647 217 C00103 65533 glucose 650867 319 C00221 64689 gluconic acid 693148 333 C00800 6857417 galactinol 1048207 204 C01235 11727586 fumaric acid 390775 245 C00122 444972 fucose 578299 160 C02095 439650 fructose-6-phosphate 804279 315 C05345 440641 fructose 639442 307 C02336 439709 erythrose 443306 205 C01796 439574 erythronic acid 512029 217 2781043 enolpyruvate NIST 234394 217 C00074 1005 cytosin 486724 254 C00380 597 cytidine 937026 223 C00475 6175 citric acid 617342 273 C00158 311 capric acid 452386 229 C01571 2969 butyrolactam NIST 277199 142 12025 beta-glycerolphosphate 574470 243 C02979 2526 beta-alanine 435564 248 C00099 239 benzoic acid 339214 179 C00180 243 behenic acid 920648 117 C08281 8215 azelaic acid 610651 317 C08261 2266 aspartic acid 480387 232 C00049 5960 asparagine 553078 188 C00152 6267 arachidic acid 856421 117 C06425 10467 alpha-ketoglutarate 507392 198 C00026 51 alanine-alanine 636898 188 C00993 5484352 alanine 243971 116 C00041 5950 adipic acid 474435 111 C06104 196 adenosine 918039 236 C00212 60961 adenine 646534 264 C00147 190 acetophenone NIST 238615 105 C07113 7410 6-deoxyglucose 573663 117 C08352 441480 5-hydroxymethyl-2-furoic acid NIST 497561 123 C20448 80642 5'-deoxy-5'-methylthioadenosine 967036 236 C00170 439176 4-hydroxybenzoate 537925 223 C00156 135 3-phosphoglycerate 610734 227 C00597 724 3-hydroxypropionic acid 269265 177 C01013 68152 3-hydroxypalmitic acid 774930 233 15569776 2-ketoadipic acid 331841 100 2-deoxytetronic acid 433456 189 150929 2'-deoxyguanosine 962182 295 C00330 187790 2,3-dihydroxypyridine 373895 240 28115 1-monopalmitin 901749 129 C01885 14900 1-hexadecanol 679596 299 C00823 2682 1,2,4-benzenetriol 521803 239 C02814 10787 2,5-dihydroxypyrazine NIST 397526 241 23368901 METABOLITES_END #END