#METABOLOMICS WORKBENCH chalermrojCS_20231106_091704 DATATRACK_ID:4443 STUDY_ID:ST002972 ANALYSIS_ID:AN004881 PROJECT_ID:PR001849 VERSION 1 CREATED_ON November 6, 2023, 9:38 am #PROJECT PR:PROJECT_TITLE Alkane production PR:PROJECT_SUMMARY Part 3/3 of XR/lactose system demonstrated in E. coli PR:INSTITUTE Vidyasirimedhi Institute of Science and Technology PR:LAST_NAME Sutthaphirom PR:FIRST_NAME Chalermroj PR:ADDRESS 555, Moo 1, Payubnai, Wangchan, Rayong, 21210, Thailand PR:EMAIL chalermroj.s_s19@vistec.ac.th PR:PHONE (+66)063-9639915 #STUDY ST:STUDY_TITLE Alkane T5h ST:STUDY_SUMMARY Untargeted metabolomics of alkane producing strains at time 5 h of bioconversion ST:INSTITUTE Vidyasirimedhi Institute of Science and Technology ST:LAST_NAME Sutthaphirom ST:FIRST_NAME Chalermroj ST:ADDRESS 555, Moo 1, Payubnai, Wangchan, Rayong, 21210, Thailand ST:EMAIL chalermroj.s_s19@vistec.ac.th ST:PHONE (+66)063-9639915 ST:NUM_GROUPS 2 ST:TOTAL_SUBJECTS 8 #SUBJECT SU:SUBJECT_TYPE Bacteria SU:SUBJECT_SPECIES Escherichia coli SU:TAXONOMY_ID 562 SU:GENOTYPE_STRAIN BL21 (DE3) SU:CELL_BIOSOURCE_OR_SUPPLIER Novagen #FACTORS #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 - FAP-1 Strain:FAP SUBJECT_SAMPLE_FACTORS - FAP-2 Strain:FAP SUBJECT_SAMPLE_FACTORS - FAP-3 Strain:FAP SUBJECT_SAMPLE_FACTORS - FAP-4 Strain:FAP SUBJECT_SAMPLE_FACTORS - FAP-XR-1 Strain:XR SUBJECT_SAMPLE_FACTORS - FAP-XR-2 Strain:XR SUBJECT_SAMPLE_FACTORS - FAP-XR-3 Strain:XR SUBJECT_SAMPLE_FACTORS - FAP-XR-4 Strain:XR #COLLECTION CO:COLLECTION_SUMMARY Samples were quenched by cold methanol and were extracted metabolites by cold CO:COLLECTION_SUMMARY methanol method. CO:SAMPLE_TYPE Bacterial cells #TREATMENT TR:TREATMENT_SUMMARY Cells were incubated with 10 mM lactose and 5 mM myristic acid in Kpi buffer TR:TREATMENT_SUMMARY 25˚C for 5 h in the presence of blue light #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Intracellular metabolites were extracted by cold methanol method. The SP:SAMPLEPREP_SUMMARY supernatants were lyophilized and reconstituted in 50% acetonitrile/water. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME Agilent 6560 CH:COLUMN_NAME Agilent Poroshell 120 HILIC-Z (150 x 2.1 mm, 2.7 µm) CH:SOLVENT_A 10 mM ammonium acetate pH 9.0 CH:SOLVENT_B 10 mM ammonium acetate in 85% acetonitrile/water pH 9.0 CH:FLOW_GRADIENT 0 min: 96%B, 24 min: 65%B CH:FLOW_RATE 0.3 mL/min CH:COLUMN_TEMPERATURE 35˚C #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Agilent 6560 Ion Mobility MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE NEGATIVE MS:MS_COMMENTS Raw MS data files (.d) were first demultiplexed using the Agilent MS:MS_COMMENTS De-multiplexing tool (Version 1.0, Agilent Technologies). The demultiplexing MS:MS_COMMENTS data files (.DeMP.d) were recalibrated using IM–MS Reprocessor (Version MS:MS_COMMENTS B.08.00, Agilent Technologies). The reference masses used for mass calibration MS:MS_COMMENTS were m/z 112.985587 and 1033.988109. The CCS calibration was performed by the MS:MS_COMMENTS IM–MS Browser software (Version 10.0, Agilent Technologies). #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS abundance MS_METABOLITE_DATA_START Samples FAP-1 FAP-2 FAP-3 FAP-4 FAP-XR-1 FAP-XR-2 FAP-XR-3 FAP-XR-4 Factors Strain:FAP Strain:FAP Strain:FAP Strain:FAP Strain:XR Strain:XR Strain:XR Strain:XR L-Leucine 3958357 3562755 3366315 3186897 2172196 1831470 1838936 1659805 L-Phenylalanine 2714176 2563518 2492852 2507151 1358460 1324597 1194617 1069606 L-Isoleucine 2614910 2702100 2482585 2368407 1289845 1462603 1224505 1016201 Acetyl-CoA 1012824 977655 896619 700178 1497521 1288791 1065546 985346 ADP-ribose 2'-phosphate 1312747 1240972 1147114 934475 719396 729100 617181 518163 Pantoic acid 979618 909809 845671 818859 246638 1 201619 161920 α-D-Glucose 1199755 1098072 1014061 1 1 1 1 1 FAD / Flavin adenine dinucleotide 338741 379179 329290 302712 889651 850782 749189 651551 UDP-N-acetyl-D-galactosaminuronic acid 454574 451214 293548 300425 957342 941535 666915 515542 L-Tyrosine 666380 643661 586096 544237 1 1 1 1 L-Valine 521499 566374 496190 444294 252871 1 1 237509 D-4'-Phosphopantothenate 441513 395056 1 265421 543015 524062 477636 418345 Stearic acid 154664 1 1 147608 1 1 1 1 UDP-N-acetylmuramoyl-L-alanyl-D-glutamate 253636 308018 235355 213103 1 1 1 1 5-Amino-6-ribitylamino uracil 240052 241409 224852 247351 59548 82577 57070 86441 Palmitic Acid 158627 1 1 1 309740 1 198142 1 ADP / Adenosine 5'-diphosphate 248698 1 166684 186367 1 268965 1 1 D-Ala-D-Ala 164501 156597 112319 1 1 1 1 1 UDP-N-acetylmuramoyl-L-alanyl-D-glutamyl-6-carboxy-L-lysyl-D-alanyl- D-alanine 228557 1 1 1 166982 1 1 109302 UMP / Uridine 5-monophosphate 73647 34222 62416 1 1 1 1 1 S-Acetylphosphopantetheine 69222 71015 1 27066 1 1 1 1 NAD 30792 1 49493 61746 1 18376 1 1 Cyclic adenosine diphosphate ribose 35175 1 1 1 1 12284 1 1 Phosphoglycolic acid 17598 1 1 1 1 1 1 1 D-Sorbitol 1 1 1 1 1 8335072 7408288 55580 1,5-Anhydrosorbitol 1 1 1 1 995398 982967 782710 699523 Uridine 1 660960 640859 581906 1 309016 1 1 2,3-Bis(3-hydroxytetradecanoyl)-β-D-glucosaminyl 1-phosphate 1 33702 1 1 124283 128353 158983 182950 Guanosine 1 1 72060 1 1 90594 81135 68532 Xanthine 1 1 1 19083 1 23120 1 1 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name Mass Retention Time DT CCS Ion Species L-Leucine 131.095 3.1989975 14.278361 130.31259 [M-H]- L-Phenylalanine 165.0796 2.9893622 15.532857 139.2205 [M-H]- L-Isoleucine 131.095 3.382506 14.163478 129.25629 [M-H]- Acetyl-CoA 809.1225 10.117327 16.891247 284.8186 [M-2H]-2 ADP-ribose 2'-phosphate 639.033 9.247358 24.769444 210.17612 [M-H]- Pantoic acid 148.0738 2.331515 16.225317 143.24977 [M+CH3COO]- α-D-Glucose 180.0636 4.549526 14.809285 131.87872 [M-H]- FAD / Flavin adenine dinucleotide 785.1511 6.3940244 28.050545 237.20181 [M-H]- UDP-N-acetyl-D-galactosaminuronic acid 621.0564 9.658229 26.47034 224.8075 [M-H]- L-Tyrosine 181.0741 3.6735678 16.140684 143.76567 [M-H]- L-Valine 117.079 3.9396446 13.467975 124.16849 [M-H]- D-4'-Phosphopantothenate 299.0762 11.234335 17.944178 155.61594 [M-H]- Stearic acid 284.2714 2.531477 20.262714 176.2362 [M-H]- UDP-N-acetylmuramoyl-L-alanyl-D-glutamate 879.1753 12.712223 31.36425 264.839 [M-H]- 5-Amino-6-ribitylamino uracil 276.111 2.4413347 18.964638 165.1068 [M-H]- Palmitic Acid 256.2399 2.1871698 19.324286 168.85948 [M-H]- ADP / Adenosine 5'-diphosphate 427.0271 10.661384 21.354164 182.95694 [M-H]- D-Ala-D-Ala 160.085 5.47627 15.1719055 136.274 [M-H]- UDP-N-acetylmuramoyl-L-alanyl-D-glutamyl-6-carboxy-L-lysyl-D-alanyl- D-alanine 1193.3324 14.361842 20.2985 340.77536 [M-2H]-2 UMP / Uridine 5-monophosphate 324.0343 8.529928 18.618465 160.96141 [M-H]- S-Acetylphosphopantetheine 400.1047 5.0447764 21.28664 182.75458 [M-H]- NAD 663.1037 9.523409 26.231596 222.46588 [M-H]- Cyclic adenosine diphosphate ribose 541.0584 9.484358 23.524673 200.32837 [M-H]- Phosphoglycolic acid 155.9818 10.293 14.877218 133.87306 [M-H]- D-Sorbitol 182.0795 3.7508342 14.6867485 130.68117 [M-H]- 1,5-Anhydrosorbitol 164.0687 3.5147104 14.552673 130.43387 [M-H]- Uridine 244.0695 2.5915473 16.894127 147.87216 [M-H]- 2,3-Bis(3-hydroxytetradecanoyl)-β-D-glucosaminyl 1-phosphate 711.4269 2.4950898 31.277746 265.06805 [M-H]- Guanosine 283.0912 2.8657444 18.584377 161.5926 [M-H]- Xanthine 152.0334 3.377973 13.355713 120.32379 [M-H]- METABOLITES_END #END