#METABOLOMICS WORKBENCH ppjutur_20200204_061726 DATATRACK_ID:1916 STUDY_ID:ST001316 ANALYSIS_ID:AN002191 PROJECT_ID:PR000893 VERSION 1 CREATED_ON February 10, 2020, 12:28 pm #PROJECT PR:PROJECT_TITLE Carbon Partitioning Metabolomic Studies PR:PROJECT_TYPE MS Qualitative Analysis PR:PROJECT_SUMMARY Qualitative metabolomic studies in response to very-low CO2 and high CO2 in PR:PROJECT_SUMMARY Microchloropsis gaditana PR:INSTITUTE International Centre for Genetic Engineering and Biotechnology PR:DEPARTMENT Integrative Biology PR:LABORATORY Omics of Algae PR:LAST_NAME Jutur PR:FIRST_NAME Pannaga Pavan PR:ADDRESS 2nd Floor, New Building, ICGEB, Aruna Asaf Ali Marg, New Delhi - 110067 PR:EMAIL jppavan@icgeb.res.in PR:PHONE +91 11 26741358 PR:PROJECT_COMMENTS http://www.icgeb.res.in/ #STUDY ST:STUDY_TITLE Time-course experiment of Microchloropsis gaditana cells supplemented with CO2 ST:STUDY_TYPE Time-course experiment ST:STUDY_SUMMARY Experiments were conducted with Microchloropsis gaditana supplemented with ST:STUDY_SUMMARY very-low CO2 and high CO2. Sampling was done on the following time points: Day ST:STUDY_SUMMARY 3, 6 and 9. ST:INSTITUTE International Centre for Genetic Engineering and Biotechnology ST:DEPARTMENT Integrative Biology ST:LABORATORY Omics of Algae ST:LAST_NAME Jutur ST:FIRST_NAME Pannaga Pavan ST:ADDRESS 2nd Floor, New Building, ICGEB, Aruna Asaf Ali Marg, New Delhi - 110067 ST:EMAIL jppavan@icgeb.res.in ST:PHONE +91 11 26741358 ST:STUDY_COMMENTS Former name of species: Nannochloropsis gaditana Lubián #SUBJECT SU:SUBJECT_TYPE Plant SU:SUBJECT_SPECIES Nannochloropsis gaditana SU:TAXONOMY_ID 72520 SU:GENOTYPE_STRAIN NIES 2587 #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 CO2 Suppl (VLC) VLC_3 Condition:Control CO2 Concentration (ppm)=300; Sample Collection (in Days)=3; RAW_FILE_NAME=VLC-Day-3.mzData.xml SUBJECT_SAMPLE_FACTORS CO2 Suppl (VLC) VLC_6 Condition:Control CO2 Concentration (ppm)=300; Sample Collection (in Days)=6; RAW_FILE_NAME=VLC-Day-6.mzData.xml SUBJECT_SAMPLE_FACTORS CO2 Suppl (VLC) VLC_9 Condition:Control CO2 Concentration (ppm)=300; Sample Collection (in Days)=9; RAW_FILE_NAME=VLC-Day-9.mzData.xml SUBJECT_SAMPLE_FACTORS CO2 Suppl (HC) HC_3 Condition:Treatment CO2 Concentration (ppm)=30000; Sample Collection (in Days)=3; RAW_FILE_NAME=HC-Day-3.mzData.xml SUBJECT_SAMPLE_FACTORS CO2 Suppl (HC) HC_6 Condition:Treatment CO2 Concentration (ppm)=30000; Sample Collection (in Days)=6; RAW_FILE_NAME=HC-Day-6.mzData.xml SUBJECT_SAMPLE_FACTORS CO2 Suppl (HC) HC_9 Condition:Treatment CO2 Concentration (ppm)=30000; Sample Collection (in Days)=9; RAW_FILE_NAME=HC-Day-9.mzData.xml #COLLECTION CO:COLLECTION_SUMMARY Marine microalgae Microchloropsis gaditana NIES 2587 is procured from Microbial CO:COLLECTION_SUMMARY Culture Collection, National Institute for Environmental Studies (NIES), CO:COLLECTION_SUMMARY Tsukuba, Japan. The strain was grown in minimal medium F/2 (Guillard and Ryther, CO:COLLECTION_SUMMARY 1962) under a light regime of 16:8 h and an illumination of 150 µmol m−2 CO:COLLECTION_SUMMARY s−1 photosynthetically active radiation (PAR) in a multi-cultivator MC 1000-OD CO:COLLECTION_SUMMARY (Photon Systems Instruments, Czech Republic) with a flow rate of 800 mL min-1 CO:COLLECTION_SUMMARY with continuous bubbling of air at 24 °C. CO:SAMPLE_TYPE Algae #TREATMENT TR:TREATMENT_SUMMARY Microchloropsis gaditana cells were grown in a Multicultivator in the presence TR:TREATMENT_SUMMARY of very-low CO2 (300 ppm) and high CO2 (30,000 ppm) for 9 days with an TR:TREATMENT_SUMMARY illumination intensity of 150 uE. TR:TREATMENT_COMPOUND CO2 #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Quenched cells were resuspended in 1 mL of ice-cold SP:SAMPLEPREP_SUMMARY methanol/ethanol/chloroform(2:6:2), followed by sonication of resuspended cells SP:SAMPLEPREP_SUMMARY in sonication bath for 15 min. Later, these samples were centrifuged at SP:SAMPLEPREP_SUMMARY 10,000×g for 15 min at 4 °C to get rid of cell debris. The supernatant was SP:SAMPLEPREP_SUMMARY filtered using a 0.2-µm filter. One hundred microlitres of supernatant was SP:SAMPLEPREP_SUMMARY taken and dried under nitrogen stream. The dried leftover was dissolved in 10 SP:SAMPLEPREP_SUMMARY µL of freshly prepared methoxyamine hydrochloride solution (40 mg mL−1 in SP:SAMPLEPREP_SUMMARY pyridine) and incubated at 30 °C for 90 min with shaking. To the above SP:SAMPLEPREP_SUMMARY solution, 90 µL of N-methyl-N-(trimethylsilyl)trifluoroacetamide was added and SP:SAMPLEPREP_SUMMARY incubated at 37 °C for 30 min. The samples were centrifuged at 14,000×g for 3 SP:SAMPLEPREP_SUMMARY min, and the supernatant was taken for the GC-MS/MS analysis. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY GC-triple quadrupole analysis was performed on an HP-5 gas chromatograph with CH:CHROMATOGRAPHY_SUMMARY standard liners containing glass wool in split mode (1:5) at 250°C injector CH:CHROMATOGRAPHY_SUMMARY temperature. The GC was operated at constant flow of 1 ml/min helium on a 30-m, CH:CHROMATOGRAPHY_SUMMARY 0.25-mm i.d., 0.25-μm HP-5 column, a start temperature of 60°C, 3 min CH:CHROMATOGRAPHY_SUMMARY isothermal, temperature ramping by 5°C/min to 180°C, 3 min isothermal and CH:CHROMATOGRAPHY_SUMMARY finally temperature ramping of 10°C/min to 310°C. CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Agilent 7890A CH:COLUMN_NAME Agilent HP5-MS (30m x 0.25mm, 0.25 um) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Agilent 7890A MS:INSTRUMENT_TYPE Triple quadrupole MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS NA #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS Peak area MS_METABOLITE_DATA_START Samples VLC_3 VLC_6 VLC_9 HC_3 HC_6 HC_9 Factors Condition:Control Condition:Control Condition:Control Condition:Treatment Condition:Treatment Condition:Treatment L-Alanine 2753288 20836730 3351986 85724174 72809832 73916376 L-Valine 568745.84 754206.44 123640.4 610783.576 902574.92 1013851.28 L-Leucine 92730.3 62809.3232 1186947.84 25307981 25714165 1052921.646 L-Proline 17650266 45604945 1020033.3 1135018.872 1205246.619 1144366.086 Glycine 2472808 709695.896 650348.504 779923.6432 867955.608 4371179 D-Glucose 1202279.25 3514207 111205185 3924068 1018965.047 9781138 D-Fructose 30997537 28132817 56812249 215134.296 132319.9561 107839.1569 D-Galactose 57056472 60350550 63585768 27030895 5319005 3057564 D-Mannose 25333.91796 45625.78041 62809.3232 30662.8192 38081.2432 36350.2776 Sucrose 80065419 1211675.92 1038579.36 66553003 125317542 13672418 Maltose 1041546.73 1115236.408 3047268 59347.392 53165.372 47972.4752 Acetate 1204257.496 98170298 165544698 32311815 1232496.963 216675899 Succinate 64243.55184 168150.944 127695.8051 868746.9066 913455.2752 997827.4842 Benzoic acid 30786.4596 62067.4808 90480.04472 52918.0912 53264.28432 30415.5384 D-Lactic acid 10331676 29113949 980468.372 7140861 786352.944 994316.0968 Hydroxybutyric acid 531653.72 6341295 7276647 79129.856 98417.7584 123937.137 Glycerol 37745171 48245716 27783976 12689675 15619799 8932030 Thiourea 49827.0812 74480.97696 99431.60968 30910.1 38130.69936 54154.4952 GABA 17661471 19692658 17465268 709695.896 736896.784 902574.92 Sorbitol 2982958 4496696 8828040 1008411.102 1157274.144 82878118 Dibenzoxepine 8973832 8354680 7868691 14193863 6678232 6502793 Prenol phosphate 3907612 4000782 3896655 8967792 4381828 4371179 Palmitic acid 10859096 16000365 21745223 14252690 25733786 24358051 ALA 75525799 54381397 80618719 37252300 18005594 3700141 Stearic acid 1199311.88 65752031 4673910 23549913 79645672 76203622 Phytol 10832355 15859577 15413254 17839275 3739047 5781216 D-Trehalose 736896.784 956976.696 22834321 98417.7584 3743880 6462489 Ergostene 168150.944 234916.76 628093.232 1053416.208 29889756 19040482 Ethanolamine 13839769 15097673 9469512 816026.64 926066.596 903564.0432 L-Threonine 4178041 1150350.282 1013851.28 1006111.391 1068253.056 1152007.063 Myo-inositol 9468261 28337395 17974911 744933.41 2670151 5714209 Hexadecanedioic acid 21698314 310090.1232 227545.3194 655294.12 764963.1548 862762.7112 Alpha-tocopherol 48961.5984 61820.2 94461.2656 662712.544 1192783.667 6223374 Oxalic acid 803662.6 890210.88 1026957.162 3499921 717114.32 681505.8848 Docosanoic acid 160732.52 185460.6 210683.2416 15902906 764097.672 1043277.695 Dihydroquinazoline 1018796.896 1140459.05 12401720 236400.4448 801189.792 5138048 Glyceryl glucoside 863751.8344 970666.1611 1068994.898 1093228.417 1152081.247 9898937 Arachidic acid 150841.288 206798.4602 231949.3904 2984397 1038579.36 1078391.569 Propionic acid 63798.4464 186029.3458 213972.0762 1069880.164 2561840 36823272 Linoleic acid 206776.205 230836.6268 509398.448 12016035 973297.2288 1177647.609 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name HMDB ID PubChem ID KEGG ID L-Alanine HMDB0000161 5950 C00041 L-Valine HMDB0000883 6287 C00183 L-Leucine HMDB0000687 6106 C00123 L-Proline HMDB0000162 145742 C00148 Glycine HMDB0000123 750 C00037 D-Glucose HMDB0000122 5793 C00031 D-Fructose HMDB0000660 439709 C02336 D-Galactose HMDB0000143 439357 C00984 D-Mannose HMDB0000169 18950 C00159 Sucrose HMDB0000258 5988 C00089 Maltose HMDB0000163 10991489 C00208 Acetate HMDB0000042 176 C00033 Succinate HMDB0000254 1110 C00042 Benzoic acid HMDB0001870 243 C00180 D-Lactic acid HMDB0001311 61503 C00256 Hydroxybutyric acid HMDB0000710 10413 C00989 Glycerol HMDB0000131 753 C00116 Thiourea HMDB0034155 2723790 C14415 GABA HMDB0000112 119 C00334 Sorbitol HMDB0000247 5780 C00794 Dibenzoxepine Prenol phosphate Palmitic acid HMDB0000220 985 C00249 ALA HMDB0000161 5950 C00041 Stearic acid HMDB0000827 5281 C01530 Phytol HMDB0002019 5280435 C01389 D-Trehalose Ergostene Ethanolamine HMDB0000149 700 C00189 L-Threonine HMDB0000167 6288 C00188 Myo-inositol HMDB0000211 C00137 Hexadecanedioic acid HMDB0000672 10459 C19615 Alpha-tocopherol HMDB0001893 14985 C02477 Oxalic acid HMDB0002329 971 C00209 Docosanoic acid HMDB0000944 8215 C08281 Dihydroquinazoline Glyceryl glucoside Arachidic acid HMDB0002212 10467 C06425 Propionic acid HMDB0000237 1032 C00163 Linoleic acid HMDB0000673 5280450 C01595 METABOLITES_END #FACTORS #END