#METABOLOMICS WORKBENCH Cristina_20171010_080737 DATATRACK_ID:1251 STUDY_ID:ST000889 ANALYSIS_ID:AN001451 PROJECT_ID:PR000618 VERSION 1 CREATED_ON November 7, 2017, 12:46 pm #PROJECT PR:PROJECT_TITLE GC-MS Quercus ilex analysis PR:PROJECT_TYPE MS qualitative analysis PR:PROJECT_SUMMARY GC-MS analysis of samples from Quercus ilex. The objetive of the study is to PR:PROJECT_SUMMARY obtain a better understanding of the molecular mechanisms mediating phenotypes PR:PROJECT_SUMMARY of interest and the selection of elite genotypes to be used in restoration and PR:PROJECT_SUMMARY reforestation programs, especially in a future climate change scenario. PR:INSTITUTE Universidad de Córdoba PR:DEPARTMENT Department Biochemistry and Molecular Biology PR:LABORATORY Agroforestry and Plant Biochemistry, Proteomics and Systems Biology PR:LAST_NAME López-Hidalgo PR:FIRST_NAME Cristina PR:ADDRESS Campus de Rabanales; Edificio C6, Planta Baja PR:EMAIL n12lohic@uco.es PR:PHONE 626894948 PR:FUNDING_SOURCE This work was supported by the University of Cordoba and financial support from PR:FUNDING_SOURCE the Spanish Ministry of Economy and Competitiveness (Project BIO2015-64737-R2). PR:PUBLICATIONS Metabolic network reconstruction from multi-omics data integration in Quercus PR:PUBLICATIONS ilex #STUDY ST:STUDY_TITLE GC-MS analysis of Quercus ilex organs (leaves, roots and acorns) ST:STUDY_TYPE Untargeted metabolomics ST:STUDY_SUMMARY Qualitative metabolomics study on leaves, roots and acorns from Quercus ilex ST:STUDY_SUMMARY plantlets. We analyzed polar(metanol:water) and apolar (chloroform) fractions. ST:INSTITUTE Universidad de Córdoba ST:DEPARTMENT Department Biochemistry and Molecular Biology ST:LABORATORY Agroforestry and Plant Biochemistry, Proteomics and Systems Biology ST:LAST_NAME López-Hidalgo ST:FIRST_NAME Cristina ST:ADDRESS Campus de Rabanales; Edificio C6, Planta Baja ST:EMAIL n12lohic@uco.es ST:PHONE 626894948 #SUBJECT SU:SUBJECT_TYPE Plant SU:SUBJECT_SPECIES Quercus ilex SU:TAXONOMY_ID 58334 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS - Acorn1 Sample Type:Acorn SUBJECT_SAMPLE_FACTORS - Acorn2 Sample Type:Acorn SUBJECT_SAMPLE_FACTORS - Acorn3 Sample Type:Acorn SUBJECT_SAMPLE_FACTORS - Leaf1 Sample Type:Leaf SUBJECT_SAMPLE_FACTORS - Leaf2 Sample Type:Leaf SUBJECT_SAMPLE_FACTORS - Leaf3 Sample Type:Leaf SUBJECT_SAMPLE_FACTORS - Root1 Sample Type:Root SUBJECT_SAMPLE_FACTORS - Root2 Sample Type:Root SUBJECT_SAMPLE_FACTORS - Root3 Sample Type:Root #COLLECTION CO:COLLECTION_SUMMARY Germinated embryo, leaves and roots from four-months plantlets were collected CO:COLLECTION_SUMMARY separately, weighted and individually frozen in liquid nitrogen. #TREATMENT TR:TREATMENT_SUMMARY Mature acorns from Holm oak (Quercus ilex L. subsp. ballota [Desf.] Samp.) were TR:TREATMENT_SUMMARY collected from a tree located in Aldea de Cuenca (province of Córdoba, TR:TREATMENT_SUMMARY Andalusia, Spain). Acorns were germinated and seedlings grew in a chamber under TR:TREATMENT_SUMMARY controlled conditions (a 12h photoperiod, a temperature of 21± 1ºC, a relative TR:TREATMENT_SUMMARY humidity of 60 ± 5% and an irradiance of 200 µmol m-2 s-1, Echevarría-Zomeño TR:TREATMENT_SUMMARY et al., 2009). #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Metabolites were extracted from each type of organs under study (leaves, roots SP:SAMPLEPREP_SUMMARY and acorns), as described by (Valledor et al., 2014). 600 µL of cold (4oC) SP:SAMPLEPREP_SUMMARY metabolite extraction buffer (methanol: chloroform: water; 5:2:2) were added to SP:SAMPLEPREP_SUMMARY 15 mg of dried tissue (frozen, lyophilized weight). Powdered samples were SP:SAMPLEPREP_SUMMARY extracted by vortexing for 10 s and sonicating in an ultrasonic bath at 4oC and SP:SAMPLEPREP_SUMMARY maximum frequency (40 kHz). Samples were centrifuged at 20.000 g for 4 min at SP:SAMPLEPREP_SUMMARY 4oC and the supernatant were transferred to 2 mL microcentrifuge tubes that SP:SAMPLEPREP_SUMMARY contained 400 µL of phase separation mix (chloroform: water; 1:1). Tubes with SP:SAMPLEPREP_SUMMARY metabolites were centrifuged at 20.000 g for 4 min at 4oC. The two phases were SP:SAMPLEPREP_SUMMARY clearly defined with a sharp interface. Polar and non-polar metabolites, upper SP:SAMPLEPREP_SUMMARY and lower layers respectively were transferred to new tubes. These two fractions SP:SAMPLEPREP_SUMMARY were washed again (200 µL of cold (4oC) chloroform for polar layer and with 200 SP:SAMPLEPREP_SUMMARY µL of cold (4oC) water for non-polar layer), centrifuged, and fractioned again. SP:SAMPLEPREP_SUMMARY Polar and non-polar layers were saved to new tubes and dried completely with a SP:SAMPLEPREP_SUMMARY micro concentrator Speedvac (Eppendorf Vacuum Concentrator Plus/5301). #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Agilent CH:COLUMN_NAME Agilent HP5-MS (30m × 0.25mm, 0.25 um) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:MS_COMMENTS - MS:INSTRUMENT_NAME Agilent 5975B MS:INSTRUMENT_TYPE Single quadrupole MS:MS_TYPE EI MS:ION_MODE POSITIVE #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS area MS_METABOLITE_DATA_START Samples Acorn1 Acorn2 Acorn3 Leaf1 Leaf2 Leaf3 Root1 Root2 Root3 Factors Sample Type:Acorn Sample Type:Acorn Sample Type:Acorn Sample Type:Leaf Sample Type:Leaf Sample Type:Leaf Sample Type:Root Sample Type:Root Sample Type:Root Pyruvic acid 0.000225086 0.000227532 0.000176324 0.000245691 0.000395084 0.000132614 0.000231924 0.000198613 0.000263381 L-Glutamic acid 0.009383847 0.011204216 0.009133578 0.023887896 0.018008481 0.018756116 0.018264871 0.017043787 0.016613909 Glucose 0.095106561 0.098597102 0.111436703 0.0559699 0.126006618 0.140069351 0.132825489 0.138491434 0.134375896 Alanine 0.013948962 0.015858275 0.011990025 0.005087434 0.004946619 0.003933144 0.01077464 0.011748393 0.011002695 Succinic acid 0.001664365 0.001965047 0.001608074 0.004448648 0.003313011 0.002389399 0.003730435 0.003615832 0.003653065 Aspartic acid 0.099544867 0.084462552 0.08322488 0.011434859 0.014895665 0.000810978 0.001740672 0.001653684 0.001654706 L-Serine 0.004248093 0.003585349 0.00311388 0.002126383 0.000133084 0.002046952 0.00344721 0.003690937 0.003315248 L(+)-Ascorbic acid 0 0 0 0.007123559 0.009692817 0.00799697 0 0 0 L-Phenylalanine 0.002158919 0.002888964 0.002517905 0 0 0 0 0 0 Urea 0.0000743 0.000257448 0.000217054 0.0000931 0.000246652 0.000199631 0.000215544 0.000227863 0.000188087 Sucrose 0.13219812 0.143758712 0.14705413 0.02124418 0.014829479 0.015712269 0.103147589 0.092766163 0.112469841 Glucose-6-phosphate 0.0000859 0.000136519 0.000112847 0.001058203 0.001338813 0.001209718 0.000473495 0.000579872 0.000391872 Glycerol 0.002941963 0.004723008 0.00287408 0.002798503 0.004080653 0.002696516 0.005862164 0.006095605 0.006506148 Fructose 0.003233623 0.005619345 0.004196509 0 0 0 0.000952496 0.000984398 0.001021309 Fumaric acid 0.001705578 0.001965047 0.001696236 0.003357119 0.001072761 0.001177688 0.001423644 0.001208096 0.001738493 Leucine 0.001347343 0.002940676 0.001639812 0.000349431 0.000482541 0.000280446 0 0 0 Galactose 0.01797514 0.024166632 0.01918404 0.003142822 0.003688113 0.002601191 0.018890402 0.018722381 0.019636166 myo-Inositol 0.086229949 0.083773062 0.087456653 0.06156464 0.08454103 0.059851294 0.055120637 0.048566711 0.047274822 L-Proline 0.003423836 0.003181997 0.003032771 0.001368415 0.001552144 0.001113874 0.002950815 0.002585327 0.003240482 Malic acid 0.12554066 0.114455377 0.132948219 0.130792759 0.130013403 0.123016584 0.161807707 0.155771384 0.148492043 Asparagine 0.007481716 0.008722051 0.00804037 0.003387498 0.003856097 0.002387616 0.010064088 0.010751628 0.012604908 Citric acid 0.001293449 0.001310031 0.001766765 0.136351733 0.14139228 0.137447351 0.024552724 0.025660983 0.024853168 Glycolic acid 0.000570639 0.000551592 0.000493707 0.00030039 0.000456924 0.000370155 0.001711592 0.002014235 0.001458117 Valine 0.006340437 0.005515922 0.005289717 0.000620534 0.00068762 0.000341936 0.004867777 0.008948519 0.007418426 D-Cellobiose 0 0 0 0.002273183 0.003561059 0.003324302 0 0 0 Lactic acid 0.002884899 0.002171894 0.001763239 0.002803623 0.001528239 0.001945222 0.019760575 0.018397203 0.021211108 L-Threonine 0.000339213 0.000482643 0.000316678 0.000306793 0.000297426 0.000313075 0.001058755 0.001755345 0.001315249 Gluconic acid 0.000621363 0.000720517 0.000684137 0.000512753 0.000356869 0.000436243 0 0 0 Maltose 0.034238362 0.014755091 0.02154678 0.000525146 0.000930556 0.000781745 0.000481951 0.000407512 0.0006454 Oxalic acid 0.003075112 0.002999282 0.002539064 0.002152468 0.001473387 0.003247829 0.000676149 0.000885547 0.000541134 Sorbose 0.000557958 0.000706727 0.000312446 0.00071183 0.000805933 0.000842482 0.127051422 0.132645834 0.128751206 Palmitic acid 0.003148027 0.003433661 0.002986927 0.001995825 0.001696991 0.002126236 0.003457072 0.003620513 0.003284022 Glyceric acid 0.000488214 0.000430931 0.000159044 0.000398772 0.001795961 0.001696492 0.000172007 0.00018057 0.000163315 Arabinose 0.000478703 0.000479196 0.000391439 0 0 0 0.000464007 0.000582327 0.000480401 D(-)-Quinic acid 0.079889511 0.083428317 0.07828781 0.136351733 0.141677345 0.137896436 0.050370389 0.044963746 0.051444982 Xylulose 0.0000891 0.000168236 0.000104031 0.000118074 0.000150959 0.0000997 0.00013405 0.000131288 0.000106993 D-(+)-Galacturonic acid 0.000104617 0.000124108 0.0000882 0.017681699 0.007522806 0.013500926 0.013766914 0.014299305 0.009000753 4-Aminobutyric acid (GABA) 0.056429893 0.047574825 0.052897169 0.001258845 0.001367102 0.001001542 0.006362238 0.006039013 0.007361601 Mannitol 0.002092344 0.002237396 0.002196996 0.000585806 0.000385402 0.000228779 0.00123238 0.001167238 0.001413789 L-Isoleucine 0.000424809 0.002999282 0.000955676 0.000242592 0.000391921 0.000583329 0.002628083 0.002379211 0.002450071 cis-Aconitic acid 0.001344173 0.001158344 0.001167264 0.001033876 0.000703688 0.000896316 0.000632687 0.000519912 0.000728306 L-Rhamnose 0.018070247 0.012824518 0.014705413 0.055001339 0.050752983 0.050937535 0.006824884 0.006989843 0.007554768 Oleic acid 0.004311497 0.005584871 0.005360246 0.005438283 0.003888588 0.004446679 0.006712496 0.007197697 0.006440189 Sorbitol 0.000596001 0.000461958 0.000507813 0.000208061 0.000138482 0.000168785 0.000451149 0.000496942 0.000484398 Salicylic acid 0 0 0 0 0 0 0.000933461 0.000824412 0.000810302 Glucaric acid 0.001230045 0.001410008 0.001414118 0 0 0 0 0 0 Galactaric acid 0.001547067 0.001630644 0.00156223 0.001850762 0.001085473 0.001388389 0.000321611 0.000345698 0.000323607 Galactonic acid 0.012585768 0.014410346 0.013012704 0.001100997 0.003502165 0.000714475 0 0 0 Maleic acid 0.004660221 0.004860906 0.004408097 0.019272965 0.009475763 0.007413349 0.013540445 0.013667987 0.012087224 Gallic acid 0.016897266 0.015996173 0.015022796 0.008819427 0.004668857 0.00670561 0.001528188 0.00182855 0.001390889 Stearic acid 0.000925704 0.00159617 0.001276585 0.00106985 0.001067972 0.001588343 0.002641768 0.002232248 0.002805032 Linoleic Acid 0.042520267 0.051710555 0.070251199 0.109548892 0.074350272 0.072809208 0.041213576 0.046834279 0.019566238 Ribonic acid 0.001366364 0.001351401 0.001184897 0.000801084 0.000697928 0.000709877 0.000480244 0.000466092 0.000419288 D-Erythrose 0.000213039 0.000230979 0.000209825 0.000188936 0.000188489 0.00018698 0.0000973 0.000103295 0.000112091 Maltotriose 0.005008946 0.008515204 0.005430776 0.000600452 0.001288181 0.001074663 0.000413592 0.000499124 0.000640359 Pyroglutamic acid 0.008654697 0.006136463 0.006947162 0 0 0 0.004048795 0.004468834 0.003819925 Melibiose 0.005991713 0.00334058 0.005430776 0.001191369 0.001886983 0.001912743 0.000244282 0.000178961 0.000330314 Catechin 0.006594055 0.004929855 0.006171336 0.058100218 0.021164892 0.038245339 0.059259023 0.064415634 0.055342684 Viburnitol 0.041529865 0.073085964 0.05430776 0.107000578 0.083983245 0.089356059 0.020253323 0.028947149 0.034895431 Epigallocatechin 0 0 0 0.010977509 0.018590712 0.017312681 0.003746992 0.003881178 0.003426823 Tridecane 0.017792474 0.010658668 0.029095857 0.000359298 0.007808246 0.004010303 0.009605598 0.008651487 0.010863739 Anthraquinone 0 0 0 0.001586214 0.003341128 0.002725863 0 0 0 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name retention index quantified m/z PubChem ID KEGG ID Pyruvic acid 11.59 147,217,133 C00022 L-Glutamic acid 25.23 246,147,128 C00025 Glucose 31.35 319,205,147 C00031 Alanine 12.13 116,190,75 C00041 Succinic acid 18.4 147,247,75 C00042 Aspartic acid 23.32 232,218,147 C00049 L-Serine 19.74 204,218,147 C00065 L(+)-Ascorbic acid 32.13 332,147,205 C00072 L-Phenylalanine 25.25 218,192,266 C00079 Urea 16.82 147,189,171 C00086 Sucrose 43.56 361,217,437 C00089 Glucose-6-phosphate 39.08 387,299,357 C00092 Glycerol 17.6 205,218,133 C00116 Fructose 29.33 217,437,147 C00095 Fumaric acid 23.19 156,147,232 C00122 Leucine 17.39 158,147,232 C00123 Galactose 31.2 319,205,147 C00124 myo-Inositol 31.65 305,318,217 C00137 L-Proline 17.87 142,147,75 C00148 Malic acid 22.7 147,233,245 C00149 Asparagine 26.24 231,116,132 C00152 Citric acid 29.5 273,147,347 C00158 Glycolic acid 11.21 147,205,177 C00160 Valine 15.78 144,218,75 C00183 D-Cellobiose 44.79 204,361,217 C00185 Lactic acid 10.69 147,117,191 C00186 L-Threonine 20.38 218,291,117 C00188 Gluconic acid 33.3073 292,319,147 C00198 Maltose 44.89 361,204,217 C00208 Oxalic acid 13.35 147,133,220 C00209 Sorbose 31.08 217,307,103 C00247 Palmitic acid 33.5 313,117,129 C00249 Glyceric acid 19.03 292,147,189 C00258 Arabinose 26.34 307,217,103 C00259 D(-)-Quinic acid 30.37 345,255,147 C00296 Xylulose 26.62 205,147,263 C00310 D-(+)-Galacturonic acid 39.8 217,204,292 C00333 4-Aminobutyric acid (GABA) 23.37 174,304,147 C00334 Mannitol 31.91 319,205,217 C00392 L-Isoleucine 17.94 158,147,218 C00407 cis-Aconitic acid 27.69 147,229,375 C00417 L-Rhamnose 33.96 204,319,220 C00507 Oleic acid 36.6 339,129,117 C00712 Sorbitol 32.05 319,147,217 C00794 Salicylic acid 27.97 267,370,193 C00805 Glucaric acid 33.63 333,292,305 C00818 Galactaric acid 39.8 217,204,292 C00879 Galactonic acid 30.89 205,275,147 C00880 Maleic acid 18.19 147,245,75 C01384 Gallic acid 32.17 458,281,443 C01424 Stearic acid 37.0349 341,117,129 C01530 Linoleic Acid 18.36 294,263,109 C01595 Ribonic acid 28.39 292,217,147 C01685 D-Erythrose 21.49 147,201,117 C01796 Maltotriose 44.19 361,204,217 C01835 Pyroglutamic acid 23.19 156,147,232 C01879 Melibiose 44.27 204,217,147 C05402 Catechin 46.1 368,355,204 C06562 Viburnitol 32.5 217,191,204 C08259 Epigallocatechin 46.65 456,355,345 C12136 Tridecane 9.82 112,98,85 C13834 Anthraquinone 46.26 369,267,204 C16207 METABOLITES_END #END