At 20 in an Agilent 6400 Triple Quadrupole LC/MS process (Agilent Systems > 자유게시판

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At 20 within an Agilent 6400 Triple Marimastat Quadrupole LC/MS program (Agilent Systems, Santa Clara, CA, United states of america). The cell stage in the binary gradient elution mixture consisted of (A) aqueous formic acid (0.1 , v/v) and (B) acetonitrile, along with the sample separation was accomplished working with the subsequent gradient: 98 A and 2 B from 0 to 2 min, 100 B from 3 to seven min, and ninety eight A and 2 B from 8 to ten min. The stream fee was 0.2 mL/min. All samples ended up maintained at four throughout analysis in the multiple reaction monitoring manner to maximize sensitivity. The resource temperature was set at 100 , though the desolvation fuel temperature was three hundred . The gasoline flow rate was 3.0 L/min. Details had been analyzed employing the Agilent MassHunter Qualitative Analysis B.04.00 software (Agilent Systems, Santa Clara, CA, United states). Specifications for glutamine, glutamate, alanine, and theanine were dissolvedLi et al. BMC Plant Biology (2016) 16:Page fifteen ofin methanol:water (three:one, v/v) for just a ultimate concentration of 100 g/mL. The diluted specifications were being blended to make calibration curves. The glutamine, glutamate, alanine, and theanine concentrations had been identified primarily based around the calibration curves. The Student's t-test was accustomed to review the metabolite concentrations inside the YG and W phases with those people from the G phase.Quantification of leaf metabolites by GC-TOF-MSAdditional filesAdditional file 1: Gene Ontology enrichment classifications of the differentially expressed genes up-regulated among the yellow-green and re-greening levels. (XLSX 14 kb) Supplemental file 2: Gene Ontology enrichment classifications on the differentially expressed genes down-regulated concerning the yellow-green and re-greening levels. (XLSX fifteen kb) More file three: Unigenes included in carotenoid biosynthesis. (XLSX eleven kb) Additional file four: Unigenes involved in chlorophyll biosynthesis. (XLSX ten kb) Extra file 5: Unigenes involved in theanine biosynthesis. (XLSX eleven kb) Additional file 6: Primers employed for quantitative reverse transcription polymerase chain reaction analyses. (XLSX eleven kb)Each individual 100-mg leaf sample was submerged in five hundred L methanol:water (3:one, v/v) and homogenized PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/18111632 in the ball mill at sixty five Hz for 3 min. The mixture was then centrifuged at twelve,000 rpm for 10 min at four . A 350-L aliquot in the supernatant was transferred into a glass sampling vial and vacuum-dried at room temperature for 1.5 h. The dried residue was handled with 80 L methoxyamine (20 mg/ mL in pyridine) and incubated at 80 for twenty min, followed by the addition of 80 L BSTFA (made up of one TCMS, v/v) and an incubation at 70 for one h. The GC-TOF-MS assessment was carried out utilizing an Agilent 7890 fuel chromatography technique coupled by using a Pegasus HT time-of-flight mass spectrometer. The system involved a DB-5MS capillary column coated with five diphenyl cross-linked with 95 dimethylpolysiloxane (30 m ?250 m interior diameter, 0.twenty five m movie thickness; J W Scientific, Folsom, CA, United states of america). A 1-L aliquot of analyte was injected in the splitless mode. Helium was used since the provider fuel. The front inlet purge circulation was 3 mL/min, even though the fuel flow charge by means of the column was twenty mL/min. The temperature was originally established at fifty ?C for 1 min, and after that elevated to 320 at a amount of ten / min. The PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/9547713 temperature was held at 320 for 5 min. The injection and transfer line temperatures were being 280 , when the ion source temperature was 220 . The strength stage was established at -70 eV while in the electron effect method. The mass spectrometry info in just the m/z variety of eighty five?00 were acquir.