General Information
- Experiment ID
- OEX024423
- Experiment Name
- In vitro activity assays
- Experiment Type
- Metabolomic
- Description
- KEYWORDS: Salvia genus; Metabolomics; LC-MS Factors: Species Diversity: Difference
- Protocol
- Sample collection: Each plant was separated into root and leaf samples, with 3 biological replicates. The 2 parts of each plant were crushed into homogeneous powders respectively and sieved by 40-mesh sieve. Extraction: All samples were ground into a fine powder, and 10 mg prepared powder was extracted with 1 mL of 70% methanol (v/v) containing warfarin (5 μg/mL) as the internal reference for 1 h in an ultrasonic bath (53 kHz, 350 W) at 4 °C. After centrifuging at 12,000 x g at 4 °C for 30 min, the supernatant was used for UHPLC-QTOF-MS analysis. Chromatography: An Acquity UPLC system (Waters) coupled with a Xevo G2-XS QTOF mass spectrometer (Waters) was used for the metabolic analysis. The samples were first separated using an Acquity UPLC T3 column (2.1 mm x 100 mm, 1.8 μm). The column temperature was kept constant at 40 °C, and the flow rate was 0.40 mL/min with an injection volume of 1.0 µL. The mobile phases for gradient elution consisted of 0.1% (v/v) formic acid/water (solvent A) and 0.1% (v/v) formic acid/acetonitrile (solvent B). The elution gradients were 98-80% A over 0-7 min, 80-78% A over 7-11 min, 78-40% A over 11-20 min, 40-35% A over 20-25 min, 25-28 min at 35% A, 35-5% A over 28-30 min, 30-33 min at 5% A and final re-equilibration at 98% A for 5 min. Mass spectrometry: A Xevo G2-XS with an electrospray ionization source was used to collect MS data. MS was performed in both positive ion and negative ion modes under 30 V cone voltage, with a capillary voltage of 3.0 kV (positive ion mode) or 2.5 kV (negative ion mode). The desolvation temperature was set at 450 °C with the desolvation gas flow rate of 600 L/h, and the source temperature was set at 150 °C with a cone gas flow rate of 50 L/h. All data were collected in the MSE mode, with the following parameters: MSE range, 50-1200 m/z; MSE low energy, 6 eV; and MSE high energy, 15-30 eV. To calibrate the instrument, a sodium formate solution (0.5 mM) was used. Continuous acquisition of leucine enkephalin was used as an external standard for mass correction. All data were viewed in MassLynx v4.2 (Waters). Data transformation: The raw data were first converted to mzML format (Ms convert Gui) and the analysis base file (ABF Converter; https://www.reifycs.com/AbfConverter/) format before being imported into the MS-DIAL v4.60 software. The parameter settings were as follows: the tolerances for MS1 and MS2 were 0.01 Da and 0.02 Da, respectively. The mass range of MS1 and MS/MS was set between 100 and 1000, with the MS/MS amplitude cutoff at 800. Retention time range was set between 1.0 and 29.5 min, with a retention time tolerance of 0.15 min. The width of the mass slice was 0.1 Da. Adduct types such as [M-H]-, [M+HCOO]-, [M+Na-2H]-, [M+K-2H]-, [2M-H]- and [2M+FA-H]- were selected for the negative ion mode. [M+H]+, [M+Na]+, [M+K]+, [M+NH4]+ and [2M+H]+ were selected for the positive ion mode. Each of the obtained feature tables was split into two peak tables based on the retention time ranges, which were according to the accumulation level of phenolic acids (1.0-18.5 min for both root and leaf samples) and ATDs (16.5-29.5 min for root samples; 14.0-29.5 min for leaf samples). Metabolite identification: Reference standards were applied to identify the known abietane-type diterpenoids. Diagnostic neutral losses were used to identify the chemical modifications of rings and methyl groups at C-20 from abietane-type diterpenoids.
Experiment information
- Attributes
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ion_mode Negative retention_time 1.0-29.5 (50-1200) instrument_type ESI-QUAD-TOF experimental_method MS platform Xevo G2-XS QTof - Project
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1
Project ID Project name Create Date OEP004258 Functional divergence of CYP76AKs shapes the chemodiversity of abietane-type diterpenoids in genus Salvia 2023-07-14 - Samples
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36
Sample ID Sample Name Organism Tissue Create Date OES315988 NEG-11-Hydroxysugiol-ancNode1 Saccharomyces cerevisiae Mixture 2023-07-14 OES315989 NEG-11-Hydroxysugiol-ancNode2 Saccharomyces cerevisiae Mixture 2023-07-14 OES315990 NEG-11-Hydroxysugiol-ancNode3 Saccharomyces cerevisiae Mixture 2023-07-14 OES315991 NEG-11-Hydroxysugiol-ancNode4 Saccharomyces cerevisiae Mixture 2023-07-14 OES315992 NEG-11-Hydroxysugiol-ancNode5 Saccharomyces cerevisiae Mixture 2023-07-14 OES315993 NEG-11-Hydroxysugiol-ancNode6 Saccharomyces cerevisiae Mixture 2023-07-14 OES315994 NEG-11-Hydroxysugiol-pESC-His Saccharomyces cerevisiae Mixture 2023-07-14 OES315995 NEG-11-Hydroxysugiol-SaCYP76AK18 Saccharomyces cerevisiae Mixture 2023-07-14 OES315996 NEG-11-Hydroxysugiol-SaCYP76AK22 Saccharomyces cerevisiae Mixture 2023-07-14 OES315997 NEG-11-Hydroxysugiol-SaCYP76AK7 Saccharomyces cerevisiae Mixture 2023-07-14 OES315998 NEG-11-Hydroxysugiol-ScCYP76AK18 Saccharomyces cerevisiae Mixture 2023-07-14 OES315999 NEG-11-Hydroxysugiol-ScCYP76AK22 Saccharomyces cerevisiae Mixture 2023-07-14 OES316000 NEG-11-Hydroxysugiol-ScCYP76AK7 Saccharomyces cerevisiae Mixture 2023-07-14 OES316001 NEG-11-Hydroxysugiol-SmCYP76AK1 Saccharomyces cerevisiae Mixture 2023-07-14 OES316002 NEG-11-Hydroxysugiol-SmCYP76AK2 Saccharomyces cerevisiae Mixture 2023-07-14 OES316003 NEG-11-Hydroxysugiol-SmCYP76AK3 Saccharomyces cerevisiae Mixture 2023-07-14 OES316004 NEG-11-Hydroxysugiol-SmCYP76AK5 Saccharomyces cerevisiae Mixture 2023-07-14 OES316005 NEG-11-Hydroxysugiol-SoCYP76AK6 Saccharomyces cerevisiae Mixture 2023-07-14 OES316006 NEG-11-Hydroxysugiol-SpCYP76AK18 Saccharomyces cerevisiae Mixture 2023-07-14 OES316007 NEG-11-Hydroxysugiol-SpCYP76AK22 Saccharomyces cerevisiae Mixture 2023-07-14 OES316008 NEG-11-Hydroxysugiol-SpCYP76AK7 Saccharomyces cerevisiae Mixture 2023-07-14 OES316009 NEG-11-Hydroxysugiol-SrCYP76AK7 Saccharomyces cerevisiae Mixture 2023-07-14 OES316010 NEG-11-Hydroxysugiol-SrCYP76AK8 Saccharomyces cerevisiae Mixture 2023-07-14 OES316011 NEG-Ferruginol-pESC-His Saccharomyces cerevisiae Mixture 2023-07-14 OES316012 NEG-Ferruginol-SmCYP76AH3 Saccharomyces cerevisiae Mixture 2023-07-14 OES316013 NEG-Ferruginol-SmCYP76AH3-ancNode1 Saccharomyces cerevisiae Mixture 2023-07-14 OES316014 NEG-Ferruginol-SmCYP76AH3-ancNode2 Saccharomyces cerevisiae Mixture 2023-07-14 OES316015 NEG-Ferruginol-SmCYP76AH3-ancNode3 Saccharomyces cerevisiae Mixture 2023-07-14 OES316016 NEG-Ferruginol-SmCYP76AH3-ancNode4 Saccharomyces cerevisiae Mixture 2023-07-14 OES316017 NEG-Ferruginol-SmCYP76AH3-ancNode5 Saccharomyces cerevisiae Mixture 2023-07-14 OES316018 NEG-Ferruginol-SmCYP76AH3-ancNode6 Saccharomyces cerevisiae Mixture 2023-07-14 OES316019 NEG-Ferruginol-SmCYP76AH3-ScCYP76AK18 Saccharomyces cerevisiae Mixture 2023-07-14 OES316020 NEG-Ferruginol-SmCYP76AH3-SmCYP76AK2 Saccharomyces cerevisiae Mixture 2023-07-14 OES316021 NEG-Ferruginol-SmCYP76AH3-SmCYP76AK3 Saccharomyces cerevisiae Mixture 2023-07-14 OES316022 NEG-Ferruginol-SmCYP76AH3-SmCYP76AK5 Saccharomyces cerevisiae Mixture 2023-07-14 OES316023 NEG-Ferruginol-SmCYP76AH3-SpCYP76AK22 Saccharomyces cerevisiae Mixture 2023-07-14 - Runs
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36
Run ID Run Name Sample Data Num Create Date OER423078 NEG-11-Hydroxysugiol-ancNode1 NEG-11-Hydroxysugiol-ancNode1 1 2023-07-16 OER423079 NEG-11-Hydroxysugiol-ancNode2 NEG-11-Hydroxysugiol-ancNode2 1 2023-07-16 OER423080 NEG-11-Hydroxysugiol-ancNode3 NEG-11-Hydroxysugiol-ancNode3 1 2023-07-16 OER423081 NEG-11-Hydroxysugiol-ancNode4 NEG-11-Hydroxysugiol-ancNode4 1 2023-07-16 OER423082 NEG-11-Hydroxysugiol-ancNode5 NEG-11-Hydroxysugiol-ancNode5 1 2023-07-16 OER423083 NEG-11-Hydroxysugiol-ancNode6 NEG-11-Hydroxysugiol-ancNode6 1 2023-07-16 OER423084 NEG-11-Hydroxysugiol-pESC-His NEG-11-Hydroxysugiol-pESC-His 1 2023-07-16 OER423085 NEG-11-Hydroxysugiol-SaCYP76AK18 NEG-11-Hydroxysugiol-SaCYP76AK18 1 2023-07-16 OER423086 NEG-11-Hydroxysugiol-SaCYP76AK22 NEG-11-Hydroxysugiol-SaCYP76AK22 1 2023-07-16 OER423087 NEG-11-Hydroxysugiol-SaCYP76AK7 NEG-11-Hydroxysugiol-SaCYP76AK7 1 2023-07-16 OER423088 NEG-11-Hydroxysugiol-ScCYP76AK18 NEG-11-Hydroxysugiol-ScCYP76AK18 1 2023-07-16 OER423089 NEG-11-Hydroxysugiol-ScCYP76AK22 NEG-11-Hydroxysugiol-ScCYP76AK22 1 2023-07-16 OER423090 NEG-11-Hydroxysugiol-ScCYP76AK7 NEG-11-Hydroxysugiol-ScCYP76AK7 1 2023-07-16 OER423091 NEG-11-Hydroxysugiol-SmCYP76AK1 NEG-11-Hydroxysugiol-SmCYP76AK1 1 2023-07-16 OER423092 NEG-11-Hydroxysugiol-SmCYP76AK2 NEG-11-Hydroxysugiol-SmCYP76AK2 1 2023-07-16 OER423093 NEG-11-Hydroxysugiol-SmCYP76AK3 NEG-11-Hydroxysugiol-SmCYP76AK3 1 2023-07-16 OER423094 NEG-11-Hydroxysugiol-SmCYP76AK5 NEG-11-Hydroxysugiol-SmCYP76AK5 1 2023-07-16 OER423095 NEG-11-Hydroxysugiol-SoCYP76AK6 NEG-11-Hydroxysugiol-SoCYP76AK6 1 2023-07-16 OER423096 NEG-11-Hydroxysugiol-SpCYP76AK18 NEG-11-Hydroxysugiol-SpCYP76AK18 1 2023-07-16 OER423097 NEG-11-Hydroxysugiol-SpCYP76AK22 NEG-11-Hydroxysugiol-SpCYP76AK22 1 2023-07-16 OER423098 NEG-11-Hydroxysugiol-SpCYP76AK7 NEG-11-Hydroxysugiol-SpCYP76AK7 1 2023-07-16 OER423099 NEG-11-Hydroxysugiol-SrCYP76AK7 NEG-11-Hydroxysugiol-SrCYP76AK7 1 2023-07-16 OER423100 NEG-11-Hydroxysugiol-SrCYP76AK8 NEG-11-Hydroxysugiol-SrCYP76AK8 1 2023-07-16 OER423101 NEG-Ferruginol-pESC-His NEG-Ferruginol-pESC-His 1 2023-07-16 OER423102 NEG-Ferruginol-SmCYP76AH3 NEG-Ferruginol-SmCYP76AH3 1 2023-07-16 OER423103 NEG-Ferruginol-SmCYP76AH3-ancNode1 NEG-Ferruginol-SmCYP76AH3-ancNode1 1 2023-07-16 OER423104 NEG-Ferruginol-SmCYP76AH3-ancNode2 NEG-Ferruginol-SmCYP76AH3-ancNode2 1 2023-07-16 OER423105 NEG-Ferruginol-SmCYP76AH3-ancNode3 NEG-Ferruginol-SmCYP76AH3-ancNode3 1 2023-07-16 OER423106 NEG-Ferruginol-SmCYP76AH3-ancNode4 NEG-Ferruginol-SmCYP76AH3-ancNode4 1 2023-07-16 OER423107 NEG-Ferruginol-SmCYP76AH3-ancNode5 NEG-Ferruginol-SmCYP76AH3-ancNode5 1 2023-07-16 OER423108 NEG-Ferruginol-SmCYP76AH3-ancNode6 NEG-Ferruginol-SmCYP76AH3-ancNode6 1 2023-07-16 OER423109 NEG-Ferruginol-SmCYP76AH3-ScCYP76AK18 NEG-Ferruginol-SmCYP76AH3-ScCYP76AK18 1 2023-07-16 OER423110 NEG-Ferruginol-SmCYP76AH3-SmCYP76AK2 NEG-Ferruginol-SmCYP76AH3-SmCYP76AK2 1 2023-07-16 OER423111 NEG-Ferruginol-SmCYP76AH3-SmCYP76AK3 NEG-Ferruginol-SmCYP76AH3-SmCYP76AK3 1 2023-07-16 OER423112 NEG-Ferruginol-SmCYP76AH3-SmCYP76AK5 NEG-Ferruginol-SmCYP76AH3-SmCYP76AK5 1 2023-07-16 OER423113 NEG-Ferruginol-SmCYP76AH3-SpCYP76AK22 NEG-Ferruginol-SmCYP76AH3-SpCYP76AK22 1 2023-07-16
Author Information
- Create Date
- 2023-07-14
- Last Modified
- 2023-07-18
- Submission
- Wancheng Chen
- Shanghai University of Traditional Chinese Medicine