Return to study ST002429 main page
MB Sample ID: SA242869
Local Sample ID: | OilPalm_Salt_10_14DAT_R2_NEG |
Subject ID: | SU002518 |
Subject Type: | Plant |
Subject Species: | Elaeis guineensis Jacq. |
Taxonomy ID: | NCBI:txid51953 |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU002518 |
Subject Type: | Plant |
Subject Species: | Elaeis guineensis Jacq. |
Taxonomy ID: | NCBI:txid51953 |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
OilPalm_Salt_10_14DAT_R2_NEG | SA242869 | FL030405 | 14 days | Group |
Collection:
Collection ID: | CO002511 |
Collection Summary: | The oil palm plants used in this study were clones regenerated out of embryogenic calluses obtained from the leaves of an adult plant—genotype AM33, a Deli x Ghana from ASD Costa Rica, as previously reported by [6]. Before starting the experiments, plants were standardized accordingly to the developmental stage, size, and number of leaves. They were in the growth stage known as bifid saplings, and the experiment was performed in March 2018 in a greenhouse at Embrapa Agroenergy in Brasília, DF, Brazil (S-15.732°, W-47.900°). The main environmental variables (temperature, humidity, and radiation) fluctuated according to the weather conditions and underwent monitoring throughout the entire experimental period using the data collected at a nearby meteorological station (S-15.789°, W-47.925°). We collected the apical leaves from control and stressed plants (0.0 and 2.0 g of NaCl per 100 g of substrate) 12 days after imposition of the treatments (DAT). |
Sample Type: | Plant |
Treatment:
Treatment ID: | TR002530 |
Treatment Summary: | The experiment consisted of five salinity levels (0.0, 0.5, 1.0, 1.5, and 2.0 g of NaCl per 100 g of substrate (a mixture of vermiculite, soil, and the Bioplant commercial substrate (Bioplant Agrícola Ltd.a., Nova Ponte, MG, Brazil), in a 1:1:1 ratio, on a dry basis), with four replicates in a completely randomized design. The substrate mixture was fertilized using 2.5 g L−1 of the N-P2O5-K2O formula (20-20-20). |
Sample Preparation:
Sampleprep ID: | SP002524 |
Sampleprep Summary: | Leaves harvested from control and stressed plants were immediately immersed in liquid nitrogen and stored at −80 °C until metabolite extraction: four plants for treatments. Before solvent extraction, all samples underwent grounding in liquid nitrogen. The solvents used were methanol grade UHPLC, acetonitrile grade LC-MS, formic acid grade LC-MS, sodium hydroxide ACS grade LC-MS, all from Sigma-Aldrich, and water treated in a Milli-Q system from Millipore. We employed a protocol to extract the metabolites in three phases (polar, non-polar, and protein pellet). Aliquots of 50 mg of ground sample were transferred to 2 mL microtubes, and then 1 mL of a mixture of 1:3 (v:v) methanol/methyl tert-butyl ether (MTBE) at −20 °C was added. Homogenization on an orbital shaker at 4.0 °C and ultrasound treatment in an ice bath were each performed for 10 min. As the next step, we added 500 μL of a mixture of 1:3 (v:v) methanol/water to each microtube. After centrifugation (15,300× g at 4.0 °C for 5 min), an upper non-polar (green) and a lower polar (brown) phase and a protein pellet remained in each microtube. After transferring both fractions separately to 1.5 mL microtubes, they were submitted to a Speed vac system (Centrivap, Labconco) to be vacuum dried. Finally, the dry-fraction, resuspended in 500 μL of 1:3 (v:v) methanol and water mixture and transferred to vials, were now ready for analysis. |
Combined analysis:
Analysis ID | AN003953 | AN003954 |
---|---|---|
Analysis type | MS | MS |
Chromatography type | Reversed phase | Reversed phase |
Chromatography system | Shimadzu Nexera X2 | Shimadzu Nexera X2 |
Column | Waters Acquity BEH HSS T3 (100 x 2.1mm, 1.8um) | Waters Acquity BEH HSS T3 (100 x 2.1mm, 1.8um) |
MS Type | ESI | ESI |
MS instrument type | QTOF | QTOF |
MS instrument name | Bruker maXis Impact qTOF | Bruker maXis Impact qTOF |
Ion Mode | POSITIVE | NEGATIVE |
Units | Peak intensity | Peak intensity |
Chromatography:
Chromatography ID: | CH002926 |
Chromatography Summary: | Solvent A was 0.1% (v:v) formic acid in water and solvent B was 0.1% (v:v) formic acid in acetonitrile/methanol (70:30, v:v). The gradient elution used, with a flow rate of 0.4 mL min–1, was as follows: 0–1 min isocratic, 0% B; 1–3 min, 5% B; 3–10 min, 50% B; 10–13 min, 100% B; 13–15 min isocratic, 100% B; then, 5 min rebalancing was conducted to the initial conditions. |
Instrument Name: | Shimadzu Nexera X2 |
Column Name: | Waters Acquity BEH HSS T3 (100 x 2.1mm, 1.8um) |
Column Temperature: | - |
Flow Gradient: | 0–1 min isocratic, 0% B; 1–3 min, 5% B; 3–10 min, 50% B; 10–13 min, 100% B; 13–15 min isocratic, 100% B; then, 5 min rebalancing was conducted to the initial conditions. |
Flow Rate: | 0.4 mL/min |
Solvent A: | 100% water; 0.1% formic acid |
Solvent B: | 70% acetonitrile/30% methanol; 0.1% formic acid |
Chromatography Type: | Reversed phase |
MS:
MS ID: | MS003688 |
Analysis ID: | AN003953 |
Instrument Name: | Bruker maXis Impact qTOF |
Instrument Type: | QTOF |
MS Type: | ESI |
MS Comments: | The rate of acquisition spectra was 3.00 Hz, mass range m/z 70–1200 for the polar fraction analysis and m/z 300–1600 for the lipidic fraction. High-resolution mass spectrometry was used for detection (MaXis 4G Q-TOF MS, Bruker Daltonics) equipped with an electrospray source in positive (ESI-(+)-MS) and negative (ESI-(−)-MS) modes. The settings of the mass spectrometer were as follows: capillary voltage, 3800 V; dry gas flow, 9 L min−1; dry temperature, 200 °C; nebulizer pressure, 4 bar; final plate offset, 500 V. For the external calibration of the equipment, we used a sodium formate solution (10 mM HCOONa solution in 50:50 v:v isopropanol and water containing 0.2% formic acid) injected through a six-way valve at the beginning of each chromatographic run. Ampicillin ([M+H] + m/z 350.1186729 and [M-H]- m/z 348.1028826) was the internal standard for later peak normalization on data analysis. |
Ion Mode: | POSITIVE |
MS ID: | MS003689 |
Analysis ID: | AN003954 |
Instrument Name: | Bruker maXis Impact qTOF |
Instrument Type: | QTOF |
MS Type: | ESI |
MS Comments: | The rate of acquisition spectra was 3.00 Hz, mass range m/z 70–1200 for the polar fraction analysis and m/z 300–1600 for the lipidic fraction. High-resolution mass spectrometry was used for detection (MaXis 4G Q-TOF MS, Bruker Daltonics) equipped with an electrospray source in positive (ESI-(+)-MS) and negative (ESI-(−)-MS) modes. The settings of the mass spectrometer were as follows: capillary voltage, 3800 V; dry gas flow, 9 L min−1; dry temperature, 200 °C; nebulizer pressure, 4 bar; final plate offset, 500 V. For the external calibration of the equipment, we used a sodium formate solution (10 mM HCOONa solution in 50:50 v:v isopropanol and water containing 0.2% formic acid) injected through a six-way valve at the beginning of each chromatographic run. Ampicillin ([M+H] + m/z 350.1186729 and [M-H]- m/z 348.1028826) was the internal standard for later peak normalization on data analysis. |
Ion Mode: | NEGATIVE |