Metabolomic analysis was performed on random samples from each group, including six samples from the control group, six from the HTN group, six from the OSA group, and seven samples from the complication group. The supernatant of the salivary samples was thawed at 4°C and mixed in equal portions of 100 μL with 400 μL cold methanol/acetonitrile (1:1, V/V) to remove protein. The mixture was centrifuged for 15 min (14,000 g, 4°C) and the resulting supernatant was dried in a vacuum centrifuge. For liquid chromatography-mass spectrometry (LC-MS) analysis, the samples were re-dissolved in 100 μL acetonitrile/water (1:1, V/V) solvent. Analyses were performed using an ultra-high-performance liquid chromatography (UHPLC) system (1290 Infinity LC, Agilent Technologies) coupled to a quadrupole time-of-flight mass spectrometer (AB Sciex TripleTOF 6600). The samples were separated on the Agilent 1290 Infinity LC by hydrophilic interaction liquid chromatography (HILIC) column with a column temperature of 25°C, flow rate of 0.5 mL/min, injection volume of 2 μL, and mobile phase composition of 25 mM ammonium acetate, 25 mM ammonia, water, and acetonitrile. QC samples were inserted into the sample queue to monitor and evaluate the stability of the system and the reliability of the experimental data.
1290 infinity lc
Manufactured by Agilent Technologies
Sourced in United States, Ireland
The 1290 Infinity LC is a high-performance liquid chromatography (HPLC) system designed for analytical and preparative applications. It features a modular design, allowing for the integration of various components to meet specific analytical needs. The 1290 Infinity LC provides reliable, accurate, and reproducible results for a wide range of applications.
Automatically generated - may contain errors
Lab products found in correlation
Tripletof 6600, by AB Sciex (14 mentions)
Acquiy uplc beh 1.7 μm column, by Waters Corporation (5 mentions)
Acquiy uplc beh 1.7 m column, by Waters Corporation (5 mentions)
Tripletof 5600, by AB Sciex (5 mentions)
Acquity uplc beh 1.7 μm column, by Waters Corporation (4 mentions)
Qtrap 5500, by AB Sciex (4 mentions)
Accurate mass q tof ms, by Agilent Technologies (3 mentions)
Uplc beh amide column, by Waters Corporation (3 mentions)
Multiquant software, by AB Sciex (3 mentions)
Tripletof 6600 mass spectrometer, by AB Sciex (3 mentions)
6460 qqq ms, by Agilent Technologies (2 mentions)
Acquity uplc beh amide column, by Waters Corporation (2 mentions)
Acquiy uplc beh column, by Waters Corporation (2 mentions)
6550 ifunnel lcq tof ms, by Agilent Technologies (2 mentions)
6545 qtof ms, by Agilent Technologies (2 mentions)
Qtrap 6500, by AB Sciex (2 mentions)
Waters uplc beh c18, by Waters Corporation (2 mentions)
6500 qtrap ms, by AB Sciex (2 mentions)
Xcalibur, by Thermo Fisher Scientific (1 mentions)
Q exactive orbitrap, by Thermo Fisher Scientific (1 mentions)
149 protocols using 1290 infinity lc
1
Salivary Metabolomic Profiling of Cardiometabolic Conditions
2
LC-MS Analysis of Reaction Products
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LC–MS analyses of the reaction product were carried out by using a system consisting of 6460 QQQ MS (Agilent, USA) and 1290 Infinity LC (Agilent, USA). Samples were filtered (0.45 μm, Millipore) and 10 μl was directly injected. The separation was performed using a ZORBOX-C18 reverse phase column (4.6 × 250 mm; 5 μm; Agilent, USA) protected by a pre-column with acetonitrile-water pre-mixed (60:40, vol/vol) solvent as the mobile phase at a flow rate of 0.6 ml/min. Mass spectrometry was performed in ESI source negative mode (50–400). The ion source temperature was set at 300°C. The capillary voltage and fragmentor were set at 3.5 kV and 135 V, respectively. Nebulizer gas was set at 45 psi. Data acquisition is carried out by Mass hunter.
3
Ultra-High Performance Liquid Chromatography-Mass Spectrometry
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Analyses were performed using a UHPLC (1290 Infinity LC, Agilent Technologies, Santa Clara, CA, USA) coupled to a quadrupole time-of-flight (AB Sciex TripleTOF 6600).
4
UHPLC-QTRAP Metabolite Analysis
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Utilizing a UHPLC (1290 Infinity LC; Agilent Technologies) coupled to a QTRAP (AB Sciex 5500), analyses of this study were performed. The mobile phase contained A = 25 mM CH3COONH4 + 0.08% FA in water and B = 0.1% FA in ACN. Samples were in the automatic sampler at 4°C, and the column temperatures were kept at 40°C constantly. The gradients were at a flow rate of 250 µL/min. A 4 µL aliquot of each sample was injected. The gradient was 90% B linearly reduced to 70% in 0–12 minutes, and then was reduced to 50% in 12–18 minutes and reduced to 40% in 18–25 minutes, and kept for 25–30 minutes. Then the B was raised to 90% in 30–30.1 minutes and kept for 30.1–37 minutes.
The quality control (QC) samples were applied to examine and evaluate the repeatability and stability and of this system. At the same time, the standard mixture of AA metabolites was set for the correction of chromatographic retention time.
In ESI cationic mode, settings of conditions were as below: Ion Source Gas1 (Gas1) as 40, Ion Source Gas2 (Gas2) as 40, curtain gas (CUR) as 30, source temperature: 500°C, IonSpray Voltage Floating (ISVF) ± 5500 V. Adopt the MRM mode detection ion pair.
The quality control (QC) samples were applied to examine and evaluate the repeatability and stability and of this system. At the same time, the standard mixture of AA metabolites was set for the correction of chromatographic retention time.
In ESI cationic mode, settings of conditions were as below: Ion Source Gas1 (Gas1) as 40, Ion Source Gas2 (Gas2) as 40, curtain gas (CUR) as 30, source temperature: 500°C, IonSpray Voltage Floating (ISVF) ± 5500 V. Adopt the MRM mode detection ion pair.
5
Targeted Metabolite Analysis of Phloem Sap
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Targeted metabolites from the phloem sap were determined by a high-throughput and multiplexed LC/MS/MRM method (Wei et al., 2010 (link)) with detail in the Supplementary Material . Metabolomics profiling was analyzed using a UPLC-ESI-Q-TRAP-MS system (UHPLC, 1290 Infinity LC, Agilent Technologies, Santa Clara, CA, United States) coupled with QTRAP 5500 (AB SCIEX, Framingham, MA, United States).
6
Targeted Metabolomics via UHPLC-QTRAP
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Targeted metabolomics was performed using a UHPLC (1290 Infinity LC, Agilent Technologies) coupled to a QTRAP (AB Sciex 5500). The mobile phase consisted of 0.08% FA in 25 mM ammonium formate and 0.1% FA in ACN. For each sample, 2 μl aliquots were automatically injected at 4°C with a column temperature of 40°C, and the flow rate was 250 μl/min.
The gradient was as follows: 0-12 min, with liquid B changing linearly from 90% to 70%; 12-18 min, from 70% to 50%; 18-25 min, from 50% to 40% and kept for 5 min, and then liquid B was increased to 90% in 30-30.1 min and kept for 30.1-37 min. In ESI positive modes, the conditions were set as follows: source temperature 500℃; ion source gas 1 (Gas1): 40 psi; ion source gas 2 (Gas2): 40 psi; curtain gas (CUR): 30 psi; and ion spray voltage floating (ISVF): 5500 V. MRM mode was adopted to detect the amino acids and derivatives.
The gradient was as follows: 0-12 min, with liquid B changing linearly from 90% to 70%; 12-18 min, from 70% to 50%; 18-25 min, from 50% to 40% and kept for 5 min, and then liquid B was increased to 90% in 30-30.1 min and kept for 30.1-37 min. In ESI positive modes, the conditions were set as follows: source temperature 500℃; ion source gas 1 (Gas1): 40 psi; ion source gas 2 (Gas2): 40 psi; curtain gas (CUR): 30 psi; and ion spray voltage floating (ISVF): 5500 V. MRM mode was adopted to detect the amino acids and derivatives.
7
Targeted Metabolomics Analysis by GC-MS and LC-MS
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Targeted metabolomics analysis was performed by Aptbiotech Co., Ltd. (Shanghai, China) using GC–MS method. The samples were thawed at 4 ℃, and 100 μL aliquots were mixed with 400 μL cold methanol/acetonitrile (1:1, v:v) to precipitate protein. The mixture was centrifuged for 20 min (14,000 × g, 4 ℃). And then the supernatant was dried in a vacuum centrifuge. For liquid chromatography-mass spectrometry (LC–MS) analysis, the samples were redissolved in 100 μL acetonitrile/water (1:1, v:v). After vortex for 1 min, the tubes were centrifuged for 15 min (14,000 × g, 4 ℃), and the supernatants were collected for LC–MS analysis. The analysis was carried out using an ultra-high-performance liquid chromatography system (1290 Infinity LC, Agilent Technologies, Palo Alto, CA, USA) coupled with to a QTRAP (5500, AB Sciex, Framingham, MA, USA).
8
Untargeted Metabolomic Profiling of Liver
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The aforementioned liver sample was added into the pre-cold solvent of acetonitrile and methanol (acetonitrile/methanol/water = 2:2:1), followed by vortexed for 30 s, homogenized with a homogenizer, sonicated for 30 min with ice-bath, and incubated at −20 °C for 10 min. The mixture was freeze centrifugated at 14,000× g for 20 min. The supernatant was dried in a vacuum centrifuge, and re-dissolved in 100 μL acetonitrile solvent (acetonitrile /water = 1:1) for LC-MS/MS analysis. To examine the stability and repeatability, QC samples were prepared by pooling an equal aliquot of each sample and analyzed together with experimental samples. LC-MS/MS analysis was performed using a UHPLC (1290 Infinity LC, Agilent Technologies) coupled to a quadrupole time-of-flight (AB Sciex TripleTOF 6600) in Shanghai Applied Protein Technology Co., Ltd., according to the manufacturer’s instruction.
9
Metabolite Extraction and Analysis Protocol
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After grinding samples in liquid nitrogen, 1 cm3 of acetonitrile: methanol: water solvent (2:2:1, v/v) was added in 0.06 g of the sample and vortexed for 60 s. After sonication twice at low temperature for 0.5 h, the extracts were left at -20 °C for 1 h to precipitate the protein [44 (link)]. Afterward, they were filtered through a filter tube at 14,000 rcf and centrifuged at 4 °C for 20 min; the supernatant was freeze-dried and stored at -80 °C. LC/MS analyses were performed using a UHPLC (1290 Infinity LC, Agilent Technologies) coupled to a quadrupole time-of-flight mass spectrometer (AB Sciex Triple TOF 6600) in Shanghai Applied Protein Technology Co., Ltd. The raw MS data (wiff. scan files) were converted to MzXML files using Proteo Wizard MS Convert before importing them into freely available XCMS software [45 (link), 46 (link)]. Detailed procedures are available in Additional file 1 .
10
Metabolic Profiling of Lysine-Treated Cells
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Logarithmic phase cells were washed with PBS and re-suspended in M9 media with or without L-lysine. After 6 h incubation, the cells were pelleted (5 min at 140,00g, 4°C), washed with cold PBS, and snap frozen in liquid nitrogen. Sextuplicate samples were collected and sent for analysis by Metabolon-associated energy metabolism (Applied Protein Technology, Shanghai, China). A homogenate of 100 mg of sample mixed with 1 ml of cold methanol/acetonitrile/H2O (2:2:1, v/v/v) was sonicated at a low temperature (30 min/once, twice) and then centrifuged for 20 min (140,00g, 4°C). The supernatant was dried in a vacuum centrifuge. For LC-MS analysis, the dried samples were dissolved in 100 μl acetonitrile/water (1:1, v/v), adequately vortexed and then centrifuged (140,00 rpm, 4°C, 15 min). The supernatants were collected for the LC-MS/MS analysis. Analyses were performed using an UHPLC (1290 Infinity LC, Agilent Technologies) coupled to a QTRAP (AB Sciex 5500).
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