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An Optimized High Throughput Clean-Up Method Using Mixed-Mode SPE Plate for the Analysis of Free Arachidonic Acid in Plasma by LC-MS/MS.

Wang W, Qin S, Li L, Chen X, Wang Q, Wei J - Int J Anal Chem (2015)

Bottom Line: The calibration curve ranged from 10 to 2500 ng/mL with sufficient linearity (r (2) = 0.9999).The recoveries were in the range of 99.38% to 103.21% with RSD less than 6%.The limit of detection is 3 ng/mL.

View Article: PubMed Central - PubMed

Affiliation: School of Material Science and Engineering, Tianjin Polytechnic University, Tianjin 300387, China.

ABSTRACT
A high throughput sample preparation method was developed utilizing mixed-mode solid phase extraction (SPE) in 96-well plate format for the determination of free arachidonic acid in plasma by LC-MS/MS. Plasma was mixed with 3% aqueous ammonia and loaded into each well of 96-well plate. After washing with water and methanol sequentially, 3% of formic acid in acetonitrile was used to elute arachidonic acid. The collected fraction was injected onto a reversed phase column at 30°C with mobile phase of acetonitrile/water (70 : 30, v/v) and detected by LC-MS/MS coupled with electrospray ionization (ESI) in multiple reaction monitoring (MRM) mode. The calibration curve ranged from 10 to 2500 ng/mL with sufficient linearity (r (2) = 0.9999). The recoveries were in the range of 99.38% to 103.21% with RSD less than 6%. The limit of detection is 3 ng/mL.

No MeSH data available.


Related in: MedlinePlus

Product ion mass spectra of arachidonic acid.
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Related In: Results  -  Collection


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fig2: Product ion mass spectra of arachidonic acid.

Mentions: The analysis was accomplished with a Shimadzu LC-20A binary HPLC system with an autoinjector coupled with API4000+ triple quadrupole tandem mass spectrometer (AB SCIEX, MA, USA). AB SCIEX Analyst software (version 1.5.1) was used for data acquisition. The HPLC column was a 3 μm, 2.1 mm × 50 mm Venusil ASB C18 (Agela Technologies) operated at 30°C under an isocratic condition with mobile phase of acetonitrile/water (75 : 25, v/v). Flow rate was 0.2 mL/min and injection volume was 5 μL. The target compounds eluted from the HPLC column were introduced directly into the MS source. Electrospray ionization (ESI) with negative ion mode was selected for arachidonic acid and that with positive ion mode was selected for phospholipids, respectively. Quantitative analysis was performed under MRM mode by calculating the peak areas. The optimal MS parameters were listed in Table 1. The ions were detected by multiple reaction monitoring (MRM), monitoring the [M + H]+ transition of the m/z precursor ion to the m/z of the product ion for arachidonic acid. These MS/MS transitions utilized for analysis were m/z 303/259.1 and 303/205.1. An example of the mass spectra of arachidonic acid was shown in Figure 2. A UV detector at 254 nm wavelength was applied for the detection of proteins.


An Optimized High Throughput Clean-Up Method Using Mixed-Mode SPE Plate for the Analysis of Free Arachidonic Acid in Plasma by LC-MS/MS.

Wang W, Qin S, Li L, Chen X, Wang Q, Wei J - Int J Anal Chem (2015)

Product ion mass spectra of arachidonic acid.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4383463&req=5

fig2: Product ion mass spectra of arachidonic acid.
Mentions: The analysis was accomplished with a Shimadzu LC-20A binary HPLC system with an autoinjector coupled with API4000+ triple quadrupole tandem mass spectrometer (AB SCIEX, MA, USA). AB SCIEX Analyst software (version 1.5.1) was used for data acquisition. The HPLC column was a 3 μm, 2.1 mm × 50 mm Venusil ASB C18 (Agela Technologies) operated at 30°C under an isocratic condition with mobile phase of acetonitrile/water (75 : 25, v/v). Flow rate was 0.2 mL/min and injection volume was 5 μL. The target compounds eluted from the HPLC column were introduced directly into the MS source. Electrospray ionization (ESI) with negative ion mode was selected for arachidonic acid and that with positive ion mode was selected for phospholipids, respectively. Quantitative analysis was performed under MRM mode by calculating the peak areas. The optimal MS parameters were listed in Table 1. The ions were detected by multiple reaction monitoring (MRM), monitoring the [M + H]+ transition of the m/z precursor ion to the m/z of the product ion for arachidonic acid. These MS/MS transitions utilized for analysis were m/z 303/259.1 and 303/205.1. An example of the mass spectra of arachidonic acid was shown in Figure 2. A UV detector at 254 nm wavelength was applied for the detection of proteins.

Bottom Line: The calibration curve ranged from 10 to 2500 ng/mL with sufficient linearity (r (2) = 0.9999).The recoveries were in the range of 99.38% to 103.21% with RSD less than 6%.The limit of detection is 3 ng/mL.

View Article: PubMed Central - PubMed

Affiliation: School of Material Science and Engineering, Tianjin Polytechnic University, Tianjin 300387, China.

ABSTRACT
A high throughput sample preparation method was developed utilizing mixed-mode solid phase extraction (SPE) in 96-well plate format for the determination of free arachidonic acid in plasma by LC-MS/MS. Plasma was mixed with 3% aqueous ammonia and loaded into each well of 96-well plate. After washing with water and methanol sequentially, 3% of formic acid in acetonitrile was used to elute arachidonic acid. The collected fraction was injected onto a reversed phase column at 30°C with mobile phase of acetonitrile/water (70 : 30, v/v) and detected by LC-MS/MS coupled with electrospray ionization (ESI) in multiple reaction monitoring (MRM) mode. The calibration curve ranged from 10 to 2500 ng/mL with sufficient linearity (r (2) = 0.9999). The recoveries were in the range of 99.38% to 103.21% with RSD less than 6%. The limit of detection is 3 ng/mL.

No MeSH data available.


Related in: MedlinePlus