Precision Relative Standard Deviation Values Research Articles

Establishment of simultaneous analysis validation for Perilla frutescens var. acuta extract

In this study, the UV spectra maximum absorption wavelengths and molecular weights of major compounds 1–3 were identified in Perilla frutescens var. acuta extract using high-performance liquid chromatography (HPLC)-DAD and HPLC-MS/MS. Luteolin-7-O-diglucuronide (1), scutellarin (2), and rosmarinic acid (3) were identified, and simultaneous analysis validation was performed for quantification using HPLC-DAD. All linearities showed R2 values of 0.9993 or higher, and the limits of detection and quantification were measured in the ranges of 4.4802–5.4133 and 13.5764–16.4038 µg/mL, respectively. The relative standard deviation (RSD) values for accuracy ranged from 0.09 to 0.89%, and the recovery rates were in the range of 99.36–101.96%. The precision RSD values within the laboratory were in the range of 0.05–1.04%, and the RSD values for repeatability evaluation were below the guideline criteria of 2%. The content change of the three marker compounds within 24 h was 0.36–0.69%, confirming the chemical stability of the extract within 24 h, which was below the guideline criteria of 2.0% established by the Ministry of Food and Drug Safety guidelines. The content analysis of the extract according to ethanol concentration showed that luteolin-7-O-diglucuronide had the highest content of 302.22 µg/mL in the 0% ethanol extract, while scutellarin and rosmarinic acid showed the highest contents of 103.61 and 118.77 µg/mL, respectively, in the 60% ethanol extract.

A novel and robust analytical method for the quantification of perospirone in human plasma using an LC–MS/MS system with self-internal standard calibration

ObjectivesThis study aims to establish a novel method for measuring perospirone in human plasma for therapeutic drug monitoring (TDM) by liquid chromatography–mass spectrometry (LC–MS) coupled with an automatic liquid chromatograph mass spectrometer coupler 9500 (LC–MS/MS-Mate 9500), which has been equipped with self-internal standard (SIS) calibration technology. Design & methodsA novel and attractive analytical calibration method designed for perospirone, calibration with SIS, was reported. After protein precipitation with acetonitrile-cyclopentanol (9:1, v/v) containing 1% NH3·H2O, LC–MS quantification of perospirone was performed by multiple reaction monitoring in the positive mode with quantitative and qualitative analysis of the ion pairs m/z 427.30 → 177.15 and 427.30 → 166.15 for perospirone and SIS. Chromatographic separation was accomplished in < 2.0 min on an Hypersil GOLDTM C18 column (2.1 mm × 50 mm, 3.0 μm) using a mobile methanol phase and 0.1% formic acid in water. ResultsThis method showed good selectivity because no interfering peaks were observed in the plasma samples during the 2.0-min run time. The calibration curve range was 0.05–20 ng/mL, with a correlation coefficient of ≥ 0.9995. Intraday and interday accuracies were 98.3%–107.9%, respectively, with precision relative standard deviation values of < 10%. The matrix effects ranged from 92.7% to 96.1%, and extraction recoveries were between 97.3% and 108.8%. Finally, this method was successfully applied to routine clinical TDM for 142 patients. The perospirone plasma concentrations of the patients ranged between 0.07 and 10.96 ng/mL. ConclusionsThis bioanalytical method can be used for the quantification of perospirone in human plasma by LC–MS/MS-Mate 9500 using perospirone itself as the SIS.

A validated LC-MS/MS assay for the quantitative determination of curdione in rabbit plasma and its application to a pharmacokinetic study after administration of zedoary turmeric oil and bioavailability of the oil.

A selective and sensitive liquid chromatography tandem mass spectrometry method was developed for the first time for the identification and quantification of curdione in rabbit plasma after vaginal drug administration and intravenous administration of zedoary turmeric oil (ZTO) solution (10 mg/kg). The analysis was performed on a triple-quadrupole tandem mass spectrometer with multiple reaction monitoring mode via electrospray ionization source in positive ionization mode. After mixing with internal standard diazepam, plasma samples were extracted with ethyl ether-acetic ether (1:1, v/v). Chromatographic separation was carried out on a C18 column with gradient elution using a mixture of water and acetonitrile (both containing 0.1% formic acid) as mobile phases. Linearity ranged over 1.06-106 and 10.6-530 ng/mL (r ≥ 0.995) with the lower limit of quantfication 1.06 ng/mL. The intra- and inter-day precision relative standard deviation values were <12% and the accuracy relative error was from -10.6 to -6.1% at all quality control sample levels. The method was applied to a study of the pharmacokinetics of curdione after vaginal drug administration and intravenous administration of ZTO.

Quantitation of glutathione by liquid chromatography/positive electrospray ionization tandem mass spectrometry

Glutathione (GSH) is a tripeptide composed of glutamate, cysteine, and glycine. It is present in practically all cells and has several important roles, such as preventing the oxidation of the sulfhydryl groups of proteins within a cell. Evidence for GSH deficiency or depletion has been found in a variety of diseases and toxicity-related studies, including diabetes and induction of oxidative stress to form reactive oxygen species which cause DNA, lipid, and protein oxidations. A simple, selective, and sensitive analytical method for measuring low levels of GSH in biological fluids would therefore be desirable to conduct GSH deficiency or depletion-related mechanistic toxicity studies. Here a method for both low- and high-level quantitation of GSH from cultured cells and rat liver tissues via liquid chromatography/positive electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) has been developed. The lower limit of quantitation (LOQ) of the method was 5 ng/mL. The method is linear over a wide dynamic concentration range of 5.0 to 5000.0 ng/mL, with a correlation coefficient R2 > 0.99. The intra-day assay precision relative standard deviation (RSD) values for all quality control (QC) samples were < or =16.31%, with accuracy values ranging from 94.13 to 97.80%. The inter-day assay precision RSD values for all QC samples were < or =15.94%, with accuracy values ranging from 94.51 to 100.29%. With this method, low levels of GSH from diethyl maleate (DEM)-treated mouse lymphoma cells, and GSH in rat liver tissues, were quantified.

Affinity Liquid Chromatography Method for the Quantification of Immunoglobulin G in Bovine Colostrum Powders

An affinity liquid chromatography (LC) method for the determination of bovine immunoglobulin G (IgG), using protein G coupled to an agarose support, was modified to permit the quantification of IgG in colostrum-based powders. Sample preparation included pH adjustment to 4.6 to precipitate casein and denatured whey protein. The method was applied to a range of colostrum powders and was compared with the alternative independent methods of surface plasmon resonance immunoassay, radial immunodiffusion, and reversed-phase LC. The method was rapid, and performance parameters included a working range of 10-150 microg IgG and precision relative standard deviation values of <10%.

Direct Determination of Free Methionine in Soy-Based Infant Formula

A method was developed for the direct determination of free methionine in soy-based infant formula, with analyte separation and quantitation by reversed-phase liquid chromatography (LC), and UV absorbance at 214 nm, respectively. Sample preparation required only dilution with mobile phase and syringe filtration. Using a 0.02M KH2PO4 mobile phase (pH adjusted to 2.9 with 85% o-phosphoric acid) and 0.7 mL/min flow rate, methionine eluted at approximately 8 min, and total run time was 14 min after column regeneration with acetonitrile-water. System linearity was demonstrated as peak area versus analyte concentration, ranging from 80 to 120% of the formula specification for free methionine (r > 0.999, and all residuals < 0.45%). Intermediate precision relative standard deviation values were < 1.5% for ready-to-feed and reconstituted powder samples, and recoveries ranged from 98.0 to 103.5% for inter-method comparison with an amino acid analyzer method. The limit of quantitation was 3 mg methionine/L in the "as fed" infant formula. Despite the relatively weak UV absorptivity of methionine, the 214 nm signal was sufficiently intense in the 30-65 mg/L (201-436 microM) range to afford quantitation by peak area proportionation versus a 2-point external standard calibration. This direct UV detection after reversed-phase LC separation provides a simple and accurate method for determining free methionine without derivatization.

Simultaneous determination of four anthracyclines and three metabolites in human serum by liquid chromatography–electrospray mass spectrometry

A sensitive and very specific method, using liquid chromatography–electrospray mass spectrometry (LC–ES-MS), was developed for the determination of epirubicin, doxorubicin, daunorubicin, idarubicin and the respective active metabolites of the last three, namely doxorubicinol, daunorubicinol and idarubicinol in human serum, using aclarubicin as internal standard. Once thawed, 0.5-ml serum samples underwent an automated solid-phase extraction, using C 18 Bond Elut cartridges (Varian) and a Zymark Rapid-Trace robot. After elution of the compounds with chloroform–2-propanol (4:1, v/v) and evaporation, the residue was reconstituted with a mixture of 5 m M ammonium formate buffer (pH 4.5)–acetonitrile (60:40, v/v). The chromatographic separation was performed using a Symmetry C 18, 3.5 μm (150×1 mm I.D.) reversed-phase column, and a mixture of 5 m M ammonium formate buffer (pH 3)–acetonitrile (70:30, v/v) as mobile phase, delivered at 50 μl/min. The compounds were detected in the selected ion monitoring mode using, as quantitation ions, m/ z 291 for idarubicin and idarubicinol, m/ z 321 for daunorubicin and daunorubicinol, m/ z 361 for epirubicin and doxorubicin, m/ z 363 for doxorubicinol and m/ z 812 for aclarubicin (I.S.). Extraction recovery was between 71 and 105% depending on compounds and concentration. The limit of detection was 0.5 ng/ml for daunorubicin and idarubicinol, 1 ng/ml for doxorubicin, epirubicin and idarubicin, 2 ng/ml for daunorubicinol and 2.5 ng/ml for doxorubicinol. The limit of quantitation (LOQ) was 2.5 ng/ml for doxorubicin, epirubicin and daunorubicinol, and 5 ng/ml for daunorubicin, idarubicin, doxorubicinol and idarubicinol. Linearity was verified from these LOQs up to 2000 ng/ml for the parent drugs ( r≥0.992) and 200 ng/ml for the active metabolites ( r≥0.985). Above LOQ, the within-day and between-day precision relative standard deviation values were all less than 15%. This assay was applied successfully to the analysis of human serum samples collected in patients administered doxorubicin or daunorubicin intravenously. This method is rapid, reliable, allows an easy sample preparation owing to the automated extraction and a high selectivity owing to MS detection.