Pharmacokinetics of gastrodin and its metabolite p-hydroxybenzyl alcohol in rat blood, brain and bile by microdialysis coupled to LC-MS/MS.

Gastrodin is a pharmacologically active substance isolated from Gastrodia elata Blume with sedation, anti-convulsion and anti-epilepsy activities. A rapid and sensitive liquid chromatography technique coupled to tandem mass spectrometry (LC-MS/MS) system was developed to determine gastrodin and its metabolite p-hydroxybenzyl alcohol (HBA) in rat blood, brain and bile collected using microdialysis technique. The analytes were separated using a reversed phase column (4.6 mm x 150 mm, 5 microm). The mobile phase for column separation was 30% methanol with a flow rate of 0.6 mL/min. As a post-column addition, 1% ammonium hydroxide solution (in methanol) was additionally pumped via a T-connection using a chromatographic pump (BAS PM-80, USA) at a flow rate of 0.2 mL/min after the column separation. A LC-MS/MS system equipped with a negative electrospray ionization (ESI) source in multiple reaction monitoring (MRM) mode was used to monitor m/z 285.0-->122.9 and m/z 123.0-->105.0 transitions for gastrodin and HBA, respectively. The lower limit of quantification (LLoQ) for gastrodin and HBA were 0.5 and 2 ng/mL, respectively. The calibration curves were linear over the range of 0.5-5,000 ng/mL and 2-1,000 ng/mL for gastrodin and HBA with a coefficient of determination >0.995, respectively. This selective and sensitive method is useful for the determination of gastrodin and HBA and in the pharmacokinetic studies of these compounds.

Oral bioavailability of curcumin in rat and the herbal analysis from Curcuma longa by LC-MS/MS.

This study presents a validated liquid chromatography technique coupled with tandem mass spectrometry (LC-MS/MS) to measure curcumin in rat plasma and provide curcuminoids analysis from the extract of Curcumin longa L. This method was applied to investigate the pharmacokinetics of curcumin in a freely moving rat. The analytes were separated by a reversed phase C18 column (150x4.6 mm I.D., particle size 5 microm) and eluted with acetonitrile-1mM HCOOH mobile phase (70:30, v/v) with a flow rate of 0.8 ml/min in rat plasma and herbal extracts. Multiple reaction monitoring (MRM) was used to monitor the transition of the deprotonated molecule m/z of 367 [M-H]- to the product ion 217 for curcumin, a m/z of 337-217 for demethoxycurcumin and a m/z of 265-224 for honokiol (internal standard) analysis. The limit of detection (LOD) and quantification (LOQ) of curcumin in the rat plasma were 1 and 5 ng/ml, respectively. The method was linear in the range of 5-1000 ng/ml with a coefficient of correlation greater than 0.996 in the rat plasma. After curcumin (500 mg/kg, p.o.) administration, the maximum concentration (Cmax) and the time to reach maximum concentration (Tmax) were 0.06+/-0.01 microg/ml and 41.7+/-5.4 min, respectively. The elimination half-life (t1/2,beta) were 28.1+/-5.6 and 44.5+/-7.5 min for curcumin (500 mg/kg, p.o.) and curcumin (10 mg/kg, i.v.), respectively. The oral bioavailability was about 1%.

Analysis of brain distribution and biliary excretion of a nutrient supplement, gastrodin, in rat.

Gastrodin is a bioactive constituent of rhizome in Gastrodia elata Blume (Orchidaceae) The aim of this study is to develop a rapid and sensitive liquid chromatographic method coupled to microdialysis sampling system to measure the unbound of gastrodin in rat blood, brain and bile. Microdialysis probes were simultaneously inserted into the jugular vein, brain striatum and bile duct of each anesthetized rat for sampling after the administration of gastrodin (100 or 300 mg kg(-1)) through the femoral vein. Separation of unbound gastrodin from various biological fluids was applied to an RP-select B column (250 mm x 4.6 mm i.d., 5 microm). The mobile phase consisted of acetonitrile-50 mM potassium dihydrogen phosphate buffer-triethylamine (5:95:0.1, v/v/v, adjusted to pH 2.5 with orthophosphoric acid) with a flow rate of 1 mL min(-1). The UV detector wavelength was set at 221 nm. Fifteen minutes after the administration, the gastrodin reached the peak concentration in brain and bile. In addition, the results indicate that gastrodin penetrates the blood-brain barrier (BBB) and goes through hepatobiliary excretion.

Pharmacokinetics of (-)-epigallocatechin-3-gallate in conscious and freely moving rats and its brain regional distribution.

A liquid chromatography technique coupled with tandem mass spectrometry (LC-MS/MS) electrospray ionization was used to measure (-)-epigallocatechin-3-gallate (EGCG) in rat plasma. This method was applied to investigate the pharmacokinetics of EGCG in a conscious and freely moving rat by an automated blood sampling device. Multiple reaction monitoring (MRM) was used to monitor the transition of the deprotonated molecule m/z of 457 [M - H]- to the product ion 169 for EGCG and the m/z of 187 to 164 for the internal standard. The limit of quantification (LOQ) of EGCG in rat plasma was determined to be 5 ng/mL, and the linear range was 5-5000 ng/mL. The protein binding of EGCG in rat plasma was 92.4 +/- 2.5%. The brain distribution result indicated that EGCG may potentially penetrate through the blood-brain barrier at a lower rate. The disposition of EGCG in the rat blood was fitted well by the two-compartmental model after intravenous administration (10 mg/kg, iv). The elimination half-life of EGCG was 62 +/- 11 and 48 +/- 13 min for intravenous (10 mg/kg) and oral (100 mg/kg) administration, respectively. The pharmacokinetic data indicate that the oral bioavailability of EGCG in a conscious and freely moving rat was about 4.95%.

Measurement and pharmacokinetic study of plumbagin in a conscious freely moving rat using liquid chromatography/tandem mass spectrometry.

The aim of the present study is to develop an automated blood sampling (ABS) method coupled to a liquid chromatography-tandem mass spectroscopy (LC-MS/MS) method to evaluate the oral bioavailability of plumbagin in a conscious freely moving rat. Plumbagin, an herbal ingredient, was isolated from Plumbago zeylanica L. The separation was performed using a reversed phase C18 (150mmx4.6mm I.D.; 5microm) column and was eluted with the mobile phase of water-acetonitrile (40:60, v/v) at a flow-rate of 0.8ml/min. Multiple reaction monitoring (MRM) was used to monitor the transition of the deprotonated molecule m/z 187 [MH](-) to the product ion m/z 159 [MHCO](-) for the plumbagin analysis. The calibration curve was linear over the concentration range of 10-2000ng/ml with a coefficient estimation of 0.995. The intra- and inter-day variations (% relative standard deviation; RSD and % bias) of the assay for rat plasma samples were less than 17%. The limit of detection and the limit of quantification were 5 and 10ng/ml, respectively. Recovery of plumbagin from the rat plasma was about 80%. This LC-MS/MS method has been validated to study the pharmacokinetics of plumbagin in rats. The oral bioavailability (AUC(PO)/Dose(PO))/(AUC(IV)/Dose(IV)) of plumbagin was about 38.7+/-5%.

Determination and identification of plumbagin from the roots of Plumbago zeylanica L. by liquid chromatography with tandem mass spectrometry.

Plumbagin (5-hydroxy-2-methyl-1,4-naphthoquinone) is an herbal ingredient which is isolated from the root of Plumbago zeylanica L. This herb is a semi-climbing subshrub distributed in thickets or grassland at low elevations of Taiwan. The crushed roots of P. zeylanica L. were ground from lumps to powder and boiled with H2O, 50% EtOH, or 95% EtOH. Chromatographic separation of plumbagin from the herb was carried out using a ZORBAX Extend-C18 column (150 x 4.6 mm I.D.; 5 microm) that was eluted with the mobile phase of water-methanol (10:90, v/v). Multiple reaction monitoring (MRM) was used to monitor the transition of the deprotonated molecule m/z 187 [M-H]- to the product ion m/z 159 [M-H-CO]- for plumbagin analysis. The limit of quantification was determined to and accuracy of 1 ng/ml. Furthermore, the mass fractions of plumbagin in P. zeylanica L. for H2O, 50% EtOH and 95% EtOH were 0.24 +/- 0.04, 3.92 +/- 0.87 and 13.4 +/- 1.59 g/kg, respectively. These results present a reliable liquid chromatography coupled with tandem mass spectrometric (LC-MS/MS) method for the determination of plumbagin form herbal medicines.