HPLC Analysis and Pharmacokinetic Study of Paeoniflorin after Intravenous Administration of a New Frozen Dry Powder Formulation in Rats

A simple and specific high performance liquid chromatographic (HPLC) method with UV detection using picroside II as the internal standard was developed and validated to determine the concentration of paeoniflorin in rat plasma and study its pharmacokinetics after an single intravenous administration of 40 mg kg^?1 paeoniflorin to Wistar rats. The analytes of interest were extracted from rat plasma samples by ethyl acetate after acidification with 0.05 mol L^?1 NaH₂PO₄ solution (pH 5.0). Chromatographic separation was achieved on an Agilent XDB C₁₈ column (250 × 4.6 mm I.D., 5 μm) with a Shim-pack GVP-ODS C₁₈ guard column (10 × 4.6 mm I.D., 5 μm) using a mobile phase consisting of acetonitrile–water–acetic acid (18:82:0.4, v/v/v) at a flow rate of 1.0 mL min^?1. The UV detection was performed at a wavelength of 230 nm. The linear calibration curves were obtained in the concentration range of 0.05–200.0 μg mL^?1 in rat plasma with the lower limit of quantification (LLOQ) of 0.05 μg mL^?1. The intra- and inter-day precisions in terms of % relative standard deviation (RSD) were lower than 5.7 and 8.2% in rat plasma, respectively. The accuracy in terms of % relative error (RE) ranged from ?1.9 to 2.6% in rat plasma. The extraction recoveries of paeoniflorin and picroside II were calculated to be 69.7 and 56.9%, respectively. This validated method was successfully applied to the pharmacokinetic study of a new paeoniflorin frozen dry power formulation. After single intravenous administration, the main pharmacokinetic parameters t _1/2, AUC_0-∞, CL_TOT, V _Z, MRT_0-∞ and V _ss were 0.739 ± 0.232 h, 43.75 ± 6.90 μg h mL^?1, 15.50 ± 2.46 L kg^?1 h^?1, 1.003 ± 0.401 L kg^?1, 0.480 ± 0.055 h and 0.444 ± 0.060 L kg^?1, respectively.     

Pharmacokinetic Study of ZS-1, a Targeted Peptide to NCI-H1299, in Rats Following Intravenous Administration

To investigate the pharmacokinetics of ZS-1 following intravenous injection in rats, ZS-1 was administered at doses of 20, 30 and 45 mg kg^?1, respectively. Blood samples were collected at 0.5, 3, 8, 12, 15, 20, 30, 40 and 45 min. ZS-1 in rat plasma was measured by LC. The limit of detection (LOD) was 0.02 μg mL^?1. The relative standard deviation (RSD) of intra- and inter-day precisions were <10%, and the accuracy of intra- and inter-day were >94%. The mean extraction recovery of ZS-1 was 86.1%. After intravenous injection at doses of 20, 30 and 45 mg kg^?1, the concentration–time curves of ZS-1 fitted well to one compartment model. Area under the concentration–time curves (AUC) increased with dose. Clearance rates (CL) and elimination half-lives (T _1/2) had no significant difference between different dose groups (P > 0.05). ZS-1 was stable in plasma after at 25 °C for 2, 4, 6 h, after three freeze–thaw cycles, after ?20 °C for a month, and after ?80 °C for 3 months. The accuracy of ZS-1 was between 96.8 and 106.9%. The results indicated there was no significant degradation. These data indicated that the method for analysis of ZS-1 was reliable and the pharmacokinetic data could guide dosing regimens to be tested in future clinical pharmacokinetic study.     

LC with Fluorescence Detection for the Tissue Pharmacokinetics of 2-Methoxyestradiol Solution and Liposome After Intravenous Administrations to Rats

2-Methoxyestradiol is currently in phase II clinical trials as a chemotherapeutic agent. An LC method with fluorescence detection was developed for determination of methoxyestradiol in rat lung. Sample was extracted with ethyl acetate and separated on a C₁₈ column (150 mm × 4.6 mm, 5 μm) with a mobile phase consisting of potassium dihydrogen phosphate-acetonitrile-triethylamine (55:45:0.3, v/v/v, pH 3.0). The excitation and emission wavelength were set at 285 and 325 nm, respectively. Standard curves were linear over the concentration range of 0.25–64 μg g^?1. The method was found to be precise, accurate and specific and can be applied to tissue pharmacokinetics of methoxyestradiol in rats.     

High-Performance Liquid Chromatographic Method for Determination and Pharmacokinetic Study of Chlorogenic Acid in the Plasma of Rats After Administration of the Chinese Medicinal Preparation Luying Decoction

A simple and sensitive high-performance liquid chromatographic method has been developed for determination of chlorogenic acid in rat plasma. Chlorogenic acid was extracted from plasma samples with methanol. HPLC analysis of the extracts was performed on a C₁₈ column (250 mm × 4.6 mm i.d., 5 µm particles). The mobile phase was acetonitrile −1% formic acid (9:91, v/v). The calibration plot was linear over the range 0.0420–2.10 µg mL⁻¹ and the lower limit of quantification was 0.0420 µg mL⁻¹. The method was reproducible and reliable with intra-day precision better than 8.2%, inter-day precision better than 9.1%, accuracy within ±8.3%, and mean extraction recovery above 84.4%. The validated method was successfully applied to pharmacokinetic studies of chlorogenic acid in rat plasma after administration of Luying decoction.     

LC Tissue Distribution Study of Paeonol in Rats after Oral Administration

Paeonol, an important constituent of the traditional Chinese medicine Cortex Moutan, has a variety of bioactivity. A simple and sensitive HPLC?CUV method has been developed for analysis of paeonol in different rat tissue (heart, liver, spleen, lung, kidney, and brain). Bio-samples were prepared by simple protein precipitation, and separation of paeonol was achieved on a C₁₈ column with methanol?C2% glacial acetic acid solution 70:30 (v/v) as mobile phase at a flow rate of 1.0 mL min^?1. UV detection was at 274 nm and the column temperature was 30 °C. Linearity was good between 0.025 and 5.0 ??g mL^?1 (r ² ?? 0.9990) for tissue samples. Inter-day and intra-day accuracy (as relative error, RE) and precision (as relative standard deviation, RSD) were <5.94 and 6.05%, respectively. The limit of detection was 0.025 ??g mL^?1 and extraction recovery for all samples was ??85.86%. The method was successfully applied to a tissue-distribution study after oral administration of 40 mg kg^?1 paeonol to healthy Sprague?CDawley rats. The study showed that paeonol was quickly distributed and eliminated after oral administration; liver and kidney were the major distribution tissues of paeonol in rats, and paeonol quickly passed through the blood?Cbrain barrier. It was also found there was no long-term accumulation of paeonol in rat tissues.     

RP-HPLC Determination and Pharmacokinetic Comparison of Cinnamic Acid in Rat Plasma After Administration of Di-Gu-Pi Decoction and Pure Cinnamic Acid

IntroductionTraditionally,Di-Gu-Pi has been used as a naturaltherapeutic agent for the treatment of diabetes,hemor-rhagic inframmation,hypertension,ulcers,and feverunder the guidance of the theory of Traditional ChineseMedicinal(TCM)science[1].This fact h…     

Simultaneous LC–MS Analysis and of Wogonin and Oroxylin A in Rat Plasma,and Pharmacokinetic Studies After Administration of the Active Fraction from Xiao-Xu-Ming Decoction

A rapid and specific high-performance liquid chromatographic method coupled with electrospray ionization mass spectrometric detection has been developed and validated for identification and quantification of wogonin and oroxylin A in rat plasma. Wogonin, oroxylin A, and diazepam (internal standard) were extracted from plasma samples by liquid–liquid extraction with ethyl acetate. Chromatographic separation was achieved on a C₁₈ column with acetonitrile–0.6% aqueous formic acid 35:65 (v/v) as mobile phase at a flow rate of 0.2 mL min⁻¹. Detection was performed with a single-quadrupole mass spectrometer in selected-ion-monitoring (SIM) mode. Linearity was good within the concentration range 14.4–360 ng mL⁻¹ for wogonin and 10.8–271 ng mL⁻¹ for oroxylin A; the correlation coefficients (r ²) were 0.9999. The intra-day and inter-day precision, as RSD, was below 12.4%, and accuracy ranged from 81.1 to 111.9%. The lower limit of quantification was 14.4 ng mL⁻¹ for wogonin and 10.8 ng mL⁻¹ for oroxylin A. This method was successfully used in the first pharmacokinetic study of wogonin and oroxylin A in rat plasma after oral administration of the active fraction from Xiao-xu-ming decoction.

     

LC Determination and Pharmacokinetic Study of Sinomenine in Dog Plasma

A simple HPLC method has been developed for determination of sinomenine in dog plasma and has been used to evaluate the pharmacokinetics of sinomenine tablets in dogs. Chromatographic separation was performed on a reversed-phase column with 0.78% (w/v) NaH₂PO₄-acetonitrile, 88:12 (v/v), as mobile phase, delivered at a flow rate of 1.5 mL min^?1. Detection was performed at 265 nm. The limit of quantification was 5.0 ng mL^?1. The calibration range was from 5.0 to 1000 ng mL^?1. The developed method was applied to pharmacokinetic studies of sinomenine sustained-release tablets (test preparation) and sinomenine conventional tablets (reference preparation) in six dogs. Pharmacokinetic data t _max, C _max, AUC _0-t , AUC _0-∞, and t _1/2 for both preparations were determined from plasma concentration-time profiles. The method was sufficiently sensitive, simple, and repeatable for use in pharmacokinetic studies.     

LC Determination and Pharmacokinetic Study of Gemfibrozil in Human Plasma

A simple and sensitive LC method for the quantitative determination of gemfibrozil in human plasma samples is described. Mometasone furoate was used as the internal standard. Plasma samples were pretreated by protein precipitation using methanol. Separation was performed at 40 °C on a YMC^® ODS-A reverse phase column (5 μm particle size, 150 mm × 4.6 mm i.d.) using 0.2% (v/v) triethylamine in water (adjusting to pH 4.0 with phosphoric acid) and acetonitrile (45:55, v/v) as mobile phase which was delivered at 1.5 mL min^?1. Ultraviolet detection was performed at 230 nm. The linear concentration range for gemfibrozil was 0.25–50 μg mL^?1. The detection limit of this method was 0.1 μg mL^?1. Intra- and inter-assay RSD ranged from 0.63 to 2.04% and 1.37 to 4.27%, respectively. The method was sensitive, simple and repeatable enough to be used in pharmacokinetic studies.     

LC Analysis of Isepamicin in Plasma Samples Post-Inhalation with Fluorescence Detection and Its Application to a Pharmacokinetic Study

A simple and novel LC method has been developed for determination of isepamicin (ISP) in rat plasma, an aminoglycoside antibiotic agent. After protein precipitation and clean-up procedure to remove lipophilic contaminants, ISP is derivatized by pre-column with 9-fluorenylmethyl chloroformate for fluorescence detection. Chromatographic separations are achieved using a C18 column and mobile phase consisting of water and acetonitrile (68/32, v/v). Amikacin was used as an internal standard. The calibration curve was linear over a concentration range of 0.625–15 μg mL^?1. The limit of quantification was 0.45 μg mL^?1. The intra- and inter-day variabilities of ISP were both less than 5%. Both derivatives were stable for at least a week at ambient condition. This assay procedure should have useful application in therapeutic drug monitoring of ISP. The limit of detection was 0.10 μg mL^?1. The specificity, assay linearity, low level assay linearity and assay repeatability were also investigated. The established method provides a reliable bioanalytical method to carry out isepamicin pharmacokinetics in rat plasma.     

甲基原薯蓣皂苷静脉注射给予大鼠后体内代谢物的LC-MS~n分析

采用液相色谱-多级离子阱质谱法(LC-MSn)法,检测大鼠生物样本中的甲基原薯蓣皂苷(MPD)及其代谢产物,以推测MPD在大鼠体内的代谢途径。对大鼠静脉注射给予40 mg/kg的MPD,并收集尿液、血浆、胆汁和粪便等样本,经固相萃取(SPE)净化提取后,采用Phenomenex Gemini C18色谱柱,以甲醇(B)-水(A)为流动相进行梯度洗脱,流速1.0 mL/min。采用正负离子检测。在生物样本中共检测到14个代谢产物。通过与对照品的色谱行为和多级质谱特征相比对,鉴定了其中6个代谢产物,分别为Protodioscin(M3),26-O-β-D-Glucopyrannosyl-(25R)-furan-5-ene-3β,22α,26-trihydroxy-3-O-α-L-rhamnopyranosyl-(1→2 or 4)-β-D–glucopyranoside(M5),MPD(M0),Pseudoprotodioscin(M7),26-O-β-D-Glucopyrannosyl(25R)-furan-5-ene-3β,26-dihydroxy-22-methoxy-3-O-α-L-rhamnopyranosyl-(1→4)-β-D-glucopyranoside(M9),26-O-β-D-Glucopyrannosyl(25R)-furan-5-ene-3β,26-dihydroxy-22-methoxy-3-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranoside(M10)和Dioscin(M11)。根据已有对照品的质谱裂解规律,用LC-MSn法推测了另外5个代谢物。其中Protodioscin(PD)和Dioscin是两个主要的代谢产物,其对应的生成途径是MPD在大鼠体内的主要代谢途径。代谢反应总体以一相代谢为主,主要是水解脱糖,同时检出2个二相代谢产物;代谢物结构变化主要发生在糖上,母核均无明显变化。     

Determination of Tectorigenin in Rat Plasma: Application to a Pharmacokinetic Study After Oral Administration of Tectorigenin or Its Prodrug Tectoridin

A simple, sensitive, and validated liquid chromatographic method has been developed for the determination of tectorigenin in rat plasma and application to a pharmacokinetic study after oral administration of tectorigenin or its prodrug tectoridin. The analysis was performed on a Kromasil C₁₈ analytical column using gradient elution with acetonitrile 0.1% phosphonic acid water at 0.8 mL min⁻¹. The detection wavelength for UV detection was set at 264 nm. The established method was fully validated with parameters as follows: the intra- and inter-day assay precisions (CV) of three analytes were in the range of 4.2–13.3% and accuracies were between 98.0 and 107.5%; the calibration curve was linear with r ² > 0.99 over a concentration range of 0.02–2 μg mL⁻¹; the lower limit of quantification was 0.02 μg mL⁻¹; tectorigenin showed stable in rat plasma after 12 h incubation at room temperature, 15 days storage at −80 °C and three freeze/thaw cycles, as well as in reconstitute buffer for 24 h at 25 °C; and the mean recoveries of tectorigenin were 92.3 ± 3.2, 95.5 ± 2.9 and 94.5 ± 3.0% with quality control levels of 0.02, 0.2 and 2 μg mL⁻¹, respectively. In conclusion, this method is simple, economic, and sensitive enough for in vivo pharmacokinetic studies of tectorigenin.

     

Determination of Tectorigenin in Rat Plasma: Application to a Pharmacokinetic Study After Oral Administration of Tectorigenin or Its Prodrug Tectoridin

A simple, sensitive, and validated liquid chromatographic method has been developed for the determination of tectorigenin in rat plasma and application to a pharmacokinetic study after oral administration of tectorigenin or its prodrug tectoridin. The analysis was performed on a Kromasil C₁₈ analytical column using gradient elution with acetonitrile 0.1% phosphonic acid water at 0.8 mL min^?1. The detection wavelength for UV detection was set at 264 nm. The established method was fully validated with parameters as follows: the intra- and inter-day assay precisions (CV) of three analytes were in the range of 4.2–13.3% and accuracies were between 98.0 and 107.5%; the calibration curve was linear with r ² > 0.99 over a concentration range of 0.02–2 μg mL^?1; the lower limit of quantification was 0.02 μg mL^?1; tectorigenin showed stable in rat plasma after 12 h incubation at room temperature, 15 days storage at ?80 °C and three freeze/thaw cycles, as well as in reconstitute buffer for 24 h at 25 °C; and the mean recoveries of tectorigenin were 92.3 ± 3.2, 95.5 ± 2.9 and 94.5 ± 3.0% with quality control levels of 0.02, 0.2 and 2 μg mL^?1, respectively. In conclusion, this method is simple, economic, and sensitive enough for in vivo pharmacokinetic studies of tectorigenin.     

RP-LC Determination and Pharmacokinetic Study of Ferulic Acid and Isoferulic Acid in Rat Plasma After Taking Traditional Chinese Medicinal-Preparation: Guanxinning Lyophilizer

A sensitive, simple, and accurate method for determination and pharmacokinetic study of ferulic acid and isoferulic acid in rat plasma was developed using a reversed-phase column liquid chromatographic (RP-LC) method with UV detection. Sample preparations were carried out by protein precipitation with the addition of methanol, followed by evaporation to dryness. The resultant residue was then reconstituted in mobile phase and injected into a Kromasil C₁₈ column (250 × 4.6 mm i.d. with 5 μm particle size). The mobile phase was methanol-1% formic acid (33:67, v/v). The calibration plots were linear over the range 5.780–5780 ng·mL^?1 for ferulic acid and 1.740–348.0 ng·mL^?1 for isoferulic acid. Mean recoveries were 85.1% and 91.1%, respectively. The relative standard deviations (RSDs) of within-day and between-day precision were not above 15% for both of the analytes. The limits of quantification were 5.780 ng·mL^?1 for ferulic acid and 1.740 ng·mL^?1 for isoferulic acid. This RP-LC method was used successfully in pharmacokinetic studies of ferulic acid and isoferulic acid in rat plasma after intravenous injection of Guanxinning Lyophilizer.     

RP-LC Determination and Pharmacokinetic Study of Ferulic Acid and Isoferulic Acid in Rat Plasma After Taking Traditional Chinese Medicinal-Preparation: Guanxinning Lyophilizer

A sensitive, simple, and accurate method for determination and pharmacokinetic study of ferulic acid and isoferulic acid in rat plasma was developed using a reversed-phase column liquid chromatographic (RP-LC) method with UV detection. Sample preparations were carried out by protein precipitation with the addition of methanol, followed by evaporation to dryness. The resultant residue was then reconstituted in mobile phase and injected into a Kromasil C₁₈ column (250 × 4.6 mm i.d. with 5 μm particle size). The mobile phase was methanol-1% formic acid (33:67, v/v). The calibration plots were linear over the range 5.780–5780 ng·mL⁻¹ for ferulic acid and 1.740–348.0 ng·mL⁻¹ for isoferulic acid. Mean recoveries were 85.1% and 91.1%, respectively. The relative standard deviations (RSDs) of within-day and between-day precision were not above 15% for both of the analytes. The limits of quantification were 5.780 ng·mL⁻¹ for ferulic acid and 1.740 ng·mL⁻¹ for isoferulic acid. This RP-LC method was used successfully in pharmacokinetic studies of ferulic acid and isoferulic acid in rat plasma after intravenous injection of Guanxinning Lyophilizer.

     

LC Determination of Curcumin in Dog Plasma for a Pharmacokinetic Study

A rapid, simple, and sensitive high-performance liquid chromatographic method for quantification of curcumin in dog plasma has been developed and validated. After addition of the internal standard (berberine), plasma was acidified and extracted with ethyl acetate. Analysis was performed on a C₁₈ column. The mobile phase was acetonitrile–5% acetic acid, 52:48 (v/v) and the flow rate 1.0 mL min^?1. The eluent was monitored at 425 nm. Chromatographic separation was achieved in less than 7 min and the calibration plot was linear over the concentration range 2–128 ng mL^?1. Intra- and inter-assay variability were less than 7.3%. The accuracy ranged from 98.7 to 105.0%. The method was successfully applied to a pharmacokinetic study of curcumin in dogs.     

LC Analysis of Aliphatic Primary Amines and Diamines After Derivatization with 2,6-Dimethyl-4-quinolinecarboxylic Acid N-Hydroxysuccinimide Ester

A method for sensitive simultaneous analysis of aliphatic primary amines and diamines has been developed and validated. The compounds were analyzed by reversed-phase high-performance liquid chromatography after pre-column derivatization with 2,6-dimethyl-4-quinolinecarboxylic acid N-hydroxysuccinimide ester as fluorescent probe. The derivatization reaction was performed at 50 °C for 40 min in 0.1 mol L^?1 borax buffer solution (pH 7.5). The resulting fluorophores were separated to baseline on a C₁₈ column and fluorimetrically detected at λ _ex/λ _em = 326/409 nm. Detection limits were in the range 0.50–0.02 nmol L^?1. The method was successfully used for analysis of aliphatic amines in water, human urine, and serum.     

Preclinical Pharmacokinetics and Bioavailability of Oxypeucedanin in Rats after Single Intravenous and Oral Administration

Oxypeucedanin, a furanocoumarin extracted from many traditional Chinese herbal medicines, has a variety of pharmacological effects. However, the independent pharmacokinetic characteristics and bioavailability of this compound remains elusive. In this study, a rapid, sensitive, and selective method using ultra-high performance liquid chromatography–tandem mass spectrometry (UPLC/MS/MS) was developed for evaluating the intravenous and oral pharmacokinetics of oxypeucedanin. After intravenous administration of oxypeucedanin (2.5, 5, and 10 mg/kg), and intragastric administration of oxypeucedanin (20 mg/kg), blood samples were collected periodically from the tail vein. The plasma concentration-time curves were plotted, and the pharmacokinetic parameters were calculated using a non-compartmental model analysis. After intravenous administration of oxypeucedanin (single dosing at 2.5, 5, and 10 mg/kg) to rats, the pharmacokinetics fit the linear kinetics characteristics, which showed that some parameters including average elimination half-life (T_1/2Z of 0.61~0.66 h), mean residence time (MRT of 0.62~0.80 h), apparent volume of distribution (V_Z of 4.98~7.50 L/kg), and systemic clearance (CL_Z of 5.64~8.55 L/kg/h) are dose-independent and the area under concentration-time curve (AUC) increased in a dose-proportional manner. Single oral administration of oxypeucedanin (20 mg/kg) showed poor and slow absorption with the mean time to reach the peak concentration (T_max) of 3.38 h, MRT of 5.86 h, T_1/2Z of 2.94 h, and a mean absolute bioavailability of 10.26% in rats. These results provide critical information for a better understanding of the pharmacological effect of oxypeucedanin, which will facilitate its research and development.     

LC Method for Determination of Ginkgolic Acids in Mice Plasma and Its Application to a Pharmacokinetic Study

A sensitive and simple LC method for the quantification of ginkgolic acids in mice plasma has been developed. Following acetonitrile deproteinization, samples were separated on a SinoChrom ODS-AP C₁₈ column. The mobile phase was 3% (v/v) acetic acid water solution–methanol (8:92, v/v) at a flow rate of 1.0 mL min⁻¹. Detection was at 310 nm. Calibration curve was linear over the range of 0.25–50 μg mL⁻¹ with intra- and inter-day precisions (RSD%) of less than 9.5%. The extraction recovery ranged from 87.0 to 90.2% (RSD 2.4–6.4%) for ginkgolic acids. The method was successfully applied to the pharmacokinetic study of ginkgolic acids in mice after oral dosing of 1.0 g kg⁻¹.

     

Comparative Pharmacokinetic Study of Forchlorfenuron in Adult and Juvenile Rats

Forchlorfenuron (CPPU) is a plant growth regulator extensively used in agriculture. However, studies on CPPU pharmacokinetics are lacking. We established and validated a rapid, sensitive, and accurate liquid chromatography–mass spectrometry method for CPPU detection in rat plasma. CPPU pharmacokinetics was evaluated in adult and juvenile rats orally treated with 10, 30, and 90 mg/kg of the compound. The area under the plasma drug concentration–time curve from 0 to 24 h (AUC), at the final time point sampled (AUC_0–t), and the maximum drug concentration of CPPU increased in a dose-dependent manner. The pharmacokinetic parameters AUC_0–t and absolute bioavailability were higher in the juvenile rats than in adult rats. The mean residence time and AUC_0–t of juvenile rats in the gavage groups, except for the 10 mg/kg dose, were significantly higher in comparison to those observed for adult rats (p < 0.001). The plasma clearance of CPPU in juvenile rats was slightly lower than that in the adult rats. Taken together, juvenile rats were more sensitive to CPPU than adult rats, which indicates potential safety risks of CPPU in minors.