An UHPLC/LC-MS illustrated the dynamic profiling of balanophorin B, gallic acid, and 4-hydroxycinnamic acid in rat as 3 molecular entities from Balanophora simaoensis.

An UHPLC/LC-MS was founded to detect balanophorin B (B), gallic acid (GA), 4-hydroxycinnamic acid (HC), and their in vivo profiling in rats, after oral administration of the ethanol extract of Balanophora simaoensis S. Y. Chang et Tam. The in vivo dynamic existence of 3 molecular entities in rats and the multistep biotransformation of GA were elucidated by their sensitive mass spectrometry response after efficient UHPLC and/or HPLC separation, through analyzing the bio-samples of rat plasma, bile, liver, kidneys, and excreta. The method was validated with satisfactory calibration curves having correlation coefficients r from 0.996 to 0.999 for concentration scaled from 0.100 nM to 0.100 µM, internal standard normalized matrix factors ranged from 0.923 to 0.993, sextuplicate recoveries valued from 95.0% to 103.6%, as well as accuracy and precision varied from 95.6% to 103.7%. The content of B, GA, and HC in the whole herb was of 4.66, 63.5, and 10.4 µmol/kg in dry weight, respectively. The Cmax for B, GA, and HC in rat systemic circulation was of 76.0 nM, 2.30 µM, and 51.0 µM, with tmax at 3, 2, and 2 h, respectively. B and GA stayed in rat liver over 4 hs to present a material base for the pharmacology and pharmacodynamics of the whole herb. The biotransformation of GA indicated a complicated scheme in rats. As a final metabolite from GA with total biotransformation conversion over 20%, 4-hydroxybenzaldehyde resourced from two steps of dehydroxylation and one step of reduction of GA, but not concerned with HC.

Comprehensive investigation of pharmacodyamic material basis of Wikstroemia indica (L.) C. A. Mey. by serum pharmacochemistry and bivariate correlation analysis.

In this study, the theory of serum pharmacochemistry of traditional Chinese medicine was used to analyze the constituents absorbed into serum after oral administration of Wikstroemia indica (L.) C. A. Mey. by ultra high performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS/MS). The micro-liquid dilution method was used to determine the minimum inhibitory concentration of the serum containing Wikstroemia indica. The bivariate correlation analysis method was used to study the spectral-efficiency relationship between the drug-containing serum and the antibacterial activity, and find the main antibacterial active components in serum containing Wikstroemia indica. A total of 26 serum migration components were identified or speculated in the samples, including 11 prototype components and 15 metabolites. Of which, syringic acid, caffeic acid, dihydrocaffeic acid, 4-hydroxybenzoic acid, hippuric acid, 3-hydroxy-3-(4-hydroxy-3-methoxyphenyl)propanoic acid, triumbelletin, (7R)-3-hydroxy-1-methyl-2-oxo-7-(prop-1-en-2-yl)-2,3,5,6,7,8- hexahydroazulene-4- carbaldehyde and (1S,3aS,8aS)-1,3,5-trihydroxy-1,4-dimethyl-7-(propan-2- ylidene) octahydroazulen-6(1H)-one were bacteriostatic active substances. It is the first time to study the constituents in serum containing Wikstroemia indica and reveal its antibacterial pharmacodyamic material basis. The above works provide scientific reference for the in-depth study of Wikstroemia indica.

Simultaneous determination of gallic acid, methyl gallate, and 1,3,6-tri-O-galloyl-ß-d-glucose from Turkish galls in rat plasma using liquid chromatography-tandem mass spectrometry and its application to pharmacokinetics study.

Turkish galls (TG) is a traditional Uygur medicine typically used in clinics for dental disease and chronic ulcerative colitis. In this study, a novel liquid chromatography-tandem mass spectrometry method was developed and validated for the simultaneous quantification of gallic acid, methyl gallate, and 1,3,6-tri-O-galloyl-ß-d-glucose in rat plasma, which are the major bioactive compounds of TG. After a feasible protein precipitation using acetonitrile for sample preparation, chromatographic separation was performed with a BDS Hypersil C18 column (2.1 × 100 mm, 5 µm) at 30°C, and water containing 10 mmol of ammonium acetate and acetonitrile was used as the mobile phase with a flow rate of 0.3 mL/min. The MS detector was operated in the selective reaction monitoring with negative-ionization mode. The results of the method validation, including selectivity, linearity, accuracy, precision, extraction recovery, matrix effect, and stability of the compounds in the biosamples, were all within the current acceptance criteria. The established method was successfully applied to the pharmacokinetics study of three analytes in rats after an oral administration of TG extract and laid the foundation for studying the active components and mechanism of TG in vivo.

Improved recovery of galloyl metabolites from mango (Mangifera indica L.) in human plasma using protein precipitation with sodium dodecyl sulfate and methanol.

Extraction of polyphenolic metabolites from blood fractions can be challenging since compound recovery can be limited by chemical structure, polarity, and protein-binding affinity of analytes. Gallic acid and its metabolites exhibit particularly low recoveries from plasma and can lead to an underestimation of their bioavailability from foods. A modified method to extract free gallic acid and its metabolites from human plasma aided by sodium dodecyl sulfate and acidified methanol (SDS-MeOH) was applied to extract free gallic acid and its metabolites from human plasma after a single consumption of 400 g of mango (cv. Ataulfo) pulp by 10 healthy male and female subjects. The use of SDS-MeOH facilitated extraction of significantly (p < 0.05) more pyrogallol, free gallic acid, 4-O-methylgallic acid, and ethyl gallate with recovery rates exceeding 80% in standard recovery from human blood plasma when compared to conventional methods that rely on solvent extraction or solid phase extraction. The method was reproducible and precise for standards from 50 to 500 µg/L. In pharmacokinetic plasma samples five predominant metabolites of gallic acid were tentatively characterized by HPLC-MS and absorption kinetics evaluated over 8 h for catechol-O-sulfate, 4-O-methylgallic acid-3-O-sulfate, and pyrogallol-O-sulfate, methylpyrogallol-O-sulfate, and 4-O-methylgallic acid with AUC0-8h of 9520 ± 3370, 6030 ± 1310, 5990 ± 1690, 4020 ± 1040, and 2790 ± 1190 µg/L h respectively. Plasma extraction was rapid and reproducible with superior recovery rates compared to conventional methods when evaluating polar phenolic metabolites.

The profiling and identification of the absorbed constituents and metabolites of Naoshuantong capsule in mice biofluids and brain by ultra- fast liquid chromatography coupled with quadrupole-time-of-flight tandem mass spectrometry.

Naoshuantong capsule (NSTC) is an oral traditional Chinese medicine formula used widely in the clinic for ischemic stroke. The absorbed ingredients and metabolites of NSTC have never been reported before. In this study, a method incorporating rapid resolution liquid chromatography with quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF/MS) was used to identify absorbed ingredients and metabolites after oral administration of NSTC. A total of 15 constituents were detected and identified as prototypes of NSTC. 109 metabolites related to catechin, gallic acid, paeoniflorin, chlorogenic acid, protocatechuate, typhaneoside, ß-elemene, calycosin were identified in serum, urine and brain. 19 metabolites of typhaneoside, 3 metabolites of ß-elemene, 12 metabolites of calycosin were reported for the first time. This is the first time to explore the absorption and metabolism of NSTC. The work will provide helpful information for further research of the mechanism and application of NSTC.

Grape pomace polyphenols improve insulin response to a standard meal in healthy individuals: A pilot study.

BACKGROUND & AIMS: Dietary polyphenols have beneficial effects on glucose/lipid metabolism in subjects at high risk to develop type 2 diabetes; however, the underlying mechanisms are not clear. We aimed to evaluate: 1) the acute effects of the consumption of a drink rich in polyphenols from red grape pomace (RGPD) on glucose/insulin and triglyceride responses to a standard meal in healthy individuals, and, 2) the relationship between plasma levels of phenolic metabolites and metabolic parameters. METHODS: Twelve healthy men, aged 20-40 years participated in a randomized, controlled study according to a cross-over design. After a 3-day low-polyphenol diet, all participants consumed, on two different days and separated by a one week interval, after an overnight fast, a drink rich in polyphenols (1.562 g gallic acid equivalents (GAE)) or a control drink (CD, no polyphenols), followed after 3 h by a standard meal (960 kcal, 18% protein, 30% fat, 52% CHO). Blood samples were taken at fasting, 3 h after the drink, over 5 h after the standard meal and at fasting on the next day to measure plasma concentrations of glucose, insulin, triglyceride and phenolic metabolites. RESULTS: Glycemic and triglyceride post-meal responses were similar after both the RGPD and the control drink. In contrast, postprandial insulin incremental area (iAUC0-5h) was 31% lower (p < 0.05), insulin secretion index was 18% lower (p < 0.016) and insulin sensitivity (SI) index was 36% higher (p = 0.037) after the RGPD compared to CD. Among phenolic metabolites, gallic acid correlated inversely with the insulin response (r = -0.604; p = 0.032) and positively with the SI index (r = 0.588, p = 0.037). CONCLUSIONS: RGPD consumption acutely reduced postprandial insulin levels and improved insulin sensitivity. This effect could be likely related to the increase in gallic acid levels. This drink, added to usual diet, could contribute to increase the daily intake of polyphenols, with potential health benefits. CLINICALTRIALS. GOV IDENTIFIER: NCT02865278.

Simultaneous quantification of fifteen compounds in rat plasma by LC-MS/MS and its application to a pharmacokinetic study of Chaihu-Guizhi decoction.

A simple, sensitive and selective high-performance liquid chromatography electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS) method was developed and validated for simultaneous determination and pharmacokinetic study of 15 active compounds (Saikosaponin A, Baicalin, Wogonin, Glycyrrhizic acid, Glycyrrhetinic acid, Albiflorin, Paeoniflorin, Liquiritin, Isoliquiritin, Liquiritigenin, Isoliquiritigenin, Cinnamic acid, Gallic acid, Wogonoside and Oroxylin A) in rat plasma. After a feasible protein precipitation using methanol for sample preparation, chromatographic separation was carried out with a Halo® C18 column (2.1 × 100 mm, 2.7 µm) at 35 °C, water containing 0.1% formic acid and acetonitrile were used as the mobile phase with a flow rate of 0.3 mL/min. Multiple reaction monitoring (MRM) with positive and negative ion switching mode was performed for the quantification of the standards and internal standard in plasma. All the calibration curves showed good linear regression within the linear range (r2 > 0.9923). In particular, the results of the method validation including specificity, linearity, accuracy, precision, extraction recovery, matrix effect, and stability of compounds in bio-samples were all within the current acceptance criteria. The established method was successfully applied to the pharmacokinetic study of 15 compounds in rats after oral administration of CGD and laid the foundation for studying the active components and mechanism of CGD in vivo.

Bioavailability Study of Maqui Berry Extract in Healthy Subjects.

Several health promoting effects have been reported for maqui berry, rich in anthocyanins. Direct effects of anthocyanins as well as bioactive metabolites might be involved. Within the study, bioavailability of a proprietary standardized maqui berry extract Delphinol® was investigated based on two selected anthocyanins (delphinidin-3-O-glucoside (DS) + cyanidin-3-O-sambubioside (CS)) and two breakdown products (protocatechuic acid (PCA) + gallic acid (GA)) after a single-dose supplementation in humans. Pharmacokinetic parameters were calculated from individual concentration time curves. In all 12 subjects a significant increase was noted in plasma values of DG and CS after intake of maqui berry extract. Maximum concentration of DG was observed after 1.0 ± 0.3 h and CS after 2.0 ± 1.1 h. Within 8 h, concentrations nearly returned to baseline levels. The results confirm a fast uptake and metabolism of the two selected key substances. Additionally, the phenolic acids GA and PCA were observed as breakdown products of anthocyanins. In summary, the study clearly confirms the bioavailability of maqui berry extract and its specific anthocyanin compounds and related breakdown products in healthy subjects.

Investigation of serum amino acids involved in gallic acid detoxification of formaldehyde by liquid chromatography/tandem mass spectrometry and neutral loss scan.

RATIONALE: Gallic acid is one of the most common polyphenols in natural products and human diet. The consumption of gallic acid reduces the incidence of cardiovascular diseases, chronic metabolic disorders and cancers. Most previous publications focused on the antioxidative or prooxidative properties of gallic acid. In the present work, gallic acid as a trapping agent of blood formaldehyde was investigated by liquid chromatography/tandem mass spectrometry (LC/MS/MS) and neutral loss scan. METHODS: Serum samples incubated with gallic acid were subjected to LC/MS/MS analysis using an LTQ XL ion trap mass spectrometer. The adduct ions of gallic acid-formaldehyde-amino acids were explored by investigation of their fragmentation patterns and neutral loss scan experiments. RESULTS: A series of Mannich adducts (namely, gallic acid-formaldehyde-alanine, gallic acid-formaldehyde-proline, gallic acid-formaldehyde-leucine or gallic acid-formaldehyde-isoleucine and gallic acid-formaldehyde-phenylalanine) were identified as metabolites by neutral loss scan experiments. CONCLUSIONS: This work demonstrated that serum amino acids are involved in gallic acid detoxification of formaldehyde. Because excessive formaldehyde in blood is implicated in a variety of disease pathologies, detoxification of formaldehyde, especially endogenous formaldehyde, may be another health beneficial effect of gallic acid. It also suggested that more attention should be paid to Mannich-type metabolites of polyphenol-formaldehyde-amino acids in research into the pharmacokinetics and bioavailability of polyphenols.

Identification and characterization of in vitro inhibitors against UDP-glucuronosyltransferase 1A1 in uva-ursi extracts and evaluation of in vivo uva-ursi-drug interactions.

Uva-ursi leaf is widely used to treat symptoms of lower urinary tract infections. Here, we evaluated the in vitro inhibitory effects of uva-ursi extracts on 10 major human UDP-glucuronosyltransferases (UGT) isoforms. Of the 10 tested UGT isoforms, uva-ursi extracts exerted the strongest inhibitory effect on UGT1A1-mediated ß-estradiol 3-glucuronidation with the lowest IC50 value of 8.45 ±â€¯1.56 µg/mL. To identify the components of uva-ursi extracts showing strong inhibitory effects against UGT1A1, the inhibitory effects of nine major constituents of the extracts were assessed. Among the tested compounds, gallotannin exerted the most potent inhibition on UGT1A1, followed by 1,2,3,6-tetragalloylglucose; both demonstrated competitive inhibition, with Ki values of 1.68 ±â€¯0.150 µM and 3.55 ±â€¯0.418 µM. We found that gallotannin and 1,2,3,6-tetragalloylglucose also inhibited another UGT1A1-specific biotransformation, SN-38-glucuronidation, showing the same order of inhibition. Thus, in vitro UGT1A1 inhibitory potentials of uva-ursi extracts might primarily result from the inhibitory activities of gallotannin and 1,2,3,6-tetragalloylglucose present in the extracts. However, in rats, co-administration with uva-ursi extracts did not alter the in vivo marker for UGT1A1 activity, expressed as the molar ratio of AUCSN-38 glucuronide/AUCSN-38, because plasma concentrations of gallotannin and 1,2,3,6-tetragalloylglucose may be too low to inhibit the UGT1A1-mediated metabolism of SN-38 in vivo. The poor oral absorption of gallotannin and 1,2,3,6-tetragalloylglucose in uva-ursi extracts might cause the poor in vitro-in vivo correlation. These findings will be helpful for the safe and effective use of uva-ursi extracts in clinical practice.

Fabrication of size controlled nanocomposite based on zirconium alkoxide for enrichment of Gallic acid in biological and herbal tea samples.

In this work, novel water-dispersible size controlled nanocomposite based on zirconium alkoxide as metal organic precursor was fabricated and subsequently applied for rapid, efficient and selective preconcentration of gallic acid in human plasma and herbal tea samples. The resultant nanocomposite (Fe3O4@Zr(OtBu)4@Laurate) was characterized by Fourier transform infrared, scanning electron microscope and X-ray spectrometer, while laurate forms aggregates on the surface of the nanocomposite and thereby improves sorption of gallic acid. The effects of some variables on efficiency of gallic acid from real samples were optimized by central composite design; while optimum points were achieved as follows: pH 3.5, 35.0 mg of nanocomposite, 150.0 µL of eluent and 3.0 min sonication time. Chromatographic separation was carried out on analytical Nucleosil C18 column (250 × 4.6 mm I.D., 5 µm particle size) at ambient temperature with methanol: water (40:60, v/v) pH adjusted at 3.5 follow by UV detection at 270 nm. Acceptable limit of detection (0.2 µg kg-1) and wide linear range (1.0-700.0 µg kg-1) in coincidence with reasonable enrichment factor (EF = 125) are the unique advantages, which promise this method for quantification of this compound in real samples with complicated matrices.

A C8-Modified Graphene@mSiO2 Composites Based Method for Quantification of Gallic Acid in Rat Plasma after Oral Administration of Changtai Granule and Its Application to Pharmacokinetics.

A rapid, effective extraction technique has been established for measuring the gallic acid in rat plasma by using sandwich-structured graphene/mesoporous silica composites with C8-modified interior pore-walls as adsorbent. The unique characteristics of the graphene-silica composites excluded large molecules, like proteins, from the mesopore channels as a result of size exclusion effect, leading to a direct extraction of drug molecules from protein-rich biological samples such as plasma without any other pretreatment procedure. Followed by elution and centrifugation, the gallic acid-absorbed composites were rapidly isolated before LC-MS/MS. Serving as a reliable tool for analysis of Traditional Chinese Medicine: Changtai Granule, the newly developed method was fully validated and successfully applied in the pharmacokinetic study of gallic acid in rat plasma. Extraction recovery, matrix effect and stability were satisfactory in rat plasma. According to the results of pharmacokinetic studies, Changtai Granule exhibited greater adsorption, distribution and clearance properties of gallic acid in the treatment of ulcerative colitis. Hence, this study may offer a valuable alternative to simplify and speed up sample preparation, and be useful for clinical studies of related preparations.

UHPLC-MS/MS method for the simultaneous quantitation of five anthraquinones and gallic acid in rat plasma after oral administration of prepared rhubarb decoction and its application to a pharmacokinetic study in normal and acute blood stasis rats.

Prepared rhubarb, as one of the main processed products of rhubarb, has a good effect on promoting blood circulation. In this paper we describe a rapid, sensitive, and selective ultra-fast liquid chromatography with tandem mass spectrometry method for simultaneous quantification of five anthraquinones (rhein, aloe-emodin, chrysophanol, emodin, and physcion) and gallic acid in plasma. Chromatographic separation was performed on an Extend C18 column at the temperature of 30°C using a mobile phase that consisted of 0.1% aqueous formic acid and acetonitrile. Satisfactory linearity, precision, accuracy, extraction recovery, and matrix effect have been achieved. Then, the validated method was successfully applied to a comparative pharmacokinetic study. The results might be helpful for guiding clinical application of prepared rhubarb in the future.

A UPLC-MS/MS Method for Simultaneous Determination of Free and Total Forms of a Phenolic Acid and Two Flavonoids in Rat Plasma and Its Application to Comparative Pharmacokinetic Studies of Polygonum capitatum Extract in Rats.

The principal active constituents of Polygonum capitatum are phenolic acids and flavonoids, such as gallic acid, quercitrin, and quercetin. The aim of this study was to develop and validate a method to determine the three constituents and the corresponding conjugated metabolites of Polygonum capitatum in vivo and to conduct pharmacokinetic studies on the herb, a well-known Miao medicinal plant in China. Gallic acid, quercitrin, and quercetin were analysed by ultra-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS). Protein precipitation in plasma samples was performed using methanol. For the determination of total forms of analytes, an additional process of hydrolysis was conducted using ß-glucuronidase and sulphatase. The analytes were separated on a BEH C18 column (50 mm × 2.1 mm; i.d., 1.7 µm) and quantified by multiple reaction monitoring (MRM) mode. The linear regression showed high linearity over a 729-fold dynamic range for the three analytes. The relative standard deviations of intra- and inter-day measurements were less than 9.5%, and the method was accurate to within -11.1% to 12.5%. The extraction recoveries for gallic acid, quercitrin, and quercetin were 94.3%-98.8%, 88.9%-98.8%, and 95.7%-98.5%, respectively. All samples were stable under short- and long-term storage conditions. The validated method was successfully applied to a comparative pharmacokinetic study of gallic acid, quercitrin, and quercetin in their free and total forms in rat plasma. The study revealed significantly higher exposure of the constituents in total forms for gallic acid and quercetin, while quercitrin was detected mainly in its corresponding free form in vivo. The established method was rapid and sensitive for the simultaneous quantification of free and total forms of multiple constituents of Polygonum capitatum extract in plasma.

Effects of a diet containing dried grape pomace on blood metabolites and milk composition of dairy cows.

BACKGROUND: The effect of a diet containing 15% grape pomace (GP) on the general health status and milk quality of dairy cows was assessed by plasma biochemistry and total polyphenol (TP) content, milk polyphenols, milk composition and milk protein fractions. RESULTS: Among the polyphenols measured by liquid chromatography-mass spectroscopy in GP, in feed containing GP (GP+) or not containing GP (GP-), gallic acid and epicatechin were present in the highest concentrations (67.58 and 19.23 µg mL-1 , respectively). Higher amounts of TP were also detected in the blood plasma of GP+ cows (114.06 and 83.93 mg GAE L-1 , respectively) but not in their milk (233.17 and 245.75 mg GAE L-1 , respectively). Also a significant increase was found for lactose and ß-lactoglobulin, although there was no effect on α-lactalbumin, albumin, secretory components and caseins. CONCLUSION: Inclusion of 15% GP in the diets of dairy cows is beneficial for overall normal blood constituent metabolism and helps to maintain cow health. The milk of cows fed with a GP diet preserves the normal levels of fat, protein and caseins, and has increased levels of components that make this milk a versatile ingredient material for the food industry (e.g. model whey powders, stability of lactose-rich powders). © 2016 Society of Chemical Industry.

Serum pharmacochemistry for tracking bioactive components by UPLC-Q-TOF-MS/MS combined chromatographic fingerprint for quality assessment of Sanziguben Granule.

To more reasonably and effectively control the quality of Sanziguben Granule, chromatographic fingerprinting and serum pharmacochemistry of this traditional Chinese medicine compound were performed. A comprehensive comparison and evaluation of 15 batches of Sanziguben Granule was successfully conducted by using high performance liquid chromatography (HPLC) fingerprint analysis. After administering a set amount of Sanziguben Granule orally to rats, blood samples were collected and tested 4 times at intervals of 30min, 1h, 2h, and 4h using UPLC-Q-TOF-MS/MS. The blood showed presence of gallic acid and corilagin indicating the pharmacological significance of these two chemical compounds. According to the result, above mentional chemical compounds were designated biomarkers for quality control of Sanziguben Granule. Therefore, a purposeful and efficient method for quality control of Sanziguben Granule was established in the present study.

A validated liquid chromatography-tandem mass spectrometry method for the determination of methyl gallate and pentagalloyl glucopyranose: application to pharmacokinetic studies.

Methyl gallate (MG) and pentagalloyl glucopyranose (PGG) are bioactive phenolic compounds that are widely distributed in herbs and plant foods. Their potential activities include anti-oxidant, anti-inflammatory, anti-cancer, anti-bacterial and anti-viral activities. However, knowledge concerning the pharmacokinetic characteristics of MG and PGG is limited. The purpose of this study was to develop a sensitive and reproducible ultra-performance liquid chromatography-tandem mass spectrometric (UPLC-MS/MS) method to simultaneously quantify MG and PGG in rat blood samples. The linear response ranges for MG and PGG were 0.0195-20 and 0.0390-20 µM, respectively. The lower limit of quantification was 0.0195 µM for MG and 0.0390 µM for PGG. The intra- and inter-day variances were less than 15%, and accuracy was within 80-120%. This assay was successfully applied to pharmacokinetic studies in Sprague-Dawley rats after intraperitoneal administration of MG and PGG (20 mg/kg). The values of areas under the blood concentration time curves (AUC0₋24 h) for MG and PGG were 109.9 ± 73.40 and 38.78 ± 24.53 h*µM, respectively. The maximum blood concentrations (Cmax) of MG and PGG were 34.72 ± 17.32 and 6.39 ± 4.25 µM, respectively. The time required to reach the maximum concentration (Tmax) was 0.85 ± 0.70 h for both MG and PGG. The values of the elimination rate constant (Ke), elimination half-life (t1/2), volume of distribution (Vd), clearance (Cl) and mean resident time (MRTlast) were 0.056 ± 0.032 h(-1), 17.50 ± 12.25 h, 530.95 ± 247.54 L/kg, 159.91±76.05L/h/kg, 8.71 ± 2.53 h for MG and 0.023 ± 0.012 h(-1), 38.66 ± 22.89 h, 7838.89 ± 3474.72 L/kg, 30.98 ± 21.73 L/h/kg, 12.47 ± 2.77 h for PGG, respectively. In conclusion, a UPLC-MS/MS method was fully validated over a wide linear range and used to quantify the levels of MG and PGG in pharmacokinetic studies of MG and PGG in rats. The main advantages of this method are the use of small blood volumes (10 µL), rapid analysis (5 min) and excellent recoveries.

An UHPLC-MS/MS method for simultaneous quantification of gallic acid and protocatechuic acid in rat plasma after oral administration of Polygonum capitatum extract and its application to pharmacokinetics.

ETHNOPHARMACOLOGICAL RELEVANCE: Polygonum capitatum Buch.-Ham. ex D. Don has been traditionally used by Hmong for the treatments of urinary tract infections and pyelonephritis. Gallic acid (GA) and protocatechuic acid (PCA) are regarded as two of the main bioactive compounds in the herb. MATERIALS AND METHODS: A rapid, selective and sensitive UHPLC-ESI-MS/MS method was established and validated for the quantification of GA and PCA in rat plasma after oral administration of P. capitatum extract. Concentrations of GA and PCA were determined at different time points after dosing 20 mg/kg (equivalent to 4 mg/kg of GA and 0.3 mg/kg of PCA), 60 mg/kg and 120 mg/kg of P. capitatum extract. The main pharmacokinetic parameters of GA and PCA were obtained based on the analysis of the plasma sample by non-compartmental analysis. RESULTS: After oral administration of P. capitatum extract, GA and PCA were quickly absorbed and showed a dose-dependent profile. Pharmacokinetic parameters for GA and PCA following oral administration of the extract were respectively: Cmax 246.24-806.27 and 15.73-30.72 ng/mL; Tmax 40-100 and 20-40 min. In the rats treated with P. capitatum t1/2 and Tmax of GA were prolonged by comparing with that of its pure form. CONCLUSION: Other compounds in P. capitatum extract may be metabolized to GA, which affected the pharmacokinetic profiles of GA. This pharmacokinetic study seems to be useful for a further clinical study of P. capitatum extract.

A UHPLC-MS/MS method for simultaneous determination of six flavonoids, gallic acid and 5,8-dihydroxy-1,4-naphthoquinone in rat plasma and its application to a pharmacokinetic study of Cortex Juglandis Mandshuricae extract.

Cortex Juglandis Mandshuricae is used as a folk remedy for treating cancer, diarrhea and dysentery in traditional Chinese medicine for many years. Six flavonoids (myricitrin, quercitrin, taxifolin, myricetin, quercetin and naringenin), gallic acid and 5,8-dihydroxy-1,4-naphthoquinone are major bioactive components in Cortex Juglandis Mandshuricae extract. In this study, an ultrahigh performance liquid chromatography and tandem mass spectrometry method was developed for simultaneous determination of eight ingredients in rat plasma using chloromycetin as an internal standard. Plasma samples added vitamin C (antioxygen) were acidified with hydrochloric acid and extracted by liquid-liquid extraction with ethyl acetate. Eight ingredients were separated on a Venusil ASB C18 column and detected by multiple reaction monitoring mode using electrospray ionization in the negative ion mode. The method was linear for all analytes over investigated range with all correlation coefficients greater than 0.9900. The validated lower limit of quantification was 20ng/mL for gallic acid, 5ng/mL for myricitrin, 3ng/mL for quercitrin, 10ng/mL for taxifolin, 6ng/mL for myricetin, 3ng/mL for quercetin, 2ng/mL for naringenin and 1µg/mL for 5,8-dihydroxy-1,4-naphthoquinone, respectively. Intra- and inter-day precisions (RSD%) were less than 15% and accuracy (RE%) ranged from -6.9% to 6.9%. The validated method was successfully applied to investigate the pharmacokinetics of the eight analytes after oral administration of Cortex Juglandis Mandshuricae extract to rats.

Gallic acid functions as a TRPA1 antagonist with relevant antinociceptive and antiedematogenic effects in mice.

The transient receptor potential ankyrin 1 (TRPA1) has been identified as a relevant target for the development of novel analgesics. Gallic acid (GA) is a polyphenolic compound commonly found in green tea and various berries and possesses a wide range of biological activities. The goal of this study was to identify GA as a TRPA1 antagonist and observe its antinociceptive effects in different pain models. First, we evaluated the ability of GA to affect cinnamaldehyde-induced calcium influx. Then, we observed the antinociceptive and antiedematogenic effects of GA (3-100 mg/kg) oral administration after the intraplantar (i.pl.) injection of TRPA1 agonists (allyl isothiocyanate, cinnamaldehyde, or hydrogen peroxide-H2O2) in either an inflammatory pain model (carrageenan i.pl. injection) or a neuropathic pain model (chronic constriction injury) in male Swiss mice (25-35 g). GA reduced the calcium influx mediated by TRPA1 activation. Moreover, the oral administration of GA decreased the spontaneous nociception triggered by allyl isothiocyanate, cinnamaldehyde, and H2O2. Carrageenan-induced allodynia and edema were largely reduced by the pretreatment with GA. Moreover, the administration of GA was also capable of decreasing cold and mechanical allodynia in a neuropathic pain model. Finally, GA was absorbed after oral administration and did not produce any detectable side effects. In conclusion, we found that GA is a TRPA1 antagonist with antinociceptive properties in relevant models of clinical pain without detectable side effects, which makes it a good candidate for the treatment of painful conditions.