Increase water solubility of Centella asiatica extract by indigenous bioenhancers could improve oral bioavailability and disposition kinetics of triterpenoid glycosides in beagle dogs.

A newly standardised extract of Centella asiatica (Centell-S) with better water solubility than the previous standardised extract of C. asiatica (ECa 233) was developed, and pharmacokinetic profiles of bioactive triterpenoids were investigated in beagle dogs. The test substances were administered via intravenous or oral administration with single and multiple doses for 7 days. The concentrations of major bioactive triterpenoids, including madecassoside, asiaticoside, madecassic acid, and asiatic acid, in biological samples were measured by liquid chromatography-tandem mass spectrometry. The dogs in this study showed good tolerability to all test substances, based on the physical appearance and blood chemistry 24 h after dosing. The major bioactive triterpenoids found in systemic blood circulation were madecassoside, asiaticoside, and asiatic acid; the concentration of these components ranged from 1 to 10,000 µg/L after intravenous administration of 1.0 mg/kg Centell-S. Oral administration of 10 and 20 mg/kg Centell-S generated approximately twofold higher plasma levels of both madecassoside and asiaticoside compared with equivalent doses of ECa 233. In addition, there was an accumulation of triterpenoid glycosides after multiple oral administrations of Centell-S for 7 days, while triterpenic acids showed little tendency for accumulation. Beagles had good tolerability to both standardised extracts of C. asiatica, and showed a similar pattern of bioactive triterpenoids to humans. Centell-S increased oral bioavailability of major triterpenoid glycosides and can be further developed into a phytopharmaceutical product.

Anti-Diabetic Potential of Plant-Based Pentacyclic Triterpene Derivatives: Progress Made to Improve Efficacy and Bioavailability.

Diabetes mellitus (DM) results from the inability of the pancreas to produce sufficient insulin or weakened cellular response to the insulin produced, which leads to hyperglycemia. Current treatments of DM focus on the use of oral hypoglycemic drugs such as acarbose, alpha-glucose inhibitors, sulphonylureas, thiazolidinediones, and biguanides to control blood glucose levels. However, these medications are known to have various side effects in addition to their bioavailability, efficacy, and safety concerns. These drawbacks have increased interest in the anti-diabetic potential of plant-derived bioactive compounds such as oleanolic and maslinic acids. Although their efficacy in ameliorating blood glucose levels has been reported in several studies, their bioavailability and efficacy remain of concern. The current review examines the anti-diabetic effects of oleanolic, maslinic, asiatic, ursolic, and corosolic acids and their derivatives, as well as the progress made thus far to enhance their bioavailability and efficacy. The literature for the current review was gathered from leading academic databases-including Google Scholar and PubMed-the key words listed below were used. The literature was searched as widely and comprehensively as possible without a defined range of dates.

Lupeol and its derivatives as anticancer and anti-inflammatory agents: Molecular mechanisms and therapeutic efficacy.

Lupeol is a natural triterpenoid that widely exists in edible fruits and vegetables, and medicinal plants. In the last decade, a plethora of studies on the pharmacological activities of lupeol have been conducted and have demonstrated that lupeol possesses an extensive range of pharmacological activities such as anticancer, antioxidant, anti-inflammatory, and antimicrobial activities. Pharmacokinetic studies have indicated that absorption of lupeol by animals was rapid despite its nonpolar characteristics, and lupeol belongs to class II BCS (biopharmaceutics classification system) compounds. Moreover, the bioactivities of some isolated or synthesized lupeol derivatives have been investigated, and these results showed that, with modification to C-3 or C-19, some derivatives exhibit stronger activities, e.g., antiprotozoal or anticancer activity. This review aims to summarize the advances in pharmacological and pharmacokinetic studies of lupeol in the last decade with an emphasis on its anticancer and anti-inflammatory activities, as well as the research progress of lupeol derivatives thus far, to provide researchers with the latest information, point out the limitations of relevant research at the current stage and the aspects that should be strengthened in future research.

Preparation and evaluation of PEGylated asiatic acid nanostructured lipid carriers on anti-fibrosis effects.

Liver fibrosis is a major pathological feature of chronic liver diseases, and effective therapies are limited at present. Asiatic acid (AA) is a triterpenoid isolated from Centella asiatica, which exhibits efficient anti-inflammatory and anti-oxidative activities. However, AA shows very low plasma levels after oral administration. In this study, AA loading PEGylated nanostructured lipid carriers (P-AA-NLCs) were prepared. P-AA-NLCs were characterized for particle size distribution, polydispersity index, entrapment efficiency, X-ray powder diffraction (XRD) pattern, differential scanning colorimeter (DSC), and transmission electron microscopy (TEM). The intestinal absorption, in vivo distribution, pharmacokinetics, and anti-fibrosis effects of P-AA-NLC were studied compared with that of AA-NLC. In situ single-pass intestinal perfusion model shows that there are significant differences in absorption between the free and NLCs formulation. The Peff values of P-AA-NLC were significantly enhanced in all four intestinal segments compared to AA-NLC and free AA (p < .05). fa% and Ka showed similar trends, suggesting the PEGylated NLC can improve the gastrointestinal absorption of the drug. The pharmacokinetic studies presented that P-AA-NLC prolonged blood circulation times with a 1.5-fold higher relative bioavailability compared with AA-NLC. In vivo distribution experiments demonstrated that the fluorescence concentration in the liver was higher than that in other organs and the fluorescence intensity in the liver of DIR-P-NLC was about 1.3 times that of DIR-NLC. In addition, oral administration of P-AA-NLC can significantly attenuate CCl4-induced liver fibrosis and functional impairment in a dosage-dependent manner, including an increase in the albumin (ALB) and decrease in aspartate aminotransferase (AST) and alanine transaminase (ALT). Moreover, the MDA and HYP in liver tissue were downregulated, while the SOD activity was upregulated. In conclusion, P-AA-NLC can increase gastrointestinal absorption of AA and enhance anti-liver fibrosis effects in SD rats.

Selection and evaluation of quality markers from Yinlan capsule and its LXRα-mediated therapy for hyperlipidemia.

BACKGROUND: The selection of active compounds for the quality evaluation of traditional Chinese medicine (TCM), specifically complex formulas, remains a challenge for researchers, as components selected as indexes usually have no clear relation with the therapeutic effects of interest. As a suggested resolution, quality control markers (Q-markers) showed good perspective for discriminating numerous compounds found for specific efficacies. In the presented study, the components of the Yinlan (YL) capsule, a TCM patent formula comprising four ingredients, were evaluated and selected for their lipid regulatory effects using principles for Q-marker selection. PURPOSE: The mechanism of TCM therapeutic effects involves several pathways and targets that combine to become an integrated action in the body. Therefore, it is assumed that specific compounds in YL should have good affinity for related targets and obvious effects (both up- and downregulating). Thus, a series of experiments, including cytobiology, animal-based pharmacodynamics, computer-assisted drug design, conventional content determination and pharmacokinetics, would be helpful for the selection and final confirmation of Q-markers. METHODS: The capsule was first administered to Wistar mice fed a high-fat diet and tested for their triglycerides (TG) and total cholesterol (TC) values to evaluate the effectiveness of YL. Then, liver tissue was extracted for gene expression. According to the results, the compounds in YL with good affiliation were selected and determined using UHPLC-MS-MS, and those with adequate results in the capsule were chosen as Q-marker candidates. Finally, pharmacokinetics research was performed; the candidates with desirable metabolite and bioavailability parameters were confirmed as Q-markers of YL. RESULTS: YL capsule was capable of lowering TG and TC levels. For target selection, the expression of LXR mRNA increased significantly at all three tested dosages. Downstream genes, such as LCAT, CYP7A1, and ABCA1, and intestinal FXR mRNA also showed significant increases in expression. For screening of the Q-marker candidates, 5 compounds were selected according to abovementioned results. The pharmacokinetics research demonstrated that the rats exploited lupeol and ginsenoside Rb3 in a desirable pattern with adequate bioavailability, which confirmed their roles as lipid regulatory Q-markers. CONCLUSION: The YL capsule was demonstrated to have obvious lipid regulatory effects, which are mainly exerted by targeting LXR and its related pathway. Lupeol and ginsenoside Rb3 were validated as Q-markers that represent the anti-hyperlipidemia activity of the capsule.

Fabrication of peptide-linked albumin nanoconstructs for receptor-mediated delivery of asiatic acid to the brain as a preventive measure in cognitive impairment: optimization, in-vitro and in-vivo evaluation.

The aim of the study was to evaluate the neuroprotective activity of glutathione (GU)-conjugated asiatic acid (AA) loaded albumin nanoparticles and establishing the drug targeting efficiency (DTE) of GU as a selective ligand for brain-targeted delivery. Albumin nanoparticles were prepared by desolvation technique and optimized using quality by design (QbD) approach. GU was conjugated with nanoparticles by carbodiimide reaction and characterized by its size and zeta potential using dynamic light scattering phenomenon. Dialysis bag technique was employed for in-vitro release study and in-vivo brain targeting efficiency was evaluated in Sprague-Dawley rats (75 mg/kg, i.p.). Neuroprotective activity was evaluated against scopolamine-induced dementia in rats. Resultant brain bioavailability of nanoparticles with 100.2 nm size and 71.59% entrapment efficiency (EE), was found 7-fold higher than AA dispersion with 293% DTE for the brain. Conjugated nanoparticles showed significantly high percentage correct alternation (p < .05), low escape latency time (p < .01), cholinesterase inhibition (p < .01) and ameliorated GU levels (p < .01) as compared to diseased animals. GU showed potential to enhance the brain delivery of AA with ameliorated neuroprotective activity due to enhanced bioavailability. This concept can serve as a platform technology for similar potential neurotherapeutics, whose clinical efficacy is still challenging owing to poor bioavailability.

Assessment of Toxicity and Absorption of the Novel AA Derivative AA-Pme in SGC7901 Cancer Cells In Vitro and in Zebrafish In Vivo.

BACKGROUND Asiatic acid (AA; 2α,3ß,23-trihydroxyurs-12-ene-28-oic acid) is an active compound derived from Centella asiatica, a traditional medicinal plant used widely in many Asian countries, particularly for the treatment of cancer. However, the modified AA derivative N-(2α,3ß,23-acetoxyurs-12-en-28-oyl)-l-proline methyl ester (AA-PMe) has shown markedly better anti-tumor activity than AA. MATERIAL AND METHODS We evaluated the toxicity of AA and AA-PMe on zebrafish morphology, mortality, and hatching rate and determined the effect on SGC7901 cancer cells by acute toxicity assay. AA-PMe absorption in vitro in SGC7901 cells and in vivo in zebrafish was determined by establishing a highly accurate and reproducible HPLC protocol. RESULTS In zebrafish, the toxicity of AA-PMe was lower than AA, with an acute toxic dose of AA-PMe above 25 µM, compared to acute toxicity at doses above 10 µM for AA. However, chronic toxicity of AA-PMe began occurring at doses below 25 µM but became apparent for AA at doses below 10 µM. Although low doses of AA-PMe were tolerated acutely, it became chronically toxic during zebrafish development, resulting in morphological abnormalities, including peripheral and abdominal edema, hemorrhage, abnormal body shape, enlarged yolk sac, and reduced motility. At low concentrations, absorption of AA-PMe by cells and zebrafish embryos occurred in a dose-dependent manner, but this stabilized as the concentration increased. CONCLUSIONS This pharmacokinetic study outlines the cellular and organismal effects of AA-PMe and suggests a theoretical basis that may underlie its mechanism of action.

[Oral absorption of asiatic acid nanoparticles modified with PEG].

A solvent diffusion method was used to prepare pegylated asiatic acid (AA) loaded nanostructured lipid carriers (p-AA-NLC), and the ligated intestinal circulation model was established to observe the absorption and distribution in small intestine. The concentration of AA in bile after oral administration of p-AA-NLC was detected by HPLC in healthy SD rats to indirectly evaluate the oral absorption promoting effect of PEG-modified namoparticles. The results showed that the penetration of p-AA-NLC was enhanced significantly and the transport capacity was increased greatly in small intestinal after PEG modification. As compared with the normal nanoparticles (AA-NLC), the Cmax of the drug excretion was increased by 76%, the time to reach the peak (tmax ) was decreased and the elimination half-life t1/2 was doubled in the rats after oral administration of p-AA-NLC, and the AUC0→t was 1.5 times of the AA-NLC group, indicating that the oral bioavailability of AA-NLC was significantly improved by hydrophilic modification of PEG.

Pentacyclic Triterpene Bioavailability: An Overview of In Vitro and In Vivo Studies.

Pentacyclic triterpenes are naturally found in a great variety of fruits, vegetables and medicinal plants and are therefore part of the human diet. The beneficial health effects of edible and medicinal plants have partly been associated with their triterpene content, but the in vivo efficacy in humans depends on many factors, including absorption and metabolism. This review presents an overview of in vitro and in vivo studies that were carried out to determine the bioavailability of pentacyclic triterpenes and highlights the efforts that have been performed to improve the dissolution properties and absorption of these compounds. As plant matrices play a critical role in triterpene bioaccessibility, this review covers literature data on the bioavailability of pentacyclic triterpenes ingested either from foods and medicinal plants or in their free form.

Development of a UPLC-TQ/MS Approach for the Determination of Eleven Bioactive Components in Haizao Yuhu Decoction Plus-Minus Haizao and Gancao Drug Combination after Oral Administration in a Rat Model of Hypothyroidism.

Haizao Yuhu Decoction (HYD) has been used for approximately 500 years and is well-known in Traditional Chinese Medicine for its efficacy in the treatment of thyroid-related diseases. In this study, a rapid liquid chromatography-tandem mass spectrometry method was developed for the determination of liquiritin, naringin, hesperidin, peimine, liquiritigenin, glycyrrhizic acid, bergapten, nobiletin, osthole, and glycyrrhetinic acid in rat plasma to investigate the pharmacokinetic profile of different HYD prescriptions in a rat model of hypothyroidism. The differences in pharmacokinetic parameters among the groups were compared by Student's t-test. The pharmacokinetic profile of liquiritin, naringin, hesperidin, peimine, liquiritigenin, glycyrrhizic acid, bergapten, nobiletin, osthole, and glycyrrhetinic acid showed significant differences between Haizao and Gancao anti-drug combination and other herbs in HYD. These results may contribute to the rational clinical use of HYD and reveal the compatibility profile of the Haizao and Gancao anti-drug combination.

Structural analysis of metabolites of asiatic acid and its analogue madecassic acid in zebrafish using LC/IT-MSn.

Although zebrafish has become a significant animal model for drug discovery and screening, drug metabolism in zebrafish remains largely unknown. Asiatic acid (AA) and madecassic acid (MA), two natural pentacyclic triterpenoids mainly obtained from Centella asiatica (L.) Urban, have been found to possess many pharmacological effects. This study is to probe the metabolic capability of zebrafish via investigation of the drug metabolism of AA and MA in zebrafish, using a sensitive LC/IT-MSn method. In addition, the main fragmentation pathways of AA and MA were reported for the first time. Nineteen metabolites of AA and MA were firstly identified after zebrafish was exposed to the drug, which all were the phase I metabolites and mainly formed from hydroxylation, dehydrogenation, hydroxylation and dehydrogenation, dihydroxylation and dehydrogenation, and dehydroxylation reaction. The results indicated that zebrafish possessed strong metabolic capacity, and the metabolites of AA and MA were formed via similar metabolic pathways and well matched with the known metabolic rules in vivo and in vitro, which supports the widely use of this system in drug metabolism research. This investigation would also contribute to the novel information on the structural elucidation, in vivo metabolites and metabolic mechanism of pentacyclic triterpenoids.

Biopharmaceutical and pharmacokinetic characterization of asiatic acid in Centella asiatica as determined by a sensitive and robust HPLC-MS method.

ETHNOPHARMACOLOGICAL RELEVANCE: Asiatic acid is one of the main components in the herb Centella asiatica, which is a well-known herbal medicine for its excellent pharmacological effects. To enhance the development potentials of asiatic acid as a chemopreventative agent, there is a great need to further understand its biopharmaceutical and pharmacokinetic properties. The aim of this research is to clarify the mechanisms of absorption and metabolism of asiatic acid, and explore its biopharmaceutical and pharmacokinetic properties in rats by using a sensitive and robust HPLC-MS method. MATERIALS AND METHODS: Male Sprague-Dawley rats were randomly assigned to 2 groups and administered with asiatic acid by oral and intravenous administration. Plasma concentrations of asiatic acid were determined at designated points and main pharmacokinetic parameters were estimated. The absorption of asiatic acid was investigated by using Caco-2 cell line absorption model in vitro and rat intestinal perfusion model in situ. The metabolic rate of asiatic acid was investigated by incubating it in rat liver microsome system in vitro. In addition, the solubility of asiatic acid in aqueous solution was also determined by using HPLC-MS method. RESULTS: The absolute oral bioavailability of asiatic acid is 16.25%. It was found that the permeability of asiatic acid is more than 10(-5) in the Caco-2 cell monolayer and rat intestinal perfusion model, and its main absorption region is the jejunum in rats. The metabolic rate of asiatic acid in rat liver microsomes, t1/2, is 9.493min, which shows that asiatic acid can be metabolized rapidly. The solubility of aisiatic acid was 0.1583mgmL(-1), and its poor solubility will result in low bioavailability. CONCLUSIONS: The asiatic acid in a variety of matrixes was analyzed by using a sensitive and specific HPLC-MS method, and its absolute oral bioavailability in rats was very low. Asiatic acid can be metabolized rapidly in rat liver microsomes, and has good permeability across Caco-2 monolayer cell and rat intestine perfusion. It can be deduced that the low bioavailability of asiatic acid results from poor solubility and rapid metabolism.

[LC-MS quantification and pharmacokinetics of the multi-constituents of Huangqin Tang in rat plasma after different single oral doses].

The current study aims to investigate the pharmacokinetic properties of Huangqin Tang on different oral doses. An LC-MS method for simultaneous determination of flavonoids and terpenoids in rat plasma was developed and validated. Plasma samples were treated with hydrochloric acid (containing 1% ascorbic acid), precipitated with acetonitrile, separated on a Zorbax SB-C18 column, detected by single quadruple mass spectrometry with an electrospray ionization interface, and quantified using selected ion monitoring mode. All pharmacokinetic parameters were processed by non-compartmental analysis using WinNonlin software. The results of specificity, linearity, intra-day and inter-day precisions, accuracy, and stability for LC-MS assay were suitable for the quantification of paeoniflorin, baicalin, wogonoside, baicalein, wogonin, oroxylin A, glycyrrhizic acid and glycyrrhetinic acid in rat plasma. The concentration-time profiles of baicalin, wogonoside, baicalein, wogonin, oroxylin A and glycyrrhizic acid showed double-peak phenomenon after Huangqin Tang was orally administered at 40 g x kg(-1) dose; all eight constituents in rat plasma showed good dose-exposure relationship within the dosage of 10-40 g x kg(-1); although plasma concentrations were different, the flavonoids with the same backbone showed the similar fate in the body with the corresponding dosage. In conclusion, the LC-MS assay was successfully applied for the pharmacokinetic study of multi-constituents of Huangqin Tang with different doses. Additionally, these constituents demonstrated good pharmacokinetic properties in the body.

Pharmacokinetic comparisons of two different combinations of Shaoyao-Gancao Decoction in rats: competing mechanisms between paeoniflorin and glycyrrhetinic acid.

ETHNOPHARMACOLOGICAL RELEVANCE: Shaoyao-Gancao Decoction (SGD), a well-known traditional Chinese medicine prescription, is a combination of Radix Paeoniae Alba (Paeonia lactiflora Pall, root) and Glycyrrhizae uralensis (Glycyrrhiza uralensis Fisch., root and rhizome, honeyed) for spasmolysis and emergency pain relief. Paeoniflorin (PF) and glycyrrhetinic acid (GA) are two typical active components of SGD for pain relief. AIM OF THE STUDY: To study comparative pharmacokinetics of ten bioactive compounds in SGDs with two different combinations of RP and GU, and therefore to investigate the herb-herb interaction mechanisms of Shaoyao-Gancao Decoction for better spasmolysis and emergency pain relief in rats. MATERIALS AND METHODS: Herbal IR macro-fingerprinting was implemented to provide the full chemical fingerprints of RP, GU and SGD decoctions and to investigate the variation rule of the full chemical profile of SGDs with various combinations of RP and GU. A specifically developed HPLC-MS/MS assay coupled with protein precipitation method was employed to determine the plasma concentrations of the ten analytes. Male Wistar rats were orally administered with SGD1 (RP:GU, 1:1 (w/w)) and SGD2 ((RP:GU, 4:1 (w/w)) equivalent to 9.5 g/kg body weight of GU. RESULTS: Full chemical fingerprints of RP, GU and SGDs with various combinations of RP and GU were provided in the form of IR macro-fingerprints. Except for liquiritin, there were statistically significant differences (p<0.05 or p<0.01) of these analytes between SGD1 and SGD2 in in vivo pharmacokinetic study. Compared with the results when oral administrated with SGD1, six glycosides (PF, albiflorin, oxypaeoniflorin, isoliquiritin, ononin, and glycyrrhizin) exhibited higher systematic exposure levels (AUC0-t) and slower elimination rates (CL) whereas two glycones (GA and isoliquiritigenin) were the reverse when administrated with SGD2. CONCLUSIONS: Increasing the amount of RP attenuated the inhibitory effect of GA via competing being consumed by intestinal bacteria (or ß-glucosidase) to reduce the conversion amount of glycyrrhizin to GA and subsequently to afford significantly higher bioavailability and longer efficacy of PF, glycyrrhizin, albiflorin, oxypaeoniflorin, isoliquiritin, and ononin, leading to better spasmolysis and emergency pain relief.

Analytical strategy to reveal the in vivo process of multi-component herbal medicine: a pharmacokinetic study of licorice using liquid chromatography coupled with triple quadrupole mass spectrometry.

Although various techniques have been employed to analyze drug metabolites, the metabolism of multi-component herbal medicine has seldom been fully addressed. In contrast to chemical drugs, a number of compounds in herbal medicine could get into circulation and then be metabolized. Moreover, these compounds may have metabolic interactions which make their pharmacokinetics (PK) even more complicated. The present work aims to elucidate the multi-component pharmacokinetics of a herbal medicine, and to demonstrate how PK behaviors were altered by co-existing constituents. Licorice (Glycyrrhiza uralensis Fisch.), a most commonly used herbal medicine, was chosen as a model. A strategy was proposed to compare the PK profiles of licorice extract with those of nine single compounds. These compounds were major bioactive constituents of licorice, and represented various structural types (flavanone, chalcone, isoflavone, saponin, and coumarin). We established a segmented selected reaction monitoring LC/MS/MS method to simultaneously monitor 63 licorice metabolites in rat plasma, and obtained the PK profiles of 55 metabolites. The results indicated that interactions among licorice compounds altered their PK behaviors in 4 aspects: improvement in bioavailability for aglycones (133- and 109-fold increase for liquiritigenin and isoliquiritigenin, respectively), prolongation in system circulation for glycosides (0.3h delay in T(max) for liquiritin apioside and isoliquiritin apioside), decrease of potential toxicity for saponins such as glycyrrhizic acid, and shift in plasma distribution for phase II metabolites. This is the first attempt to systematically reveal the in vivo process of licorice. Moreover, the study indicates noticeable interactions to alter pharmacokinetics among licorice compounds, which may be characteristic for herbal medicines.