Qualitative and quantitative analysis of Arnebiae Radix and Dictamni Cortex and efficacy study of herbal extracts on allergic contact dermatitis using 3D human reconstructed epidermis.

Objective: To evaluate the quality of Arnebiae Radix (AR) and Dictamni Cortex (DC) and study the efficacy of herbal extracts of these two herbs on the treatment of allergic contact dermatitis (ACD). Methods: Qualitative and quantitative analysis of effective components was performed using High Performance Thin Layer Chromatography (HPTLC), High Performance Liquid Chromatography (HPLC), and HPLC-Quadrupole Time of Flight-Mass Spectrometry (HPLC-QTOF-MS). In vitro allergic ACD 3D model was established by incubating 3D reconstructed human epidermis (RHE) with skin sensitizer, potassium dichromate. A total of 65 gene expression that were associated with ACD, which included 24 antioxidant responsive element (ARE) and 41 SENS-IS genes were quantified by qRT-PCR. More than or equal to 10 ARE genes and 18 SENN-IS genes were induced by 1.3-fold, demonstrating the successful establishment of in vitro ACD model. Oil extracts of AR and DC were applied on the in vitro ACD model to study the efficacy. Results: Batch 3 of AR and batch 2 of DC showed presence of all active ingredients with the highest concentrations. Active ingredients of the herbs were extracted using a special oil and formulated into herbal oil extracts. The herbal oil extracts were able to down regulate the induced genes in the in-vitro ACD skin model, bringing the tissue back to homeostatic status. Conclusion: The oil extracts showed the potent efficacy of using AR and DC in ACD treatment. The combination study will be done to optimize the formulation ratio which will be developed into a topical cream.

Hepatic Bioactivation of Skin-Sensitizing Drugs to Immunogenic Reactive Metabolites.

The clinical use of some drugs, such as carbamazepine, phenytoin, and allopurinol, is often associated with adverse cutaneous reactions. The bioactivation of drugs into immunologically reactive metabolites by the liver is postulated to be the first step in initiating a downstream cascade of pathological immune responses. Current mechanistic understanding and the ability to predict such adverse drug cutaneous responses have been partly limited by the lack of appropriate cutaneous drug bioactivation experimental models. Although in vitro human liver models have been extensively investigated for predicting hepatotoxicity and drug-drug interactions, their ability to model the generation of antigenic reactive drug metabolites that are capable of eliciting immunological reactions is not well understood. Here, we employed a human progenitor cell (HepaRG)-derived hepatocyte model and established highly sensitive liquid chromatography-mass spectrometry analytical assays to generate and quantify different reactive metabolite species of three paradigm skin sensitizers, namely, carbamazepine, phenytoin, and allopurinol. We found that the generation of reactive drug metabolites by the HepaRG-hepatocytes was sensitive to the medium composition. In addition, a functional assay based on the activation of U937 myeloid cells into the antigen-presenting cell (APC) phenotype was established to evaluate the immunogenicity potential of the reactive drug metabolites produced by HepaRG-derived hepatocytes. We showed that the reactive drug metabolites of known skin sensitizers could significantly upregulate IL8, IL1ß, and CD86 expressions in U937 cells compared to the metabolites from a nonskin sensitizer (i.e., acetaminophen). Thus, the extent of APC activation by HepaRG-hepatocytes conditioned medium containing reactive drug metabolites can potentially be used to predict their skin sensitization potential.

The relative antioxidant activity and steric structure of green tea catechins - A kinetic approach.

There are eight most abundant green tea catechins (GTCs) existing in four pairs of eipimers, and carbon-3 configuration represents the only steric difference within each pair. This study aimed to use a new kinetic approach to elucidate the effect of stereochemical changes on the antioxidant activity. A mixture of eight GTCs was treated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) prepared in a series of concentrations, their relative reaction rates towards scavenging DPPH were revealed by the recently introduced parameter, i.e. Dm. The 3-R configuration in (-)-epicatechin, (-)-epigallocatechin and (-)-epigallocatechin gallate gave lower Dm values demonstrating faster kinetics as opposed to their 3-S counterparts, with the only exception of the pair of (-)-catechin gallate and (-)-epicatechin gallate in which 3-S configuration was faster. These results suggested that the kinetic approach adopted in this study could reflect the different antioxidant activity of GTCs attributed by minor steric changes.

Bioavailability of Orally Administered Des-Aspartate-Angiotensin I in Human Subjects.

In an earlier single-dose escalation study to evaluate the safety and pharmacokinetics of orally administered des-aspartate-angiotensin I (DAA-I) in healthy subjects, the plasma level of DAA-I could not be determined because DAA-I is rapidly degraded in the circulation. The present study investigated the oral bioavailability of DAA-I by measuring the prostaglandin E2 metabolite (PGEM) in the plasma samples of the same trial. PGEM is a stable derivative of PGE2, which has been shown to be a biomarker of DAA-I. The data show that plasma from two of the three subjects who were orally administered the efficacious preclinical dose of 0.70 mg/kg DAA-I exhibited a significant PGEM peak at 5-6 h postdose. Plasma of subjects who were administered 0.08 and 1.5 mg/kg DAA-I, the subefficacious and two-times efficacious dose, respectively, did not exhibit a similar PGEM peak. This observation is concordant with the known in vivo actions of DAA-I, especially its hypoglycemic action where maximum efficacy occurred at a dose of 0.7 mg/kg, and decreased to nil at the two-times efficacious dose. The onset of the PGEM peak at 5-6 h postdose was closed to the 4-h onset of absorption of [C14]DAA-I seen in preclinical rat studies, albeit the absorption kinetics between rodents and humans are not identical. The occurrence of polymorphism of enzymes involved in the formation and degradation of PGE2 is common, and this has been attributed to contributing to the variation in response, onset and peak PGEM observed among the three subjects who were administered the efficacious dose.

A new parameter to simultaneously assess antioxidant activity for multiple phenolic compounds present in food products.

In this work, we established a new methodology to simultaneously assess the relative reaction rates of multiple antioxidant compounds in one experimental set-up. This new methodology hypothesizes that the competition among antioxidant compounds towards limiting amount of free radical (in this article, DPPH) would reflect their relative reaction rates. In contrast with the conventional detection of DPPH decrease at 515nm on a spectrophotometer, depletion of antioxidant compounds treated by a series of DPPH concentrations was monitored instead using liquid chromatography coupled with quadrupole time-of-flight (LC-QTOF). A new parameter, namely relative antioxidant activity (RAA), has been proposed to rank these antioxidants according to their reaction rate constants. We have investigated the applicability of RAA using pre-mixed standard phenolic compounds, and also extended this application to two food products, i.e. red wine and green tea. It has been found that RAA correlates well with the reported k values. This new parameter, RAA, provides a new perspective in evaluating antioxidant compounds present in food and herbal matrices. It not only realistically reflects the antioxidant activity of compounds when co-existing with competitive constituents; and it could also quicken up the discovery process in the search for potent yet rare antioxidants from many herbs of food/medicinal origins.