Subchronic toxicity and genotoxicity studies of Hericium erinaceus ß-glucan extract preparation.

The medicinal effects of Hericium erinaceus have been long documented in scientific studies of Eastern traditional medicine. It is widely consumed, because of its nutritional qualities and perceived health benefits. Also, it is rich in ß-glucans, which has been shown to have immunomodulating and antitumor effects. The objective of the present study was to investigate adverse effects, if any, of ß-glucan extract preparation from H. erinaceus in subchronic toxicity and genotoxicity studies. The conduct of these studies was in compliance with Good Laboratory Practice (GLP) and test guidelines established by the Organization for Economic Cooperation and Development (OECD). In the subchronic toxicity study, Sprague Dawley rats (12/sex/group) were administered (gavage) H. erinaceus ß-glucan extract preparation at dose levels of 0, 500, 1000 and 2000 mg/kg body weight (bw)/day for 90 days. Treatment with H. erinaceus ß-glucan extract preparation did not result in any toxicologically significant treatment-related changes in clinical observations, ophthalmic examinations, body weights, body weight gains, feed consumption, and organ weights. Clinical pathology including hematology, serum chemistry, urinalysisand terminal necropsy (gross or histopathology findings) did not reveal any treatment-related adverse effects. The results of genotoxicity studies as evaluated by gene mutations in Salmonella typhimurium, in vitro chromosome aberrations and in vivo micronucleus test in mice did not reveal any genotoxicity of H. erinaceus ß-glucan extract preparation. Based on the subchronic study, the no observed-adverse-effect level (NOAEL) for H. erinaceus ß-glucan extract preparation was determined as 2000 mg/kg bw/day, the highest dose tested.

[Preliminary study of the changes in blood system in pyrrolizidine alkaloid-related liver damage].

Objective: To preliminary explore the changes in blood system in pyrrolizidine alkaloids (PAs)-related liver damage. Methods: General situation, liver function, biochemical blood test, routine blood test, coagulation function markers, etc., of 77 cases with drug-induced liver damage admitted to the Zhongshan Hospital Affiliated to Fudan University from 2012 to 2019 were retrospectively analyzed. Patients' were divided into PA group, other traditional Chinese medicine group and Western medicine group according to their medication history. Simultaneously, the changes in liver function were observed in the established mice model of monocrotaline-induced liver damage. Liver tissues HE staining and blood routine indexes were observed. Results: 24 cases received PA, 24 cases received other traditional Chinese medicine, and 29 cases received western medicine. Alanine aminotransferase was lower in PA group than the other two groups (P < 0.05), and the total bilirubin and direct bilirubin were significantly lower than the other traditional Chinese medicine group (P < 0.05). The peripheral platelet count of the PA group was (84.11 ± 26.91) ×10(9)/L, which was significantly lower than the lower limit of normal, and had statistically significant difference with other traditional Chinese medicine and western medicine group (P < 0.01). Thrombocytocrit, mean platelet volume and platelet indices of PA group were statistically different from the other two groups (P < 0.05). The D-dimer level in patients with PA group was (2.62 ± 1.93) mg/L, which was higher than the upper limit of normal, and significantly higher than the D-dimer level of the other two groups of patients (P < 0.01). Meanwhile, prothrombin time was longer in PA group than that of the other two groups (P < 0.01), and platelets count were decreased significantly in the mouse model of monocrotaline-induced liver damage after alanine aminotransferase and aspartate aminotransferase elevation (P < 0.01). Conclusion: PA-related liver damage has lower peripheral platelet counts, and the peripheral platelet counts of these patients are lower than other types of drug-induced liver damage. In addition, increased D-dimer in patients with PA-related liver damage indicate a potential risk of thrombosis.

Dimeric Proanthocyanidins on the Stability of Dentin and Adhesive Biointerfaces.

A dentin biomodification strategy with selective proanthocyanidin (PAC)-enriched extracts reinforces dentin and dentin-resin interfaces. Enrichment of the extracts according to the degree of polymerization allows exploration of bioactive principles of PACs and structure-activity relationships. This study investigated the sustained dentin matrix biomodification and dentin-resin bioadhesion of 2 fractions consisting exclusively of B-type PAC dimers with or without a single galloyl motif (specifically, DIMERG and DIMERNG) and their precursor material, enriched grape seed extract (e-GSE; Vitis vinifera). The biomodification potential was determined by long-term evaluation of the apparent modulus of elasticity and collagen solubility (hydroxyproline release). Chemical characterization of the dentin matrix was performed by attenuated total reflectance-Fourier-transform infrared spectroscopy. The bioadhesive properties were assessed by a microtensile bond strength test at different time points, and macro-hybrid layers were produced to verify the degree of conversion of the adhesive resin. Fractions consisting of DIMERG, DIMERNG, and their precursor, e-GSE, increased the modulus of elasticity at all time points and reduced collagen degradation. Specimens treated with DIMERNG remained stable throughout 12 mo of storage, whereas a significant drop in the modulus of elasticity was observed for the DIMERG and e-GSE groups at 6 mo. The fractions and precursor did not affect the degree of resin conversion at the hybrid layer. Changes in infrared resonances corresponding to collagen cross-links in the dentin matrix occurred for all treatments. Higher bond strength was observed for dentin treated with e-GSE as compared with DIMERG and DIMERNG; all biointerfaces remained stable after 12 mo. Nongalloylated PACs mediate stable dentin biomodification, which includes protective activity against collagen degradation and reinforcement of the anchoring dentin matrix. Collectively, PACs with a higher degree of oligomerization offer a robust bioadhesion between the hydrophilic dentin matrix and the hydrophobic adhesive.

Effect of Bioactive Primers on Bacterial-Induced Secondary Caries at the Tooth-Resin Interface.

Secondary caries at the tooth-resin interface is the primary reason for replacement of resin composite restorations. The tooth-resin interface is formed by the interlocking of resin material with hydroxyapatite crystals in enamel and collagen mesh structure in dentin. Efforts to strengthen the tooth-resin interface have identified chemical agents with dentin collagen cross-linking potential and antimicrobial activities. The purpose of the present study was to assess protective effects of bioactive primer against secondary caries development around enamel and dentin margins of class V restorations, using an in vitro bacterial caries model. Class V composite restorations were prepared on 60 bovine teeth (n=15) with pretreatment of the cavity walls with control buffer solution, an enriched fraction of grape seed extract (e-GSE), 1-ethyl-3-(3-dimethyl aminopropyl)-carbodiimide/N-hydroxysuccinimide, or chlorhexidine digluconate. After incubating specimens in a bacterial model with Streptococcus mutans for four days, dentin and enamel were assessed by fluorescence microscopy. Results revealed that only the naturally occurring product, e-GSE, significantly inhibited the development of secondary caries immediately adjacent to the dentin-resin interface, as indicated by the caries inhibition zone. No inhibitory effects were observed in enamel margins. The results suggest that the incorporation of e-GSE into components of the adhesive system may inhibit secondary caries and potentially contribute to the protection of highly vulnerable dentin-resin margins.

Dentin biomodification potential depends on polyphenol source.

Although proanthocyanidins (PACs) modify dentin, the effectiveness of different PAC sources and the correlation with their specific chemical composition are still unknown. This study describes the chemical profiling of natural PAC-rich extracts from 7 plants using ultra high pressure/performance liquid chromatography (UHPLC) to determine the overall composition of these extracts and, in parallel, comprehensively evaluate their effect on dentin properties. The total polyphenol content of the extracts was determined (as gallic acid equivalents) using Folin-Ciocalteau assays. Dentin biomodification was assessed by the modulus of elasticity, mass change, and resistance to enzymatic biodegradation. Extracts with a high polyphenol and PAC content from Vitis vinifera, Theobroma cacao, Camellia sinensis, and Pinus massoniana induced a significant increase in modulus of elasticity and mass. The UHPLC analysis showed the presence of multiple types of polyphenols, ranging from simple phenolic acids to oligomeric PACs and highly condensed tannins. Protective effect against enzymatic degradation was observed for all experimental groups; however, statistically significant differences were observed between plant extracts. The findings provide clear evidence that the dentin bioactivities of PACs are source dependent, resulting from a combination of concentration and specific chemical constitution of the complex PAC mixtures.

Biological characterization of non-steroidal progestins from botanicals used for women's health.

Progesterone plays a central role in women's reproductive health. Synthetic progestins, such as medroxyprogesterone acetate (MPA) are often used in hormone replacement therapy (HRT), oral contraceptives, and for the treatment of endometriosis and infertility. Although MPA is clinically effective, it also promiscuously binds to androgen and glucocorticoid receptors (AR/GR) leading to many undesirable side effects including cardiovascular diseases and breast cancers. Therefore, identifying alternative progestins is clinically significant. The purpose of this study was to biologically characterize non-steroidal progestins from botanicals by investigating theirinteraction and activation of progesterone receptor (PR). Eight botanicals commonly used to alleviate menopausal symptoms were investigated to determine if they contain progestins using a progesterone responsive element (PRE) luciferase reporter assay and a PR polarization competitive binding assay. Red clover extract stimulated PRE-luciferase and bound to PR. A library of purified compounds previously isolated from red clover was screened using the luciferase reporter assay. Kaempferol identified in red clover and a structurally similar flavonoid, apigenin, bound to PR and induced progestegenic activity and P4 regulated genes in breast epithelial cells and human endometrial stromal cells (HESC). Kaempferol and apigenin demonstrated higher progestegenic potency in the HESC compared to breast epithelial cells. Furthermore, phytoprogestins were able to activate P4 signaling in breast epithelial cells without downregulating PR expression. These data suggest that botanical extracts used for women's health may contain compounds capable of activating progesterone receptor signaling.

Pharmacokinetics of 23-epi-26-deoxyactein in women after oral administration of a standardized extract of black cohosh.

Dietary supplements containing black cohosh are alternatives to conventional hormone replacement therapy in menopause. This study investigates the maximum tolerated dose of a 75% ethanol extract of black cohosh and determines the pharmacokinetics of one of its most abundant triterpene glycosides, 23-epi-26-deoxyactein. Single doses of black cohosh extract containing 1.4, 2.8, or 5.6 mg of 23-epi-26-deoxyactein were administered to 15 healthy, menopausal women. Serial blood samples and 24-h urine samples were obtained; blood chemistry, hormonal levels, and 23-epi-26-deoxyactein levels were determined. No acute toxicity or estrogenic hormone effects were observed. Pharmacokinetic analyses of 23-epi-26-deoxyactein in sera indicated that the maximum concentration and area under the curve increased proportionately with dosage, and that the half-life was ~2 h for all dosages. Less than 0.01% of the 23-epi-26-deoxyactein was recovered in urine 24 h after administration. No phase I or phase II metabolites were observed either in clinical specimens or in vitro.

Aqueous extract of Schizandra chinensis fruit causes endothelium-dependent and -independent relaxation of isolated rat thoracic aorta.

An aqueous extract of Schizandra chinensis fruit (ScEx) has long been used to promote the vascular health of postmenopausal women in Korea. This study investigated the ability of ScEx to relax rat aorta constricted with norepinephrine (NE) and the mechanism(s) of such relaxation. ScEx induced partial, endothelium-dependent relaxation. In particular, the relaxation induced by lower concentrations of ScEx (0.1 and 0.3 mg/ml) was largely endothelium-dependent, and was essentially abolished by NG-nitro-L-arginine, methylene blue, 1H-[1,2,3] oxadiazole [4,4-a] quinoxalin-1-one, indomethacin, or ICI 182,780. The results indicate that the response to ScEx involves enhancement of the nitric oxide (NO)-cGMP system, and that it occurs via estrogen receptors. The magnitude of the inhibition with these treatments decreased with increasing ScEx concentration, however, indicating that other vasorelaxation mechanisms are involved, which depend on the ScEx concentration. Calcium concentration-dependent contraction curves in high potassium depolarization medium were shifted significantly to the right and downward after incubation with ScEx (0.3 and 1.0 mg/ml), implying that ScEx is also involved in inhibition of the extracellular calcium influx to vascular smooth muscle. These data demonstrate that ScEx caused both endothelium-dependent and -independent vasorelaxation, which may contribute to understanding the cardiovascular protective effect of ScEx.

Activation of the mu-opiate receptor by Vitex agnus-castus methanol extracts: implication for its use in PMS.

The dried ripe fruit of Vitex agnus-castus L. (VAC) is widely used for the treatment of premenstrual syndrome (PMS). A previous study reported that extracts of VAC showed affinity to opiate receptors; however, functional activity was not determined. We tested two different VAC extracts in receptor binding and functional assays. Our objectives were: (1) to confirm the opiate affinity; (2) to rule out interference by free fatty acids (FFA); (3) to determine the mode of action of VAC at the mu-opiate receptor. Methanol extracts of VAC were prepared either before (VAC-M1) or after (VAC-M2) extraction with petroleum ether to remove fatty acids. Both extracts showed significant affinities to the mu-opiate receptor, as indicated by the concentration-dependent displacement of [3H]DAMGO binding in Chinese hamster ovary (CHO)-human mu-opiate receptor (hMOR) cells. The IC50 values were estimated to be 159.8 microg/ml (VAC-M1) and 69.5 microg/ml (VAC-M2). Since the defatted extract not only retained, but exhibited a higher affinity (p<0.001), it argued against significant interference by fatty acids. In an assay to determine receptor activation, VAC-M1 and VAC-M2 stimulated [35S]GTPgammaS binding by 41 and 61% (p<0.001), respectively. These results suggested for the first time that VAC acted as an agonist at the mu-opiate receptor, supporting its beneficial action in PMS.

Three diterpene glucosides and a diphenylamine derivative from Pieris formosa.

Three new diterpenoids, pierisformosides G-I (1-3) and a diphenylamine derivative 4 have been isolated from the ethanol extract of Pieris formosa leaves, compound 1 being a new type seco-ring-A kaurane diterpene glucoside. Their structures were elucidated with the aid of NMR spectroscopy including two-dimensional-NMR techniques.

ent-Kaurene diterpenoids from Isodon rubescens.

Two kaurenoids, taibairubescensins A and B, were isolated from the ethanol extract of the leaves and branches of Isodon rubescens. Their structures are designated as 2 beta, 3 beta-diacetoxy-11 beta, 13 alpha-dihydroxy-ent-kaur-16-en-15-one and 3 beta, 11 beta-diacetoxy-2 beta, 6 alpha-dihydroxy-ent-kaur-16-en-15-one, respectively, on the basis of detailed spectroscopic analyses.

Diterpenoids from Isodon leucophyllus.

The structures of two new ENT-kaurane diterpenoids and two derivatives of shikokianin isolated from leaves of Isodon leucophyllus were elucidated by 1D and 2D NMR techniques as 11alpha-acetoxyeffusanin D, 6-acetylepinodosinol, 16beta-ethoxymethyleneshikokianin, and 16alpha-ethoxymethyleneshi-kokianin.

Diterpenoid Alkaloids from Delphinium caeruleum.

A new minor diterpenoid alkaloid, caeruline, was isolated from DELPHINIUM CAERULEUM Jacq. ex Camb. Its structure was elucidated as 1 on the basis of spectral evidence. Along with this new compound, three known diterpenoid alkaloids were also isolated from this plant and identified as tatsiensine, delcosine, and lycoctonine which had not been found in this herb previously.