Pigmentation Effects of Kaliziri Standard Extracts on Melanocytes, C57BL/6 mice of Hydroquinone- Induced and Guinea Pigs of Hydrogen Peroxide-Induced Hyperpigmentation Model

Background: Vitiligo is a depigmentation disorder characterized by losing functional melanocytes, leading to skin and/or hair depigmentation. Kaliziri (Vernonia anthelmintica (L.) Willd) seeds have been traditionally used to treat pigmentation disorders in Central Asia. The extracts of these seeds increased melanin content and tyrosinase activity and upregulated melanogenesis-related proteins in B16 melanoma cells. We isolated the main active compound Kaliziri standard Extracts (encoded as CAM-Y7) from the Kaliziri seeds in a previous study. Our goal was to reveal whether CAM-Y7 promotes melanogenesis in melanocytes and restored pigmentation in animal models of vitiligo. Methods: Herein, we analyzed the melanogenic effects of CAM-Y7 on melanocytes and the skin pigmentation of hydroquinone (H.Q.)-induced mouse model and hydrogen peroxide (H2O2)-induced guinea pig model of vitiligo. Results: CAM-Y7 promotes Melanogenesis up-Regulation MITF-induced Tyrosinase Expression via p38MAPK signal pathway in melanocytes. Oral administration of CAM-Y7 progressively darkened the dorsal skin and hair of the C57BL/6 mice and guinea pigs. Lillie staining and hematoxylin-eosin staining further showed that CAM-Y7 induced melanogenesis in the epidermis and hair follicles of the animals. Conclusion: Taken together, these results suggest that CAM-Y7 regulation of melanogenesis may be mediated by the activation of p38 MAPK and restored pigmentation in an HQ-Induced C57BL/6 mouse and H2O2-induced guinea pig models of vitiligo. CAM-Y7 might be valuable and promising therapeutic as an agent for vitiligo. Subsequent clinical experiments using are needed to verify our results.

The Great Capacity on Promoting Melanogenesis of Three Compatible Components in Vernonia anthelmintica (L.) Willd.

To investigate a possible methodology of exploiting herbal medicine and design polytherapy for the treatment of skin depigmentation disorder, we have made use of Vernonia anthelmintica (L.) Willd., a traditional Chinese herbal medicine that has been proven to be effective in treating vitiligo. Here, we report that the extract of Vernonia anthelmintica (L.) Willd. effectively enhances melanogenesis responses in B16F10. In its compound library, we found three ingredients (butin, caffeic acid and luteolin) also have the activity of promoting melanogenesis in vivo and in vitro. They can reduce the accumulation of ROS induced by hydrogen peroxide and inflammatory response induced by sublethal concentrations of copper sulfate in wild type and green fluorescent protein (GFP)-labeled leukocytes zebrafish larvae. The overall objective of the present study aims to identify which compatibility proportions of the medicines may be more effective in promoting pigmentation. We utilized the D-optimal response surface methodology to optimize the ratio among three molecules. Combining three indicators of promoting melanogenesis, anti-inflammatory and antioxidant capacities, we get the best effect of butin, caffeic acid and luteolin at the ratio (butin:caffeic acid:luteolin = 7.38:28.30:64.32) on zebrafish. Moreover, the effect of melanin content recovery in the best combination is stronger than that of the monomer, which suggests that the three compounds have a synergistic effect on inducing melanogenesis. After simply verifying the result, we performed in situ hybridization on whole-mount zebrafish embryos to further explore the effects of multi-drugs combination on the proliferation and differentiation of melanocytes and the expression of genes (tyr, mitfa, dct, kit) related to melanin synthesis. In conclusion, the above three compatible compounds can significantly enhance melanogenesis and improve depigmentation in vivo.

Quantitative Determination of Marker Compounds and Fingerprint Analysis of the Seeds of Vernonia anthelmintica

In traditional Chinese medicine, the seeds of Vernonia anthelmintica (L.) Willd. have been widely used for treatment of cough, skin diseases, diarrhea, fever, schistosomiasis, amoebic dysentery, and gastrointestinal problems, especially in the treatment of vitiligo for thousands of years in China. In this study, an effective, reliable, and accurate high-performance liquid chromatography diode array detector (HPLC-DAD) method was developed for quantitative analysis of 3 marker bioactive compounds and chemical fingerprint of the seeds of V. anthelmintica. Data corresponding to common peak areas and HPLC chromatographic fingerprints were analyzed by exploratory hierarchical cluster analysis (HCA) and principal component analysis (PCA) to extract information of the most significant variables contributing to characterization and classification of the analyzed samples. Based on variety and origin, the high-performance thin layer chromatography (HPTLC) method validated the chemical fingerprint results used to screen the in vitro antioxidant activity of V. anthelmintica. The results show that the developed method has potential application values for the quality consistency evaluation and identification of similar instant V. anthelmintica samples. When considered collectively, this research results provide a scientific basis for the improvement of standardization and specification of V. anthelmintica medicinal materials and provide a pathway for the development and utilization of references for the identification of V. anthelmintica herbs.

A preliminary investigation of anticholinesterase and antioxidant properties of various extracts of Vernonia anthelmintica in relation to the treatment of Alzheimer’s disease

The complexity of the underlying pathogenesis of Alzheimer’s disease (AD) and substandard results of existing treatment, demands persistent research for the development of new therapeutics. As natural antioxidant has attracted considerable attention on this regards, the present study evaluated and validated antioxidant and anti-cholinesterase activity of Vernonia anthelmintica seeds. Different extracts were collected after sequential extraction of plant seeds by using pet ether, chloroform, ethyl acetate, methanol and water as solvents. Antioxidant activity of the extracts was tested by using superoxide, nitric oxide (NO) and 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assays, whereas, acetylcholine esterase (AChE) inhibition property was assessed by Ellman’s method. Results revealed that all the extracts produced some degree of both the activities in a concentration-dependent manner. While pet ether extract exhibited weakest, methanolic extract displayed potent radical scavenging and anti-cholinesterase properties in all the assays with IC50 of 159.71 μg/ml against AChE inhibition and 98.51, 120.22 and 170.79 μg/ml against superoxide, NO and DPPH radical scavenging assays respectively. Presence of an array of secondary metabolites with modest flavonoid and phenol content in the methanolic extract is accountable for these desired activities. Collectively, reports from our experiments covey that V. anthelmintica possess significant AChE inhibitory and antioxidant property and thus can be further evaluated in search of potential disease-modifying therapeutic for management of neurodegenerative diseases like AD.