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Poria cocos is a traditional Chinese medicine with homology of medicine and food in China. It has the effects of promoting diuresis, eliminating dampness, invigorating spleen, calming heart and so on. It is widely used in medicine, food and health care products. With the in-depth study of P. cocos, its triterpenes, polysaccharides and other major chemical components, as well as its wide range of pharmacological effects and application development research have attracted much attention. This paper systematically reviewed the chemical components and pharmacological effects of P. cocos, according to the concept of quality markers, the quality markers of P. cocos were predicted and analyzed from the aspects of the biosynthetic approach and specificity of chemical components, traditional medicinal efficacy, traditional medicinal properties, measurable components, different processing methods and so on, which provides a scientific basis for quality evaluation and product development of P. cocos.
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Objective: To summarize the prescription rules of traditional Chinese medicine (TCM) for treating alcoholic liver disease (ALD) based on TCM inheritance support system (V2.50). Methods: The literatures about TCM prescriptions for treating ALD were collected from CNKI, Wanfang database, and VIP database. The TCM inheritance platform system was used to analyze the prescription rules of TCM in the treatment of alcoholic liver disease. Results: Statistics showed that the majority of prescriptions were used to treat alcoholic fatty liver, alcoholic hepatitis, and alcoholic cirrhosis. Through "frequency statistics" analysis, 107 prescriptions were found involving 149 flavors of TCM, with a cumulative frequency of 1 195 times. Twenty-three Chinese medicines with a frequency of ≥ 15 times were used, and the cumulative frequency was 737 times (62%). The most frequently used medicines were blood-activating and stasis-removing drugs, water-diffusing and damp-permeating drugs, tonics, heat-clearing drugs, antialcoholic poisons and qi-regulating drugs. The commonly used doses of Salvia miltiorrhiza, Poria cocos, Bupleurum chinense, Glycyrrhiza uralensis, Atractylodes macrocephala, Alismatis Rhizoma, and Curcumae Radix in the top 10 medicines ranked in the frequency of medication accorded with the prescribed doses in the Pharmacopoeia of the People's Republic of China (2015 edition), while Crataegi Fructus, Artemisiae Scopariae Herba, and Puerariae Lobatae Radix exceeded the prescribed doses. In the frequency analysis of drug pairs, the combination of S.miltiorrhiza and B. chinense was the most widely used. According to the association rules of drug combination, the correlation between Curcumae Radix and S. miltiorrhiza was the strongest, that was, the probability of S. miltiorrhiza appearing with the emergence of Curcumae Radix was 88%. From the network display chart, it was indicated that S. miltiorrhiza and P. cocos were the main herbs for treatment. Through unsupervised entropy hierarchical clustering algorithm, 14 core combinations for new clustering were extracted, and seven new prescriptions can be obtained by further clustering. Conclusion: The basic principles of TCM treatment of ALD include promoting blood circulation and removing blood stasis, removing dampness, tonifying, detoxifying alcohol, and promoting qi, and with "protecting spleen and stomach function" as its purpose, which accords with the theoretical basis of traditional Chinese medicine in treating alcoholic liver disease. Core combinations and new prescriptions provide references for clinical drug use and new drug research and development, but new prescriptions must be further evaluated with the combination of traditional Chinese medicine theory and clinical practice.
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Objective: To investigate the non-polysaccharide chemical constituents of Poria cocos and their anti-complementary activity. Methods: The anti-complementary bioassay-guided isolation was carried out with the hemolysis test as guide. All isolates were evaluated for their in vitro anti-complementary activities on the classical pathway. The structures were identified by various spectroscopic data including ESI-MS, 1H-NMR, and 13C-NMR data. Results: Eleven compounds were isolated from the EtOAc fraction of P. cocos extracts, including stigmasterol (1), lupeol (2), oleanolic acid (3), ursolic acid (4), polyporenic acid C (5), tumulosic acid (6), dehydrotumulosic acid (7), 3-epi-dehydrotumulosic acid (8), pachymic acid (9), dehydropachymic acid (10), and dehydrotrametenolic acid (11). Compounds 1-4 were obtained from this plant for the first time, and compounds 3-11 showed the anti-complementary activity in different degrees. Conclusion: Triterpenoid acids are the main anti-complementary constituents in the chemical constituents of P. cocos non-polysaccharides (CH50 0.10-0.27 g/L).
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Objective: The quality markers of Poria cocos were identified based on the “in vitro-in vivo” multidimensional chemical group associated network and in vivo pharmacokinetics, and the mass spectrometry method for quantitative detection of quality markers was established to evaluate the quality of P. cocos. Methods: A quantitative analysis method of triterpenoids in P. cocos was established by ultra-high performance liquid chromatography-triple quadruple tandem mass spectrometry (UHPLC-MS/MS) while dioscin was chosen as internal reference substance. The pharmacokinetic curves of active ingredients in vivo were drawn. Seven quality markers of P. cocos including dehydrotumulosic acid, tumulosic acid, and poricoic acid B were identified based on the results of pharmacokinetics. The content of high content components and quality markers in 10 batches of P. cocos were detected and used as variables for cluster analysis of 10 batches of P. cocos. Results: There were differences in the results of clustering analysis with different indexes as variables in evaluating the quality of P. cocos. Conclusion: To a certain extent, the method of quality control of P. cocos with high content components is one-sided. In order to comprehensively and accurately control the quality of P. cocos, it is necessary to take into full account the in vivo and in vitro changes and the in vivo dynamic process of P. cocos composition, and select the effective quality markers related to its pharmacodynamics for its quality control.
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Objective: Poria cocos and Polyporus umbellatus are similar medicinal fungi in traditional Chinese medicines. A method for fingerprint analysis of monosaccharide composition of polysaccharides by HPLC combined with chemometrics methods has been developed for characterization and discrimination of them in this research. Methods: The polysaccharides were extracted by decocting in water, and then completely hydrolyzed with hydrochloride. Monosaccharides in the hydrolyzates were derivatized with 1-phenyl-3-methyl-5-pyrazolone (PMP) for HPLC analysis. More than 20 batches of P. cocos and P. umbellatus from different regions were analyzed. Results: The fingerprints of P. cocos showed five common characteristic peaks, which were identified by comparing with the reference substances. The five peaks corresponded to the derivatives of mannose, ribose, glucose, galactose, and fucose. At the same time, the fingerprints of P. umbellatus showed eight common characteristic peaks, of which seven were identified as the derivatives of mannose, ribose, rhamnose, glucose, galactose, xylose, and fucose. Moreover, the similarity of their fingerprints was respectively calculated by the Similarity Evaluation System for Chromatographic Fingerprint of TCM published by China Pharmacopoeia Committee (Version 2004A). And the data were further processed by hierarchical cluster analysis (HCA) and principal component analysis (PCA). The similarity evaluation and HCA indicated that there were no significant difference in P. cocos or P. umbellatus samples from different geographical regions, but PCA was performed to characterize the difference in monosaccharide constituents between P. cocos and P. umbellatus, and linear discriminant analysis (LDA) showed the overall correct classification rate was 100%. Conclusion: The fingerprint analysis method of monosaccharide composition of water-soluble polysaccharides can distinguish P. cocos and P. umbellatus, and can be applied for the authentication or quality control for P. cocos and P. umbellatus.
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Objective To establish a new method and validate its feasibilities by the simultaneous quantitative assay of four triterpenes in Poria cocos. Methods A new quality evaluation method of multi-components by single marker (QAMS) was established and validated for P. cocos. Pachymic acid (PA), dehydropachymic acid (DPA), dehydrotumulosic acid (DTA), and dehydrotrametenolic acid (DMA) were selected as analytes while pachymic acid was chosen as internal reference substance to evaluate the quality. The relative correction factor (RCF) of pachymic acid to the other three triterpenes were calculated. The method was evaluated by the comparison of quantity between external standard method and QAMS method. Results The contents of four triterpenes in 17 batches of P. cocos from QAMS method were not significantly different from those from external standard method. Conclusion The method with a single marker is accurate and feasible to determine PA, DPA, DTA, and DMA when some authentic standard substance are unavailable.
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Daodi herbs, as the quintessence of Chinese medicine, is famous for its genuine products, good quality and effect. The research of Daodi herbs influences both the process and development of modernization and internationlization of Chinese medicine. Poria cocos has a long history of application, there is a certain difference over the time of ancients to P. cocos, and P. cocos producing area has a certain change with the change of dynasty. Moreover, as a medicine with dual-purpose of drug and food, it has a widely development future. Therefore, this article from the perspective of the herbalogy to research and discuss the history and changes of P. cocos including the original plant awareness, the real estate area, the medicinal, the taste, the harvest processing, and the quality evaluation, in order to provide the theory basis for the further development and utilization of P. cocos.
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Objective: In order to carry out the study on proteins from Poria cocos fermentation broth, the proteins in the fermentation broth were separated and identified. Methods: Proteins were obtained by organic acid precipitation from the fermentation broth and the protein concentration was determined by Bradford method. The obtained P. cocos secreted proteins were separated on SDS-PAGE electrophoresis, subjected to in-gel digestion, then identified by mass spectrometric analysis followed by database searching. Results: The protein concentration in the fermentation broth was around 74.01 μg/mL, with the apparent molecular weight ranged from 2.8 × 104 to 1.3 × 105. A total of 52 P. cocos secreted proteins were identified, including catalase, protein kinase, alkaline protease, glucoamylase, lysozyme, and so on. Conclusion: P. cocos fermentation broth has abundant proteins, which could be a good material for the study of P. cocos protein and also a potential healthy food and beverage.
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Objective: To clone cytochrome P450 reductase (PcCPR) gene from Poria cocos and to characterize with bioinformatics methods. Methods: According to annotated transcriptome of P. cocos, the PcCPR gene was cloned through RACE, and the genomic DNA sequence was further obtained through PCR. The characteristics of the encoded protein were analyzed using bioinformatics, the 3D structure of the protein was modeled with I-TASSER server, and phylogenetic tree of CPR was carried out with MEGA. Results: The 2 514 bp full-length cDNA sequence (GenBank Accession No. KP768251) and the 5 292 bp genomic DNA sequence (GenBank Accession No. KP896487) of PcCPR was obtained, which contained four exons and three introns. PcCPR encoded a protein with 732 amino acids. The protein was predicted to be an unstable hydrophilic protein with calculated molecular weight of 81 147 and isoelectric point 5.39. PcCPR does not have a signal peptide but has a transmembrane segment (aa residues 7 to 22), and it is anchored to endoplasmic reticulum. There are two flavin binding domains and many predicted FAD and NADP binding sites, these sites are adjacent respectively at 3D structure level. The homologous analysis indicates that PcCPR has a higher similarity with CPR from basidiomycetes than CPR from ascomycetes. Conclusion: The PcCPR was successfully cloned, which will provide a foundation for researches on PcCPR and PcCPR associated metabolic.
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Objective: To develop new molecular markers for Poria cocos, and to characterize the SSR in P. cocos transcriptome. Methods: The transcriptome Ungenes and genomic scaffolds were examined by the tool of MISA. The gene annotation and gene function cluster were obtained by blasting the Unigenes which contained SSR to the nr and KEGG databases with BlastX. Results: A total of 2 075 SSRs were identified in 4.57% Unigene sequences, the density of distribution was average one SSR per 17.01 kb, and the average length of SSR was 19.59 bp. Meanwhile, those were 54.00 SSRs per Mb, and 20.74 bp in genomic sequences. Among all 241 SSR motifs found in transcriptome, (CG/CG)n which accounted for 10.97% was the most frequent repeat motif. And hexa-nucleotide repeats which accounted for 35.64% was the most group among mono- to hexa-nucleotide repeats. (ACCACG/CGTGGT)14 with the length of 84 bp was the longest SSR. Only 115 Unigenes of 1 887 Unigenes containing SSR were annotated to cellular metabolic process or nucleotide binding, etc, with GO classification. On the other hand, 1 223 Unigenes containing SSR annotated into 219 KEGG pathway maps. 314 and 297 Unigenes of them were annotated into metabolism pathways and genetic information processing pathways, respectively. Conclusion: The SSR in the transcriptome of P. cocos is rich in type, and has a high potential of polymorhpism. Associating gene function, SSR might be applied in marker-assisted breeding with the aim of specific traits.
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Objective: To prepare the recombinant Poria cocos immunomodulatory protein-1 (WCFIP1) and its antibody. Methods: The cDNA of WCFIP1 was synthesized, cloned into plasmid and transformed into Escherichia coli expression strain. The recombinant protein of WCFIP1 was expressed, purified, and injected into mice to prepare antiserum. The antibody against WCFIP1 was purified by portein-G column and the specificity was determined by Western blotting. Results: The recombinant protein of WCFIP1 and the specific antibody were prepared. Conclusion: The recombinant protein of WCFIP1 is expressed and purified from E. coli, and the specifity of the prepared antibody is determined.
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Objective: To evaluate the activities of Fuling (the dried sclerotium of Poria cocos) and its components, then its nature and flavor were attributed by traditional Chinese medicine (TCM) theory. Methods: Scopolamine was used to cause learning and memory impairment in mice, and Morris water maze test was used to observe the effects of Fuling and its components on learning and memory of mice, and shaking cage method and pentobarbital sodium were used to evaluate the sedative and hypnotic effects of Fuling and its components. Results: Water decoction of Fuling (crude drug 8.56 g/kg), ethyl acetate, petroleum ether, crude polysaccharides, and refined polysaccharides components could significantly shorten the latency of scopolamine-induced memory impairment in mice to reach the platform; Acetylcholinesterase activity in mice brain was decreased, the effects of the components washed with alcohol and water on latency were less obvious and the individual components of the cerebral index showed an increase in different proportions; Crude drug (42.8 g/kg) water decoction of Fuling could significantly prolong the sleeping time, but had no effect on sleep latency; Crude polysaccharides could significantly shorten the sleep latency while the refined polysaccharides could significantly increase sleeping time; Followed by ethyl acetate components, the petroleum ether components showed a weak antagonistic effect. Conclusion: The learning and memory improvement as well as the sedative and hypnotic effects are associated with Fuling's sweet taste which can tonify spleen to calm nerves; The ethyl acetate, petroleum ether, and polysaccharides components are the basic materials of sweet taste.
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Objective: To definite the technological parameter of carboxymethylation modified product of Poria cocos, improve the water-soluble ability of Poria cocos and to expand the processing applications of P. cocos in Chinese medicine industry. Methods: Taking the P. cocos powder as raw materials, The single-factor experiment of carboxymethylated modification was carried out to gain the greatest degree of substitution. Results: Reaction medium, 85% ethanol solution reaction temperature, 50°C, 6 h of etherification after 2 h of and molar ratio of alkali/acid controlled in 2.5 : 1-3 : 1, and the water-soluble ability was improved. Conclusion: Using ethanol as reaction medium and method of two steps of alkaline addition to obtain carboxymethylation modified product of P. cocos, the processing characteristics of modified product could be greatly improved and be used in processing applications.
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Objective To study the conditions of separation and regeneration of Poria cocos protoplast.Methods Separation and regeneration of P.cocos protoplast under different ages of hyphae,enzyme consistency,and enzymolysic time and stabillizer.Results The results showed that the yield of the protoplast was different under different ages of hyphae,enzyme concentration,enzymolysic time,and stabilizer.Conclusion For strain Z_(10-2),its optimal age of hyphae,enzyme concentration,and stabilizer is 72 h,3%,and 0.5 mol/L mannitol,respectively.The strain Z_(10-3) is 56 h,3%,and 0.6 mol/L mannitol.The frequency of regeneration of the two strains is the best and 7.5?10~(-3)(strain Z_(10-2)) and 1.3?10~(-2)(strain Z_(10-2)) separately when the stabilizer is 0.6 mol/L mannitol.