Pilose antler (Cervus elaphus Linnaeus) polysaccharide and polypeptide extract inhibits bone resorption in high turnover type osteoporosis by stimulating the MAKP and MMP-9 signaling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Pilose antler is a traditional Chinese medicine used to improve kidney function, strengthen tendons and bones, and prolong life, among other uses. It is widely employed in the treatment of osteoporosis. However, the molecular mechanisms underlying the treatment of high turnover osteoporosis are not fully understood. AIM OF THE STUDY: The present study aimed to investigate the molecular mechanism underlying pilose antler polysaccharide and polypeptide extracts in inhibiting bone resorption in high turnover osteoporosis, and compare the effects of the two components alone and in combination to explore whether they could produce synergistic enhancement effects. MATERIALS AND METHODS: The quantitative and qualitative characteristics of pilose antler polysaccharide and polypeptide extracts were detected by UV-visible spectrophotometry and high-performance liquid chromatography. A rat model of retinoic acid-induced osteoporosis was used to evaluate the inhibitory effect of the extracts on bone resorption. Enzyme-linked immunosorbent assay (ELISA) was used to detect the activity of factors related to high turnover type osteoporosis in rat serum. Western blotting was used to detect the expression of proteins related to the MAKP and MMP-9 signaling pathways in rat femurs. Fluorescence quantitative PCR was used to detect the transcription levels of genes related to the MAKP and MMP-9 signaling pathways in rat femur tissues. Hematoxylin and eosin staining were used to observe the osteoprotective effects of pilose antler polysaccharides and polypeptides. RESULTS: The yield of pilose antler polysaccharides was 8.3%, and was mainly composed of mannose, glucosamine hydrochloride, glucuronic acid, Galacturonic acid, Galactose hydrochloride, glucose, and galactose. The yield of the polypeptides was 26.2%, and eighty percent of the molecular weight of the antler polypeptides was 1.6 kDa-7kD, among which, the molecular weight of 7kD peptide accounted for 52% of the total. Both polysaccharides and peptides could reduce the activities of TRACP, OCN, ERK1, JNK, and MMP-9 in rat serum and reduce both the protein expression and gene transcription levels of ERK1, JNK, and MMP-9 in rat femur tissue with significant differences compared with the model group. Both extracts exerted significant protective effects on rat femur tissue. The effect of pilose antler polypeptides alone was better than that of polysaccharides either alone or in combination. CONCLUSIONS: Pilose antler polysaccharides, polypeptides, and their mixtures could inhibit the occurrence of bone resorption of high turnover osteoporosis by stimulating the MAKP and MMP-9 signaling pathways to reduce the expression of the ERK1, JNK, and MMP-9 genes and proteins, and could help alleviate bone loss caused by retinoic acid. Pilose antler polypeptides had a stronger effect on inhibiting bone resorption. The combination of the two components did not show synergistic enhancement effect, and the polysaccharide tended to moderate the inhibitory enhancement effect of the polypeptide.

[Detection of chondroitin sulfate in Cervi Cornu Pantotrichum and Cervi Cornu of different specifications and its application in quality evaluation].

The present study comprehensively compared the content of chondroitin sulfate in Cervi Cornu Pantotrichum(CCP) and Cervi Cornu(CC) of different specifications and explored the feasibility of chondroitin sulfate as an indicator to distinguish between CCP and CC. Twenty-two batches of CCP of different specifications(two-branched velvet antler and three-branched velvet antler) from 15 habitats, CC from 6 habitats, and 60 batches of CCP slices prepared from different parts(wax slices, powder slices, gauze slices, and bone slices) were collected. High-performance liquid chromatography(HPLC) was used to determine chondroitin sulfate content in CCP and CC of different specifications. Cluster analysis was used to classify CCP slices of different specifications. The results showed that CCP contained abundant chondroitin sulfate. The average content of chondroitin sulfate was 2.35 mg·g~(-1) in two-branched velvet antler and 1.79 mg·g~(-1) in three-branched velvet antler, significantly higher than 0.11 mg·g~(-1) in CC. Chondroitin sulfate content in wax slices, powder slices, gauze slices, and bone slices were 7.81, 8.39, 1.33, and 0.54 mg·g~(-1), respectively. Cluster analysis showed that gauze slices and bone slices could be clustered into one category and distinguished from wax slices and powder slices. CCP slices prepared from different parts could be separated well through chondroitin sulfate content. Based on the five principles of Q-marker selection, chondroitin sulfate can be used as a potential Q-marker for the identification of CCP and CC, as well as a potential quality indicator for CCP slices of different specifications(wax slices, powder slices, gauze slices, and bone slices). This research provides data support for CCP quality evaluation.

[Quality assessment of different forms of Cervi Cornu Pantotrichum based on content of free amino acids].

In this study, we analyzed the composition and content of 25 free amino acids in 32 batches of different forms of Cervi Cornu Pantotrichum(CCP; one-branched, two-branched, and three-branched) from 15 producing areas. The clustering analysis and orthogonal partial least squares discriminant analysis(OPLS-DA) were performed based on the content of 25 free amino acids. Potential differential metabolites were identified based on VIP value. The results showed that there were 25 free amino acids in CCP, and the average content of essential, non-essential, and total amino acids was 6.13, 32.99, and 39.12 mg·g~(-1), respectively. The clustering analysis and OPLS-DA demonstrated that 25 free amino acids had different content among the three forms of CCP, of which two-branched CCP samples were separately gathered into a group. Five differential components, including glutamic acid, tryptophan, ornithine, γ-aminobutyric acid, and hydroxylysine, were screened out as potential quality markers for the identification of different forms of CCP. This study provides a theoretical basis for the quality evaluation, processing, and utilization of different forms of CCP.

Mechanism of Cervi Cornu Pantotrichum in Treatment of Osteoarthritis Based on Network Pharmacology / 中国实验方剂学杂志

ObjectiveTo explore the mechanism of Cervi Cornu Pantotrichumin in the treatment of osteoarthritis by network pharmacology. MethodThe active ingredients and the corresponding targets of Cervi Cornu Pantotrichumin were screened out by a Bioinformatics Analysis Tool of Molecular mechANism of Traditional Chinese Medicine （BATMAN-TCM）. The targets related to osteoarthritis were obtained through GeneCards and Online Mendelian Inheritance in Man （OMIM）. The targets corresponding to the active ingredients and those related to osteoarthritis were intersected to reveal the common targets， and STRING was adopted to build a protein-protein interaction （PPI） network. DAVID was used for gene ontology （GO） annotation and Kyoto encyclopedia of genes and genomes （KEGG） pathway enrichment on the anti-osteoarthritis targets of Cervi Cornu Pantotrichumin， and R x64 3.6.3 was employed to produce the advanced bubble charts of GO terms and KEGG pathways. Cytoscape 3.7.2 was used to establish the “Chinese medicinal herb-active ingredient-target-signaling pathway” network. In vitro experiments were performed to detect the viability of RAW 264.7 cells exposed to oxidative stress and the tumor necrosis factor （TNF）-α level in RAW 264.7 cells with inflammation under the treatment by Cervi Cornu Pantotrichumin. ResultA total of 20 active ingredients of Cervi Cornu Pantotrichum were obtained， of which ceramide， 6'-O-β-D-glucosylgentiopicroside， cerebroside， oleuropein， sphingomyelin， and cholesterol ferulate did not meet the screening conditions. Therefore， a total of 14 active ingredients were finally screened out， and 303 and 3 093 targets of active ingredients and osteoarthritis were respectively obtained. The two target sets were taken to intersect， which revealed 92 common targets. GO annotation and KEGG pathway enrichment showed that the targets were mainly involved in redox process， positive regulation of RNA polymerase Ⅱ promoter transcription， inflammatory response， protein synthesis， osteoclast differentiation， TNF signaling pathway， signaling pathways in cancer， mammalian target of rapamycin （mTOR） signaling pathway， mitogen-activated protein kinase （MAPK） signaling pathway， and cyclic adenosine monophosphate （cAMP） signaling pathway. The results of in vitro experiments showed that a certain concentration of protein in Cervi Cornu Pantotrichum significantly increased the viability of RAW 264.7 cells exposed to H2O2-induced oxidative damage （P<0.05， P<0.01） and reduced the level of TNF-α in the RAW 264.7 cells experiencing lipopolysaccharide （LPS）-induced inflammation （P<0.05）. ConclusionBased on the network pharmacology method， the mechanism of the multi-component， multi-target and multi-pathway treatment of OA by antler antler was explained， and the anti-inflammatory and antioxidant activities of antler antler were confirmed， which provided theoretical guidance and scientific basis for further research on the treatment of OA by antler antler.

Quality assessment of different forms of Cervi Cornu Pantotrichum based on content of free amino acids / 中国中药杂志

In this study, we analyzed the composition and content of 25 free amino acids in 32 batches of different forms of Cervi Cornu Pantotrichum(CCP; one-branched, two-branched, and three-branched) from 15 producing areas. The clustering analysis and orthogonal partial least squares discriminant analysis(OPLS-DA) were performed based on the content of 25 free amino acids. Potential differential metabolites were identified based on VIP value. The results showed that there were 25 free amino acids in CCP, and the average content of essential, non-essential, and total amino acids was 6.13, 32.99, and 39.12 mg·g~(-1), respectively. The clustering analysis and OPLS-DA demonstrated that 25 free amino acids had different content among the three forms of CCP, of which two-branched CCP samples were separately gathered into a group. Five differential components, including glutamic acid, tryptophan, ornithine, γ-aminobutyric acid, and hydroxylysine, were screened out as potential quality markers for the identification of different forms of CCP. This study provides a theoretical basis for the quality evaluation, processing, and utilization of different forms of CCP.

Comparative study on chemical constituents of Cervi Cornu Pantotrichum in different forms of Cervus nipport / 中草药

Objective: In order to study the differences in chemical constituents of Cervi Cornu Pantotrichum in different forms of Cervus nipport. Methods: The content of polysaccharides, crude protein, amino acids, collagen, fatty acids, mineral elements, biogenic amines, nucleosides, and chondroitin sulfate composition of Cervi Cornu Pantotrichum in different forms of C. nipport was determined and compared by using UV spectrophotometer, Dumas azotometer, automatic amino acid analyzer, UPLC, HPLC, gas chromatograph, inductively coupled plasma mass spectrometer and atomic fluorescence photometer. Principal component analysis was used to evaluate the quality of Cervi Cornu Pantotrichum. Results: The content of polysaccharide, crude protein, amino acid, collagen, fatty acid, mineral element, biogenic amine, nucleoside and chondroitin sulfate in one-branched velvet antler was 8.40 mg/g, 44.82%, 42.24%, 23.23%, 6 234.69 mg/kg, 145.69 mg/g, 55.12 mg/kg, 2 271.87 mg/kg, and 0.74 mg/g, respectively; The content in two-branched velvet antler was 8.14 mg/g, 52.12%, 48.57%, 21.50%, 8 684.27 mg/kg, 126.40 mg/g, 76.14 mg/kg, 3 438.37 mg/kg, and 1.94 mg/g, respectively; The content in three-branched velvet antler was 8.64 mg/g, 51.86%, 45.49%, 22.31%, 9 100.78 mg/kg, 138.36 mg/g, 70.75 mg/kg, 2 507.82 mg/kg, and 1.84 mg/g, respectively. Conclusion: The chemical composition of Cervi Cornu Pantotrichum in different forms of C. nipport was different. The results of principal component analysis showed that the quality of the two-branched velvet antler was the best, the three-branched velvet antler was the second, the one-branched velvet antler was the worst. This study provides a reference for the quality evaluation and grading standards of different forms of Cervi Cornu Pantotrichum.

Research progress on chemical constituents and pharmacological effects of different Cervi Cornu Pantotrichum / 中草药

Cervi Cornu Pantotrichum has a long medicinal history in China and is often used as a tonic Chinese medicine. Because of the complex provenances, various specifications and unclear efficacy substance, the industry of Cervi Cornu Pantotrichum lacks corresponding quality standards. The paper summarizes the research progress of chemical composition and pharmacological effects of different provenances, different growth stages, different forms, different processing methods, different parts of Cervi Cornu Pantotrichum and provides reference for the in-depth application of Cervi Cornu Pantotrichum in medical and functional food industries.

Progress on carbon dots research and its application prospect in Cervi Cornu Pantotrichum / 中草药

Carbon dots are an emerging nanomaterial with excellent fluorescent properties. Compared with traditional organic dyes and semiconductor quantum dots, carbon dots possess advantages of low toxicity and good biocompatibility. At present, carbon dots are widely used in many fields such as analytical detection, fluorescence imaging and drug delivery. Cervi Cornu Pantotrichum, a rare mammalian organ with ability of full regeneration, has been used as famous traditional Chinese medicine. The biological functions and efficacy of Cervi Cornu Pantotrichum are closely related to their chemical constituents. In this paper, we briefly reviewed the progress of carbon dots research in fluorescence detection, fluorescence imaging and phototherapy, carried out the bioimaging and cell cytotoxicity test by carbon dots, and discussed the feasibility of carbon dots in biological features and chemical composition analysis of Cervi Cornu Pantotrichum.

[Effects of different processing methods on water soluble polysaccharide contents and monosaccharide compositions in Cervi Cornu Pantotrichum].

The water-soluble polysaccharides in antler velvet were extracted by water extraction and alcohol precipitation. The content was determined by a phenol-sulfuric acid colorimetry. After hydrolysis and derivatization, the various monosaccharide compositions of water-soluble polysaccharides from different parts of the antler velvet after different processing methods were analyzed by ultra high performance liquid chromatography (UPLC). The results showed that the water-soluble polysaccharide contents in wax slices, powder slices, gauze slices and bone slices of the boiled antler velvet were 1.74, 1.67, 1.03 and 1.13 g/kg, respectively, whereas those of the freeze-dried antler velvet were 2.77, 3.07, 1.22 and 3.20 g/kg, respectively. The water-soluble polysaccharide contents in in the same four parts of the antler velvet processed antler velvet without and with blood were 1.55, 1.78, 0.96, 0.77 g/kg, and 1.69, 1.64, 1.01, 1.31 g/kg, respectively. Eight monosaccharides (mannose, glucosamine, ribose, glucuronic acid, galacturonic acid, aminogalactose, glucose and galactose) were detected in the compositions of water-soluble polysaccharides in the antler velvet by using different processing methods. For the same part, the monosaccharide contents in the boiled antler velvet were lower than those in the freeze-dried antler velvet, and the monosaccharide contents (except those of glucosamine and aminogalactose) in the antler velvet processed without blood were lower than those in the antler velvet processed with blood. For different parts processed with the same method, the monosaccharide contents in wax and powder pieces were higher than those in gauze slices and bone pieces. This study provides a theoretical reference for velvet processing and product development.

Analysis on differential metabolites of Cervi Cornu Pantotrichum based on metabolomics / 中草药

Objective: To identify the differential metabolic small molecule substances in Cervi Cornu Pantotrichum (CCP) of different segments based on UPLC-QTOF/MS metabonomics technology, and analyze the metabolic pathways involved by differential metabolites combined with histological observation. Methods: The standard trident antler of sika deer (Cervus nippon) were treated by freeze-drying. The tissue morphology was observed by section. The tissue morphology was divided into three sections from top to bottom, VAU, VAM, and VAB. The differential metabolites were analyzed by UPLC-QTOF/MS technology, principal component analysis, and least squares discriminant analysis. Results: A total of 124 kinds of metabolic small molecular substances were identified in CCP. Based on the FC value of differential metabolites higher than 2.2, 16 substances with different relative contents in different parts of CCP were further screened, and the relative content of each substance in different parts were compared. Conclusion: This experiment is based on non-targeted metabonomics to analyze the composition of metabolic small molecules in freeze-dried CCP. By comparing the adjacent zones, it is found that there are some differences in endogenous metabolites among the zones, which provides data support for revealing their functions.

Quality evaluation of Cervi Cornu Pantotrichum based on fingerprint analysis and quantitative analysis of multi-components / 中草药

Objective: To establish the fingerprints of nucleosides in Cervi Cornu Pantotrichum (CCP) pieces by HPLC method, perform cluster analysis and principal component analysis (PCA), and compare the differences of six nucleosides in CCP pieces. Methods: A total of 16 batches of CCP pieces from different origins were determined by HPLC. Sixteen batches from different origins in China were collected to assess the similarities according to similarity evaluation for “chromatographic fingerprint of traditional Chinese medicine” (2012), and four kinds of decoction pieces were distinguished and compared by chemometric methods such as principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA). Results: The HPLC fingerprints of CCP nucleosides were established and the similarity was above 0.960. Six common peaks of uracil, adenine, hypoxanthine, uridine, inosine, and guanosine were identified. Among them, uracil, hypoxanthine, and inosine were different compounds, which can be used as a quality control indicator for identifying and distinguishing CCP pieces. Conclusion: The CCP nucleoside fingerprints established by the method are characterized by strong features and simple methods. The combination of six nucleosides can better control the quality, which has guiding significance and reference value for the identification and quality control of CCP.

Identification of Cervi Cornu Pantotrichum and Cervi Cornu by Allele-specific PCR / 中国实验方剂学杂志

Objective: To obtain a rapid,efficiency and convenient polymerase Chain reaction(PCR) identification method for medicinal Cervi Cornu Pantotrichum,Cervi Cornu and its common adulterates. Method: Based on three single nucleotide polymorphisms (SNP) of Cytb gene DNA sequences among Cervus nippon,C. elaphus and its adulterants,a pair of species-specific primers (LR-238.F and LR-238.R) was designed,the reaction conditions were optimized,and the PCR method for identification was explored and verified in terms of tolerance and feasibility. Result: Through the established allele-specific PCR method,under the annealing temperature of 56℃ and cycle number of 35,250 bp of fragments were amplified from DNA templates of Cervi Cornu Pantotrichum,Cervi Cornu and its subspecies in origin animal samples as well as herbal medicines. All of the adulterants species of Przewalskium albirostris,Cervus eldi,Odocoileus hemionus,Dama dama,Alces alces,Elaphurus davidianus,Capreolus pygargus,Rusa unicolor and Rangifer tarandus were negative by the PCR assay. Conclusion: The identification primer is highly specific,and the allele-specific PCR identification method established in this paper can accurately identify the medicinal Cervi Cornu Pantotrichum and Cervi Cornu.

Production of 5-hydroxymethylfurfural in processing of Cervi Cornu Pantotrichum and investigation on its influencing factors / 中草药

Objective: To investigate the causes and influencing factors of 5-hydroxymethylfurfural (5-HMF) during the processing of Cervi Cornu Pantotrichum (CCP). Methods: HPLC was used to determine the content of 5-HMF in CCP with different processing methods and different parts. Moreover, the contents of total sugar, reducing sugar, amino acid, and Ca, Mg, Fe, and Cu in velvet antler were determined, and the effects of several factors on 5-HMF production during processing were discussed by comparing the contents of above mentioned substances in velvet antler with different processing methods. Results: The contents of 5-HMF of boiled CCP were significantly higher than that of freeze-dried CCP (P < 0.05) in same parts, indicating that high temperature intensified caramelization reaction and Maillard reaction to produce more 5-HMF in boiled CCP; The contents of 5-HMF of CCP with blood were significantly higher than that of CCP without blood (P < 0.05), CCP with blood was rich in total sugar, reducing sugar and amino acid and these substances can provide sufficient substrate to produce more 5-HMF; The content of 5-HMF in wax slices was significantly higher than other parts (P < 0.05), and rich content of total sugars, reducing sugars, amino acids, and differentially distributed mineral elements all contribute to the production of 5-HMF in wax slices. Conclusion: The production of 5-HMF in processing CCP is the result of the combined action of total sugar, reducing sugar, amino acid, and mineral elements at different temperatures.

[Research progress on identification and quality control of Cervi Cornu Pantotrichum].

Cervi Cornu Pantotrichum, as a traditional Chinese medicine, has great potential for development. However, the identification and quality control system is not perfect, leading to the market chaos and chronic slow growth in deep processing of Cervi Cornu Pantotrichum. This paper gives an overview of present situation in identification and quality control system of the Cervi Cornu Pantotrichum, and analyzes present problems. Based on these results, the feasibility study scheme in identification and quality control system for Cervi Cornu Pantotrichum would be then put forward, providing ideas to establish its comprehensive evaluation system.

Research progress on identification and quality control of Cervi Cornu Pantotrichum / 中国中药杂志

Cervi Cornu Pantotrichum, as a traditional Chinese medicine, has great potential for development. However, the identification and quality control system is not perfect, leading to the market chaos and chronic slow growth in deep processing of Cervi Cornu Pantotrichum. This paper gives an overview of present situation in identification and quality control system of the Cervi Cornu Pantotrichum, and analyzes present problems. Based on these results, the feasibility study scheme in identification and quality control system for Cervi Cornu Pantotrichum would be then put forward, providing ideas to establish its comprehensive evaluation system.

Analysis on fatty acids of Cervi Cornu Pantotrichum with different processing methods by gas chromatography / 中草药

Objective: To study the differences and the trends of fatty acids content in different parts of Cervi Cornu Pantotrichum (CCP) with different processing methods. Methods: KOH-CH3OH esterification and n-hexane extraction were combined as the pretreatment method to obtain the fatty acid samples, and then the samples were analyzed by gas chromatography. Results: The fatty acids content in wax slices, powder slices, and bee slices of CCP without blood were 16.00, 10.32, and 5.51 g/kg. The fatty acids contents in wax slices, powder slices, and bee slices of CCP with blood were 14.81, 6.04, and 4.88 g/kg. The fatty acids contents in wax slices, powder slices, and bee slices of CCP with boiling processing were 9.06, 6.20, and 4.23 g/kg. The fatty acids contents in wax slices, powder slices, and bee slices of CCP with freeze-drying were 9.46, 7.54, and 6.23 g/kg. Conclusion: The contents of fatty acids in CCP with different processing methods are different. The content of fatty acids in CCP without blood is higher than that in CCP with bloods, and the content of fatty acids in CCP with boilding processing is lower than that in CCP with freeze-drying. The trend of different parts of antlers with the same processing method shows decreasing one by one from wax slices to bee slices. The main saturated fatty acids in the antlers are palmitic acid (C16:0) and stearic acid (C18:0). The mono unsaturated fatty acids are mainly oleic acid (C18:1, cis-9). The content of γ-linolenic acid (C18:3, cis-9, 12, 15), twenty C two acid (C20:2, cis-11, 14), and four arachidonic acid (C20: 4, cis-5, 8, 11, 14) in polyunsaturated fatty acids are higher than others.

Effects of different processing methods on content of crude protein and hydrolytic amino acids in Cervi Cornu Pantotrichum / 中草药

Objective: To compare the contents of protein and amino acids in different parts of Cervi Cornu Pantotrichum (CCP) with different processing methods, in order to provide a guidance for the processing and comprehensive utilization of CCP. Methods: The techniques of Dumas combustion and cation-exchange chromatography were respectively adopted to determinate the contents of crude protein and 17 amino acids in different parts of CCP with different processing methods, and the difference was compared. Results: The content of crude protein in powder slices of CCP with blood was higher than that of CCP with blood (P 0.05). The crude protein content in wax slices of CCP with freeze-drying processing was less than that with boiling processing (P 0.05). The content of crude protein and TAA in wax slices of CCP is both significantly higher than that in powder slices, bee slices (P 0.05). Conclusio:n The difference is existed in content of crude protein and amino acids in CCP with different processing methods. The wax silices of CCP are significantly higher than that of powder slices and bee slices. And the difference in powder slices and bee slices is not significantly. The distribution of crude protein and TAA in different parts of CCP with freeze-drying processing is more uniform than CCP with boiling processing.

Research progress on material basis, pharmacological effects, clinical application, and quality control of Cervi Cornu Pantotrichum / 中草药

Cervi Cornu Pantotrichum (CCP) is one of the famous Chinese materia medica, which has been widely used in clinical application with a long history. In this paper, the compositions, activities and quality control of antler were reviewed according to ancient herbal books and modern literature. By putting forward the research idea about the relationship of functional composition-activity-quality control, to elucidate the quality markers of antler. It would provide a reference for the relevant research of explaining pharmacodynamic material basis of Guilingji and formulating CCP quality evaluation criteria.

Development of multiplex PCR assay for authentication of Cornu Cervi Pantotrichum in traditional Chinese medicine based on cytochrome b and C oxidase subunit 1 genes.

This study describes a method for discriminating the true Cervus antlers from its counterfeits using multiplex PCR. Bioinformatics were carried out to design the specific alleles primers for mitochondrial (mt) cytochrome b (Cyt b) and cytochrome C oxidase subunit 1 (Cox 1) genes. The mt DNA and genomic DNA were extracted from Cervi Cornu Pantotrichum through the modified alkaline and the salt-extracting method in addition to its counterfeits, respectively. Sufficient DNA templates were extracted from all samples used in two methods, and joint fragments of 354 bp and 543 bp that were specifically amplified from both of true Cervus antlers served as a standard control. The data revealed that the multiplex PCR-based assays using two primer sets can be used for forensic and quantitative identification of original Cervus deer products from counterfeit antlers in a single step.

Ultrafiltration and concentration used for separating insulin-like growth factor-1 from Cervi Cornu Pantotrichum / 中草药

Objective: To establish the ultrafiltration process of concentrating insulin-like growth factor-1 (IGF-1) from Cervi Cornu Pantotrichum (CCP). Methods: The ultrafiltration technique was utilized to concentrate the IGF-1 with CCP freeze-dried powder as raw material. The polyethersulfone (PES) membrane and modified PES membrane were compared and studied. The effects of relative molecular weight cutoff (RMWCO), pressure, and pH value on IGF-1 ultrafiltration process were investigated. Furthermore, the cleaning methods were investigated, and the optimal conditions were obtained. Results: The optimal conditions were as follows: The RMWCO of PES membrane was 4 000, the RMWCO of modified PES membrane was 5000, the pressure was 0.3 MPa, and the pH value was 12. The recovery rate of IGF-1 and protein reached 68.32% and 69.20%, respectively by the PES membrane, while the recovery rate of IGF-1 and protein reached 57.17% and 87.43% respectively by the modified PES membrane. Four different methods were employed to clean the membrane. The results showed that the membrane immersed in 0.4% NaOH-0.1% NaClO solution for 24 h was the appropriate cleaning method, by which the PES membrane permeation flux recovery rate could reach 98.65% and the modified PES membrane permeation flux recovery rate could reach 93.75%. Conclusion: The IGF-1 could be easily concentrated and separated using the ultrafilter condensing technique. The process is easy and stable, and the membrane is easy to be cleaned and has high recovery rate, which has the high potential of industrial production and application.