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Objective: EST-SSR loci were identified and analyzed based on the transcriptome sequencing data in Polygonatum cyrtonema, in order to develop SSR markers suitable for evaluation and application of germplasm resources on P. cyrtonema. Methods: SSR loci were identified and analyzed in all of 126 544 Unigenes by using MISA tool. SSR primers were designed by using Primer 3.0 software and 50 pairs of SSR primers were randomly selected for validation test. Results: A total of 12 317 SSR loci, including the types of 2-6 nucleotide repeats with occurring frequency of 1/5.91 kb, were identified from 9 982 Unigenes in P. cyrtonema transcriptome. The distribution frequency of SSRs was 9.73%. Dinucleotide repeat was the main type, accounting for as much as 53.14% of all SSRs, followed by trinucleotide repeat (33.31%). The validation test on 50 pairs of SSR primers showed that 29 of them (58%) generated fragments with expected molecular size from P. cyrtonema. The capillary electrophoresis using fluorescence-labeled SSR markers showed that nine genotypes were identified at seven SSR loci in P. cyrtonema, which further demonstrated the validity of these SSR primers. Conclusion: There are numerous SSRs in P. cyrtonema transcriptome with high frequency, rich repeat types and relatively high polymorphic, which will provide abundant valuable candidate markers for genetic diversity analysis and genetic mapping construction in P. cyrtonema, also can be used as a technical tool for molecular identification among Polygonatum species and for molecular marker assistant breeding in superior cultivars of P. cyrtonema.
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Objective: To study the genetic diversity and geographical distribution of Polygonatum cyrtonema resources. Methods: ISSR technique was applied to analyze 118 individuals from 20 P. cyrtonema provenances in six provinces, including Anhui, Jiangxi, Fujian, Hunan, Hubei, and Zhejiang. Results: The results showed that 130 clear bands were amplified by 16 primers with 123 polymorphic bands and the average percentage of polymorphic loci (PPL) was 94.62%, PPL within provenances was 33.85%-60.00%. Nei's genetic diversity index (He) was 0.183 8, Shannon's information index (I) was 0.267 4 and gene differentiation index (Gst) was 0.529 3. There were abundant genetic diversities existing in wild resources of P. cyrtonema. The UPGMA clustering analysis revealed that individuals from the same provenance were almost clustered together firstly, explaining that the genetic differentiation among different provenances was higher than those within provenances. When genetic similarity (GS) was 0.61, 118 germplasms can be divided into four categories, including Wuyi Mountains, Wuling and Luoxiao Mountains, Dabie Mountains, Donggong, and Tianmu Mountains. Conclusion: P. cyrtonema has high genetic diversity, genetic variation was closely related to mountains, and the isolation of plains and water areas between mountains was one of the main causes of genetic differentiation among groups. This study had essentially theoretical value and practical significance for the protection of the germplasm resources and the breeding of the species.
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Objective: In this work, phylogenetic analysis was used to compare the ITS2 and psbA-trnH sequences of Polygonatum cyrtonema samples from different geographical sources, so as to explore the genetic diversity and genetic relationship of these resources. Methods: PCR method was used to amplify the regions of ITS and psbA-trnH, and the sequences of ITS2 and psbA-trnH were obtained after the amplified fragment sequences were blasted in NCBI database. The neighbor joining (NJ) and maximum parsimony (MP) methods were used to construct phylogenetic trees and Kimura two-parameter (K2-P) model was used to calculate the genetic distance of different samples. Mega and DNAman softwares were applied for mutiple alignment of ITS2 and psbA-trnH sequences of 25 samples of P. cyrtonema. Results: The lengths of ITS2 and psbA-trnH sequences of Anhui Qingyang and Fujian Taining samples of P. cyrtonema were 224 bp and 620 bp, respectively. The lengths of ITS2 and psbA-trnH of the remaining 24 samples were 225 bp and 621 bp, respectively. ITS2 and psbA-trnH had seven and four mutation points, respectively. These 25 samples were clustered into two groups based on ITS2 sequences. Five samples in Hunan and Guizhou were clustered into one group, while the other 20 samples were clustered into another group. The genetic distance showed that the samples from Huaxi and Jianhe in Guizhou Province and Jianyang in Fujian Province had the largest genetic distance. Phylogenetic tree constructed by psbA-trnH sequences were unable to distinguish 25 samples from different geographical sources. Conclusion: Phylogenetic and mutation analysis will provide the theoretic foundation to utilize the resources of P. cyrtonema, investigate their evolution, and evaluate their genuineness. The results of mutation point will also be used in the identification of related P. cyrtonema resources.
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Objective: To find a solution to the problems in the growth of tissue cultured seedlings of Polygonatum cyrtonema, such as low seedling survival rate, poor growth, and leaf disease. Method: In this study, tissue cultured seedlings of P. cyrtonema were used as testing plants. Two types of commercially-available cultivation medium commonly used in Fujian Province, as well as six types of cultivation medium mixed using peat soil, pearlite, roseite, sand, fungi residues and plant ash were used as the raw materials; Among them, a customized nutrient solution was added into the T5 and T7 cultivation media. Besides measuring the physicochemical properties of the cultivation media mentioned above, the study focused on the evaluation of their influence on the rate of grown-up seedlings, biological characteristics, growth biomass, strength index and incidence of leaf disease of P. cyrtonema. It also attempted to find the correlation between the physicochemical properties of the cultivation media and the biological characteristics, growth biomass, as well as the strength index of tissue cultured seedlings of P. cyrtonema. Results: The difference in the cultivation media had a great influence on the tissue cultured seedlings of P. cyrtonema in the rate of grown-up seedlings, biological characteristics, growth biomass, strength index and incidence of leaf disease. The difference in bulk density of the cultivation media had a notable negative correlation with the plant height of the tissue cultured seedlings of P. cyrtonema. The difference in the organic content of the cultivation media had a notable positive correlation with the fresh weight and dry weight of the tissue cultured seedlings of P. cyrtonema. Treated with the T5 cultivation medium we prepared, the tissue cultured seedlings of P. cyrtonema were in better growing condition in the rate of grown-up seedlings, morphological index, growth bomass, strength index, and no disease was found on the leaf. Conclusion: The prepared T5 was worth being widely used as the cultivation medium for growth of tissue cultured seedlings of P. cyrtonema.
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Objective: To investigate the secondary metabolites of Aspergillus ochraceus, an endophytic fungus isolated from Polygonatum Cyrtonema. Methods: Compounds were isolated and purified from the EtOAc extract by using chromatography technology and their structures were established on the basis of comprehensive spectroscopic analysis. Results: A total of 15 compounds were obtained and their structures were elucidated as 6,7-dihydroindolizin-8(5H)-one (1), polygonatine A (2), 8-oxo-5,6,7,8-tetrahydro-3-indolizinyl methyl acetate (3), 8-hydroxyketone (4), cyclo-(L-Leu-L-Ile) (5), alternariol (6), seco- patulolide C (7), n-butyl-β-D-fructopyranoside (8), Nb-acetyltryptamine (9), N-trans-cinnamoyltyramine (10), 5-hydroxy- methylfurfural (11), 5,7-dihydroxy-6,8-dimethyl-3-(4’-hydroxybenzyl) chroman-4-one (12), 5,7-dihydroxy-6-methyl-8- methoxy-3- (4’-hydroxybenzyl) chroman-4-one (13), 25R-3-β-hydroxyspirost-5-en-12-one (14), and 25S-3-β-hydroxyspirost-5-en-12-one (15). Conclusion: All chemical constituents are isolated from A. ochraceus for the first time.
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This paper reviewed the ancient and modern literatures and made a textual research on the original plant and medicinal materials of Jiuhua Polygonatum Cyrtonema. Original plant of Jiuhua P. Cyrtonema is Polygonatum cyrtonema, and the leaf order type is interleaf type. The processing method of Jiuhua P. Cyrtonema as medicine is mainly nine steaming and nine sunning. In modern times, Jiuhua P. Cyrtonema is used from medicinal materials to edible food, which can be applied for the food development to lay the foundation for the modern production of Jiuhua P. Cyrtonema. However, the further research on genuineness of Jiuhua P. Cyrtonema is also needed to do. It can be determined that P. cyrtonema from the Hunan, Guizhou, Guangxi, and Jiuhua Mountain is better, so the further research need to be done to determine whether the modern Jiuhua P. Cyrtonema can be genuine medicinal materials.
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Polygonatum cyrtonema, which is growing in Jiuhua Mountain, is genuine medicinal materials. The main processing methods of it are cutting, single-steamed, re-steamed, nine times steamed and nine times shined, wine-steamed, wine-stewed, copying and so on. Its effective components are polysaccharides and steroidal saponins, and they also have pharmacological activities such as anti-oxidation, regulating immunity, lowering blood sugar, antitumor, antibiosis, and antiviral. The development history and modern research progress of processing technology of P. cyrtonema in Jiuhua Mountain were summarized, which provides reference and basis for its processing research and lays the foundation for its standard operation rules of GMP processing.
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Objective: To develop a method for determination of monosaccharide composition of Polygonatum cyrtonema polysaccharides (PCR) by high performance anion-exchange chromatography with pulsed amperometric detection (HPAEC- PAD). Methods: The water-soluble crude polysaccharides were extracted from the P. cyrtonema Hua, then the polysaccharides were hydrolyzed with sulfuric acid, then separated by CarboPac PA1 column (250 mm × 4 mm, 10.0 μm) and with gradient elution, determination of monosaccharide composition in PCP by HPAEC-PAD. Results: The results showed that within the range of 1-100 mg/L it has good linearity (r > 0.999), The detection limitation was 0.015-0.025 mg/L, 1.35%-2.80% RSD, the recovery rates were 88.36%-111.3%. The results showed that the polysaccharides in P. cyrtonema Hua from different regions were composed of rhamnose, arabinose, galactose, glucose, mannose, and fructose. Conclusion: The proposed method has good separation effect and high sensitivity, which can be used for the study on monosaccharide composition and content determination of PCP.
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Objective To study the reproductive biology of Chinese herbal medicine Polygonatum cyrtonema Hua. Methods The flowering habit of P. cyrtonema was observed and recorded regularly. The pollen viability at different flowering periods was detected by TTC. The stigma activity at different flowering periods was measured by the benzidine-hydrogen peroxide method. The hybridization index was estimated and pollen-ovule ratio was calculated. The breeding system of P. cyrtonema was determined by six different artificial pollination treatments. Results The flower buds of P. cyrtonema appeared at April 5. They bloomed at April 10 and fruited early in May, flowering about 30–40 days. The pollen began to have viability since the anthers were not cracked, and loss viability after flowers withered. The pollen viability at the bloom day was about 73.06%±5.19%. The stigmas began to have activity before flowers opening and lost it when the flowers began to decline. The hybridization index was 4, so the breeding system of P. cyrtonema Hua was partly self-compatible and xenogamy needing pollinator according to Dafni's standard.The pollen-ovule ratio was 1799.20±445.54,therefore the breeding system of P.cyrtonema Hua was facultative xenogamy according to Cruden's standard. The results of artificial pollination experiments showed that P. cyrtonema Hua had three breeding methods,including apomixis,selfing and outcrossing.The fruit setting rate of P. cyrtonema Hua was greatly influenced by external conditions such as strong wind and rain.Generally,P.cyrtonema Hua had a low fruit-set percentage because of obvious flower and fruit dropping. The natural fruit setting rate was only 17.46%. Conclusion The coexistence of apomixes, sexual and asexual reproduction in P. cyrtonema are the survival strategy adapting to different habitat types.
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Objective To study the reproductive biology of Chinese herbal medicine Polygonatum cyrtonema Hua. Methods The flowering habit of P. cyrtonema was observed and recorded regularly. The pollen viability at different flowering periods was detected by TTC. The stigma activity at different flowering periods was measured by the benzidine-hydrogen peroxide method. The hybridization index was estimated and pollen-ovule ratio was calculated. The breeding system of P. cyrtonema was determined by six different artificial pollination treatments. Results The flower buds of P. cyrtonema appeared at April 5. They bloomed at April 10 and fruited early in May, flowering about 30–40 days. The pollen began to have viability since the anthers were not cracked, and loss viability after flowers withered. The pollen viability at the bloom day was about 73.06%±5.19%. The stigmas began to have activity before flowers opening and lost it when the flowers began to decline. The hybridization index was 4, so the breeding system of P. cyrtonema Hua was partly self-compatible and xenogamy needing pollinator according to Dafni's standard.The pollen-ovule ratio was 1799.20±445.54,therefore the breeding system of P.cyrtonema Hua was facultative xenogamy according to Cruden's standard. The results of artificial pollination experiments showed that P. cyrtonema Hua had three breeding methods,including apomixis,selfing and outcrossing.The fruit setting rate of P. cyrtonema Hua was greatly influenced by external conditions such as strong wind and rain.Generally,P.cyrtonema Hua had a low fruit-set percentage because of obvious flower and fruit dropping. The natural fruit setting rate was only 17.46%. Conclusion The coexistence of apomixes, sexual and asexual reproduction in P. cyrtonema are the survival strategy adapting to different habitat types.
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Objective: To study the chemical constituents from the aerial part of Polygonatum cyrtonema. Methods: The compounds were isolated and purified by means of chromatographic techniques and their structures were identified on the basis of spectral features. Results: Fifteen known compounds were isolated in methanol extract from the aerial part of P. cyrtonema and their structures were identified as (3R)-5,7-dihydroxy-3-(2'-hydroxy-4'-methoxybenzyl)-chroman-4-one (1), 5,7-dihydroxy-6-methyl-3-(2',4'-dihydroxybenzyl)- chroman-4-one (2), 5,7-dihydroxy-6-methyl-3-(4'-hydroxybenzyl)-chroman-4-one (3), (3S)-3,7-dihydroxy-8-methoxy-3-(3',4'- methylenedioxybenzyl)-chroman-4-one (4), apigenin (5), kaempferol (6), vanillic acid (7), trans-p-hydroxycinnamic acid (8), trans-p-hydroxycinnamic acid methyl ester (9), saliylic acid (10), (+)-syringaresinol (11), balanophonin B (12), caffeic acid (13), phenylalanine (14), and coniferaldehyde (15). Conclusion: Compounds 1, 2, 7, 11, 14, and 15 are firstly obtained from P. cyrtonema. Compounds 3, 4, and 12 are isolated from the plants of Polygonatum Mill. for the first time.
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Polygonti Rhizome belongs to the family Liliaceae. Its tuber has the medicinal functions of tonifying the kidney to arrest spontaneous emission (Bushen Yijing) and replenishing yin essence and moisturizing the viscera (Ziyin Runzao), and has been used for many years in Chinese medicine. This paper mainly summarizes the research advances on chemical constituents and pharmaco- logical activities of P. sibiricum, P. kingianum, and P. cyrtonema accepted in Chinese Pharmacopoeia, and provides the reference for their further development and utilization.
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Objective:Eighteen inorganic elements were measured from the rhizoma and fibrous root of Polygonatum Cyrtonema Hua. The contents of K, Fe,Mg,Ba,Cu,Mn and Bi of them in rhizoma were higher than that of fibrous root, where as the contents of Na,Al,Ca,Ge,P,Zn and Sr elements in rhizoma were lowere than that of fibrous. Both contained As,Hg,Pb and Cd elements. In the meantime, 16 mino acid were determined. The total content of them was 8.92% for rhizoma and 9.61% for fibrous. Cystine, cysteine, tryptophan and ornithine couldn't be detected.