Studies On The Chemical Constituents And Bioactivity Of Two Polygonatum Plants Of Qinling Mountain

Based on the theories and principles of phytochemistry and modern chemical isolation and purification methods, with the conventional and new skill, the chemical constituents and bioactivities of Polygonatum odoratum and Ploygonatum cirrhifolium of Qinling mountain were studied. Thirty compounds were isolated, four new compounds together with twenty one known compounds structure were elucidated by spectroscopic and the data reported in literature, the new compounds were C-methylated homoisoflavanone, 25S steroidal saponins and amide compound respectively. The biological activities of some compounds were tested to preliminary discuss its construction and effect relations. The best technology of extraction and isolation of polysaccharide from the rhizomes of Polygonatum odoratum and Polygonatum cirrhifolium were determined.The results are concluded as follows:1. The antisepsis and allelopathy activities of Polygonatum odoratum were studied for the first time. The ethyl acetate extracts had strong inhibition effects on the hvphae growth of Colletotrichum lagenarium, Trichothecium roseum, Verticillium dahliae with the EC50 values of 151.77μg·mL-1, 114.24μg·mL-1 and 284.12μg·mL-1 respectively in tested species. The n-butanol extracts had strong inhibitory effects on the hvphae growth of Exserohilum turcicum with the EC50 values of 137.05μg·mL-1. The petroleum extracts had ceartain inhibiton against four plant pathogenic fungi growth, its inhibiton rate reach 35%. To receiver plants of allelopathic effect, every extracts inhibited seed germination and seedling growth , the allelopathic effect intensity was related to the concentration and strengthen along with the concentration raise, but petroleum extracts inhibited root growth at high concentration and stimulated it at low concentration. The average allelopathic effect of ethyl acetate extracts was strongest, the next was that aqueous extracts, n-butanol extracts and petroleum extracts. The ethyl acetate extracts had extremely strong inhibition on the seed germination, seedling and root growth. The different receiver plant had distinct sensitive to the allelopathic effect, wheat was more sensitive than cucumber and carrot.2. The chemical constituents from the rhizomes extract of Polygonatum odoratum of Qinling mountain were studied for the first time. Nineteen compounds were isolated, the structure of three new compounds and thirteen known compounds were elucidated by spectroscopic and the data reported in literature. Three new compounds structure were identified as 4',8-dimethoxy-5,7-dihydroxy-6-methyl–homoisflavanone(Ⅰ-2), 3-O-β-D- glucopyranosyl-(1→2)-[β-D-xylopyranosyl-(1→3)]-β-D-glucopyranosyl-(1→4)-galactopyra-nosyl-25(S)-spirost-5-en-3β,12β-diol(Ⅰ-8),3-O-β-D-glucopyranosyl-(1→2)-[β-D-xylopyra- nosyl-(1→3)]-β-D-glucopyranosyl-(1→4)-galactopyranosyl-25(S)-spirost-5,14-dien-3β-ol (Ⅰ-9). Eight compounds were isolated for the first time from the genus Polygonatum, they were identified as 9,19-cyclolart-25-en-3β,24(R/S)-diol(Ⅰ-1), 4'-methoxy-5,7-dihydroxy- 6,8-dimethyl-homoisflavanone(Ⅰ-3), 4',5,7-trihydroxy-6-methyl-8-methoxy-homoisflavan- one (Ⅰ-5) and 4',5,7-t- rihydroxy-6-methyl-homoisflavanone (Ⅰ-6),α-palmitic-glycerolester (Ⅰ-10), (Z)-6-nonadecenoic acid(Ⅰ-14), methyl palmitate(Ⅰ-15)and octacosanoic acid (Ⅰ-16). Other compounds were identified as 4',5,7-trihydroxy-6,8-dimethyl-homoisflava- none (Ⅰ-4), 3-O-β-D-glucopyranosyl-(1→2)-[β-D-xylopyranosyl-(1→3)]-β-D-glucopyran- osyl-(1→4)-galactopyranosyl-25(S)–spirost-5-en-3β,14α-diol (Ⅰ-7),β-sitosterol(Ⅰ-11), dauvosterol (Ⅰ-12) and salicylic acid(Ⅰ-13). Besides, the 2D-NMR spectrum data of compoundⅠ-3 andⅠ-5 were supplement.3. The antisepsis and allelopathy activities of some compounds from Polygonatum odoratum were tested for the first time. The compoundⅠ-1 had strong inhibition effect on the growth of Colletotrichum lagenarium with inhibiton rate 100% ,and its inhibition effect correspond to erythromycin against Bacterium prodigious growth. CompoundⅠ-7 ~Ⅰ-9 both had strong inhibition against Exserohilum turcicum growth.CompoundⅠ-7 only had certain inhibition against Escherichia coli growth in testesd bacteriums, but compoundⅠ-8 and compoundⅠ-9 had strong inhibition against Bacillus cereus and Corynebacterium sepedonium growth respectively. In four C-6 methylated homoisoflavanones, CompoundⅠ-2 andⅠ-5 had strong inhibition against many bacteriums growth. The activities of compoundⅠ-2 (4',8-dimethoxy)was strongest, next was that compoundⅠ-5 (4'-hydroxy-8-dimethoxy), compoundⅠ-4 (4',8-dimethyl) and compoundⅠ-6 (4'-hydroxy). The average allelopathic effect of four C-6 methylated homoisoflavanones was similar to their antisepsis activities.4. The antisepsis and allelopathy activities of Polygonatum cirrhifolium rhizomes were studied for the first time. The n-butanol extracts had strong inhibitory effects on the hvphae growth of Monilia frugtigena, Exserohilum turcicum, and Verticillium dahliae with the EC50 values of 261.26μg·mL-1, 376.03μg·mL-1 and 430.05μg·mL-1 respectively in 10 plant patho- genic fungi. The ethyl acetate extracts had strong inhibitory effects on the hvphae growth of Valsa mali and Phytophthora infestans with the EC50 values of 386.00μg·mL-1, 865.75μg·mL-1 respectively, the toxicity were strengthen along with the concentration raise. To receiver plants of allelopathic effect, The average allelopathic effect of ethyl acetate extracts was strongest, then n-butanol extracts, aqueous extracts and petroleum extracts. The ethyl acetate extracts had extremely strong inhibition on the growth of seedling and root, and n-butanol extracts on the seed germination also. The different receiver plant had distinct sensitive to the the allelopathic effect, cucumber was more sensitive than carrot and wheat.5. The chemical constituents from the rhizomes extract of Polygonatum cirrhifolium were studied for the first time. Eleven compounds were isolated from the rhizomes of Ploy- gonatum cirrhifolium. A new compound structure was identified as Huangjing amide[N,N-bis(2,5-dihydroxybenzoyl)-2,5-dihydroxybenzamideⅡ-1]. Eight compounds were isolated for the first time from the Polygonatum cirrhifolium, they were identified as, diosgenin(Ⅱ-2), (25R/S)-spirost-5-ene-3β-ol-3-O-α-L-rhamnopyranosy (1→2)- [α-L-rham- nopyranosyl(1→4)]-β-D-glucopyranoside(Ⅱ-3), (25R)-spirost-5-ene-3β-ol-3-O-[α-L-rham- nopyranosyl(1→4)]-β-D-glucopyranoside(Ⅱ-4), n-butyl-β-D-fructopyrano-side(Ⅱ-5),β- sitosterol(Ⅱ-6),dauvosterol(Ⅱ-7), (Z)-6-nonadecenoic acid(Ⅱ-8), (Z)-6-stearic acid (Ⅱ-9). Besides, the 25S spectrum data of compoundⅡ-3 was supplement for the first time.6. The antisepsis activities of several compounds from Polygonatum cirrhifolium were tested for the first time. CompoundⅡ-3 and CompoundⅡ-4 had certain inhibition against Bacillus subtilis and Proteus vulgaris growth, but compoundⅡ-2 had strong inhibition against Bacillus cereus, Bacillus subtilis and Proteus vulgaris growth. CompoundⅡ-3 had strong inhibitory effects on the hvphae growth of Exserohilum turcicum and Botryosphaeria ribis with the EC50 values of 46.77μg·mL-1 and 57.54μg·mL-1 respectively, CompoundⅡ-4 had inhibitory effects on the hvphae growth of Exserohilum turcicum and Botryosphaeria ribis with the EC50 values of 60.50μg·mL-1 and 57.65μg·mL-1 respectively, but compoundⅡ-2 only had inhibitory effects on the hvphae growth of Fusarium oxysporum with the EC50 values of 288.40μg·mL-1.7. The best technological parameters of extraction and isolation of polysaccharides from the rhizomes of Polygonatum odoratum and Polygonatum cirrhifolium were studied for the first time. The optimal technological parameters for refining P. odoratum polysaccharide was summarized as: when the alcohol consistency came up to 95% in the polysaccharide solution (V/V), the polysaccharides precipitated out more than under other alcohol consistency, the time of polysaccharides precipitated was five hours , the pH of sample solution was 7, the times of polysaccharides precipitated was one . The best protein-free technology was: Using enzyme combine with TCA, the dosage of enzyme was 0.8% of polysaccharide solution volume (W/V) and TCA was 4% of polysaccharide solution volume (V/V). LSI296 macroporous anion exchange resin was suitable for the decolorization, the decolorization rate, polysaccharide content and polysaccharide loss rate reached 89.26%, 94.24%, 15.14% respectively. The optimal technology for extracting P. cirrhifolium polysaccharide was summarized as follows: temperature 80℃, solvent: water , time: 2h , times of extraction: 2~3 , rate of solvent to material: 1∶25. The best technological parameters for isolating with precipitating through ethanol is : when the alcohol consistency comes up to 80% in the polysaccharide solution ,the polysaccharide is precipitated out more than under other alcohol consistency. The concentration of sample solution is 1mL·g-1 (material ), pH-value is 6. The best protein -free technology was: Using enzyme combine with TCA and Sevag, the dosage of enzyme was 0.8% of polysaccharide solution volume (W/V) , TCA was 4% of polysaccharide solution volume (V/V) ,Sevag reagent (chloroform : n-butanol is 4: 1 V/V)was 1/5 of polysaccharide fluid volume. LSD208 macroporous anion exchange resin was suitable for the decolorization , the decolorization rate, polysaccharide content and polysaccharide rate reached 88.19%, 92.32%, 64.16% respectively.As a conclusion, the studies on the chemical constituents and bioactive assays of the two Ploygonatum plants elucidated their structure of chemical constituents , screened out new antibacterial compound , established the certain basis to discover precursor compounds and also proved scientific instructs to further utilize the resources of Ploygonatum odoratum and Ploygonatum cirrhifolium .