| Magnolia kwangsiensis Figlar & Noot. (M. kwangsiensis) is a rare and endanged plant, only locates in Guangxi, China. In order to protect the plant source and utilize them effectively, research on effective components from leaf, flower and fruit of M. kwangsiensis was carried out. Extraction, purification and analysis of volatile compound, fatty acid, polysaccharides and polyphenols were studied. The antitumor activity of polysaccharides and antioxidant activity of polyphenols were investigated. The main research contents and results were showed as follows:(1) Research on volatile compounds from flower of M. kwangsiensisThe volatile oils from fresh flowers of M. kwangsiensis were obtained using hydrodistillation, and analyzed by gas chromatography-mass spectrometry technology (GC-MS). The yields of volatile oils extracted from male flower and female flower were the same, both were 0.13%. Monoterpene hydrocarbons were rich in flower oils, which occupied 48.19% of male flower oil and 54.03% of female flower oil. The major compositions of volatile oil in male flower were a-limonene 18.45%, a-terpinen 12.96%, a-cadinol 12.20%, τ-muurolol 9.85%, β-ocimene 8.14%,δ-cadinene 7.18% and β-myrcene 6.35%. The major compositions of volatile oil in female flower were a-limonene 20.79%, a-cadinol 11.48%, β-ocimene 9.45%,δ-cadinene 8.54%, τ-muurolol 8.40%, a-terpinen 7.06% and p-myrcene 6.57%.The volatile compounds from fresh male flower and female flower were collected by headspace injection and analyzed by GC-MS. The constituents in male flower were β-ocimene 28.12%, a-terpinylacetate 21.35%, a-terpinen 16.36%, P-myrcene 9.87%, p-menth-1-ene 6.95% and a-phellandrene 1.26%. The constituents in female flower were a-terpinylacetate 25.88%, a-terpinen 22.40%, β-myrcene 16.38%, P-ocimene 10.73%, p-menth-1-ene 10.20% and a-phellandrene 5.09%.The constituents and contents of volatile components in male flower and female flower have certain difference, which affect the performance of the frangrance.(2) Research on volatile oil from leaf of M. kwangsiensisThe volatile oil from fresh leaf of M. kwangsiensis was extracted using hydrodistillation, and analyzed by GC-MS. The yield of volatile oil was 0.19%. Monoterpene hydrocarbons predominated, which occupied 44.62% of the fresh leaf oil. The main components were P-terpineol 28.85%, y-terpinene 18.14%, β-myrcene 15.90%, a-terpineol 5.35%, a-limonene 4.74%, a-eudesmol 4.42%, and p-menth-1-ene 2.78%.The leaves were oven-dried and air-dried, from which the volatile oils were obtained. The yield of the oven-dried leaf oil was 0.31%, the main compositions were β-terpinylacetate 27.45%, y-terpinene 15.28%, (3-myrcene 14.56%, a-eudesmol 6.15%, a-terpineol 4.91%, a-limonene 4.79%。The yield of the oil in air-dried leaf was 0.36%, the main compositions were β-terpineol 20.23%, β-ocimene 11.29%, β-myrcene 8.65%, a-limonene 7.54%, nerolidol 4.66%, a-terpineol 4.37%。The significant difference was seen on the constituents and contents of volatile oils from fresh leaf, oven-dried leaf and air-dried leaf, declaring the biotransformation was run in the process of air-dring.(3) Research on effective components from fruit of M. kwangsiensisThe volatile oils were obtained using hydrodistillation from fresh peel and aril of M. kwangsiensis fruit, the yields were 0.10% and 4.91%, respectively. The chemical constituents were analyzed by GC-MS. In the peel oil,22 compounds were identified, the major compounds were β-ocimene 30.80%, p-menth-1-ene 17.76%, a-terpinene 10.15%, β-myrcene 7.03% and a-terpineol 5.18%. In the aril oil,23 compounds were identified, major constituents were β-ocimene 56.03%, P-phellandrene 10.96%, a-terpinene 6.37%, a-phellandrene 6.16% and β-myrcene 6.04%.The volatile compounds from fresh peel and aril were extracted by headspace injection and analyzed by GC-MS. Five compounds were identified in peel, which were β-ocimene 35.96%, a-terpinene 24.99%, a-limonene 15.76%, β-myrcene 10.29% and a-phellandrene 2.96%. Seven compounds were identified in aril, which were β-ocimene 69.16%, a-terpinene 7.49%, β-myrcene 6.66%, a-phellandrene 6.17%, a-limonene 5.45%, p-menth-1-ene 3.30% and β-thujene 1.74%.The lipids were extracted by ultrasound-assisted extraction from the aril and kernel of M. kwangsiensis fruit, the yields were 33.00% and 54.05, respectively. The fatty acids of lipids were esterified and identified through GC-MS. The main fatty acids in aril were 9,12-octadecadienoic acid 49.52%, hexadecanoic acid 32.27%, octadecenoic acid 10.24% and octadecanoic acid 4.49%. While the main fatty acids in kernel were 9,12-octadecadienoic acid 56.91%, hexadecanoic acid 25.27%, octadecenoic acid 12.09% and octadecanoic acid 5.63%. The content of unsaturated fatty acids was up to 62.22% in aril, and that was 69.03% in kernel. Three odd carbon fatty acids, which were unusual in plant, were found in M. kwangsiensis aril.(4) Research on polysaccharides from leaf of M. kwangsiensisThe polysaccharides were extracted from leaf of M. kwangsiensis through hot water reflux and microwave assisted extraction, and determined by the phenol-sulfuric acid method. The yield of hot water reflux extraction was 5.09%, the yield of microwave assisted extraction was 2.48%, proving that hot water reflux extraction was more suitable method for extraction of M. kwangsiensis leaf polysaccharides. Two polysaccharide fractions (P-2 and P-3) were isolated by polyamide resin, DEAE-52 cellulose chromatography and Sephadex G-100 column chromatography in order, respectively. The polysaccharides contents of P-2 and P-3 were 96.12% and 95.07%, respectively. P-2 and P-3 were characterized by infrared spectroscopy (IR), high performance gel permeation chromatography (HPGPC) and GC-MS. P-2 was comprised of only glucose. Its molecular weight was about 11.2×103 Da and the formula might be C414H6900345. P-3 was comprised of xylose and rhamnose in the ratio of 1:4. Its molecular weight was about 7.8×103 Da, and the formula might be C319H528O220.The antitumor activities of P-2 and P-3 on the growth of human hepatocellular carcinoma (BEL-7404) cells, human lung cancer (A549) cells and human gastric carcinoma (SGC7901) cells in vitro were evaluated. P-2 and P-3 showed no inhibition on BEL-7404 cells. P-2 had some inhibition on A549 ells and SGC7901 cells, P-3 exhibited marked antitumor activities on A549cells and SGC7901 cells with IC50 value of 8.48 and 5.66μg·mL-1. The results indicted P-3 could be potentially useful as a natural and safe health food or antitumor agent.(5) Research on polyphenols from leaf of M. kwangsiensisThe analysis method of polyphenols was established with high precision and good stability. The microwave assisted extraction conditions were optimized by response surface methodology, the factors of affecting the yield of M. kwangsiensis leaf polyphenols in order were ratio of solvent to material> acetone concentration> extraction time. The yield was 2.44% under the optimal conditions:acetone concentration of 70%, ratio of solvent to material of 21 mL·g-1 and extraction time of 16 min. The ethyl acetate phase, water phase and NKA-9 purification were obtained through solvent extraction and macroporous resin purification, whose polyphenols contents were 32.11%,21.89% and 39.33%, respectively.The antioxidant activities of M. kwangsiensis leaf polyphenols were evaluated in terms of total antioxidant activity, reducing power, DPPH-scavenging activity,-OH scavenging activity and antilipid peroxidant activity. Results showed the total antioxidant activities were presented as:NKA-9 purification> crude extract> water phase> ethyl acetate phase, and reducing power decreased in the order of NKA-9 purification> water phase> ethyl acetate extract> crude extract. IC50 values of crude extract, ethyl acetate extract, water phase and NKA-9 purification on DPPH-scavenging effect were 0.099,0.067,0.135 and 0.021 mg·mL-1, respectively. When the concentration was 0.50 mg·mL-1, the scavenging rates on-OH of crude extract, ethyl acetate extract, water phase and NKA-9 purification were equivalent to 1.6,1.4,1.8 and 2.5 times of Vc, that of Vc was 26.90%. NKA-9 purification showed best antilipid peroxidant activity, with the inhibition rate of 71.17%; crude extract resulted worst antilipid peroxidant activity, with inhibition rate of 38.98%. Those results indicted that the M. kwangsiensis leaf polyphenols presented potential antioxidant activities.(6) It was revealed that M. kwangsiensis has abundant functional compositions by means of analysis of volatile compound, fatty acid, polysaccharides, polyphenols and abioactivity of polysaccharides and polyphenols. This study has great impotance to promoting large-scale cultivation and developing new natural product with biological activity from M. kwangsiensis. |
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