| Lithocarpus polystachyus(Wall.)Rehd.is a medicinal and edible plant which can be used as drug,tea and sugar.Folk often take the leaves in the boiling water,and then the color of the water will be golden brown.It tastes sweet and slightly bitter lasting.Ancients used to clear the heat and assuage the thirst.Lithocarpus polystachyus(Wall.)Rehd.contains up to 10%of non-sugar sweetener,which is high-intensity and low-calorie.Its sweetness composed mainly of phlorizin,3-hydroxy-phlorizin,trilobatin and so on.Modern pharmacological studies show that Lithocarpus polystachyus(Wall.)Rehd.can lower blood sugar,blood fat,blood pressure,and antioxidant,anti-inflammatory,antibacterial and anti-allergy etc.Lithocarpus polystachyus(Wall.)Rehd.show that more active ingredients of Lithocarpus polystachyus(Wall.)Rehd.leaves are mainly flavonoids.Therefore,this study was performed for Lithocarpus on total effective part of the preparation of high purity and HPLC fingerprint of flavonoids.ObjectiveIn this paper,the samples were investigated with the content of total flavonoids,3-hydroxy-phlorizin and phlorizin as the index,combining with the HPLC fingerprint for the effective part of Lithocarpus leaves.The samples were gotten in the different harvest time and different processing methods,which were transplanted or wild.And ultimately,the best harvest of Lithocarpus polystachyus(Wall.)Rehd.leaves was determined.At the same time,high purity multi total flavonoids are gotten by taking advantage of a polyamide and AB-8 macroporous resin preparation technique combined with It will provide a scientific basis for Lithocarpus polystachyus(Wall.)Rehd.herbs harvesting,processing and preparation.Methods1.The factors of fingerprints for the chromatographic were investigated,including the mobile phase gradient,separation of different columns,column temperature,flow rate,detection wavelength and acquisition time.According to the investigation,the best conditions for chromatographic fingerprint was determined,and then methodology study was carried out.Finally,10 different batches of sample was determined under optimum chromatographic conditions.Take the peak of phlorizin as reference to determine the common peak,the HPLC fingerprints for effective parts of Lithocarpus polystachyus(Wall.)Rehd.leaves was established.2.Using ultraviolet spectrophotometry multi spike coca leaf total flavonoid content.With 3-hydroxy-phlorizin and phlorizin content index,high performance liquid chromatography with Kromasil(C18 250mm*4.6mm,5 ?m)column was used with methanol-0.1%formic acid as the mobile phase gradient elution,detection wavelength 285nm,column temperature was room temperature,flow rate 0.8 mL-min-1.Comparison of different harvest time,different processing methods,different cultivation methods Lithocarpus polystachyus(Wall.)Rehd.leaf quality.3.Preparation of total flavonoids in high purity:to investigate the eluent concentration polyamide,polyamide-macroporous resin combined with the eluent concentration,sample volume and the proportion of polyamide,sample volume and macroporous resin Effect of proportion,eluent amount on the total flavonoids purity determine the best purification process and process validation.Results1.The chromatographic conditions on establishing HPLC fingerprint of a more effective part of Lithocarpus polystachyus(Wall.)Rehd.leaves:C18 reverse phase column Kromasil(C18 250mm*4.6mm,5 ?m);methanol-0.1%formic acid gradient elution system;column temperature was 25?;flow rate was 0.8mLˇmin-1;detection wavelength of 254nm;injection volume was 25 uL;acquisition time 140min.2.The contant of 3-hydroxy-phlorizin and phlorizin on Lithocarpus polystachyus(Wall.)Rehd.leaves is different from different harvest time and different processing methods.And fingerprint peaks change significantly.Lithocarpus polystachyus(Wall.)Rehd.leaves have a variety of ingredients,so its pharmacological effects are not the same.In order to determine the harvest period,the higher content of total flavonoids,3-hydroxy-phlorizin and the total peak area of HPLC in October was picked.Different processing methods on Lithocarpus chemical composition and content had little effect.3.Established a process for preparing high purity of total flavonoids of Lithocarpus polystachyus(Wall.)Rehd.,the process conditions:eluent of 60%ethanol,the sample amount(g)and quality polyamide(g)ratio of 0.075:1 to 0.09;1,sample amount(g)and the quality of macroporous resin(g)a ratio of 1:90,the eluent volume 5BV,total flavonoids purity of more than 90%.Conclusion1.The establishment of fingerprints on the effective part of Lithocarpus polystachyus(Wall.)Rehd.leaves suggests the material basis for its effective part,while methodological study showed that it is a good stability of the fingerprint and easy to reproduce.2.The method reflects the overall characteristics and individual characteristics of component parts on Lithocarpus polystachyus(Wall.)Rehd.Leaves,which combines a multi-index components determination with fingerprint evaluation system.It will offer new ideas about better quality control of medicines.3.Using resin combined with the preparation of high purity total flavonoids,the method is simple and practical.It can provide a reference for the late preparation. |
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