Extraction And Separation Of Xanthohumol From Hops And Its Application

Xanthohumol is the most important component of polyphenol from hops, with many physiological activities. And hops are the unique natural source of xanthohumol as so far. The main purpose of hops is brewing except for a small amounts of medicinal, So, drinking beer is the main ways for xanthohumol in diet. However, the content of xanthohumol in beer is only a few parts per million by its poor water solubility. Therefore, it has been a hot topic in hops research that people focus on how to improve the water-solubility of xanthohumol and expand its application. From the spent hops which was the residue of supercritical carbon dioxide extraction hop extract, an accepted commercial process was obtained. The content of polyphenol and xanthohumol in different hop samples was determined, furthermore, the stability of xanthohumol was studied. The inclusion effect of xanthohumol was compared using seven kinds of cyclodextrins and its derivative, meanswhile the inclusion process of xanthohumol and HP-β-CD was optimized and the structure of cyclodextrins inclusion complex was characterized. The synergism antioxidant activity of xanthohumol mixed with three acidulants were investgated and its stability in different pH conditions and six preservatives. The main research contents as follows:In comparison with High Performance Liquid Chromatography(HPLC) method, the analysis methods of xanthohumol were established by Ultraviolet spectrophotometry(UV) based on the methodology of the investigation. Three methods were used to detemined the content of polyphenol in hop samples. The results indicated that the calibration curves of three methods, namely HPLC, direct UV method and UV with chromogenic agent methods, had a good linear relation. The recoveries of the methods were ranged from 95%-102% and the repeatability or stability were all well for them. For higher percentage of xanthohumol, the test results were good correlation of the three methods, but HPLC method is slightly lower. The relative standard deviations were range from 4.03% to 20.67%. Among them, the data of UV with chromogenic agent methods is closed to HPLC methods. The test results of 20 kinds of hop samples by UV methods were far above the values detected by HPLC. It shows that the values does not reflect the content of xanthohumol in hops. However, the result also shows that it was closed to the contents of total polyphenol by Folin-Ciocalteu method, especially for UV with chromogenic agent method. The method is more suitable for the detemination of polyphenol, because it eliminated the interference of other substances and operated easily, low-cost, fast analysis.Xanthohumol was extracted and separated from Marco Polo hops by column elution method. The preparation technology was simplified through qualitative analysis and tracking active ingredrent of elution. After mixed the hop extracts and hops and diatomite, 30% methanol was used to remove impurities. Then 75% crude extract of xanthohumol was obtained from 60% methanol elution. The purity of xanthohumol up to more than 99% after recrystallization with the melting point is 166 ℃ to 167 ℃and melting process is 1 ℃.In order to improve the water-solubility of xanthohumol, The inclusion effect of xanthohumol was compared by grinding using seven kinds of cyclodextrins and its derivative. The results showed that there is a best adduction effect when mixed xanthohumol with HP-β-CD. On the basis of above results, the influences of the host-guest molecular ratio, triturate time, adduction temperature and purity on the adduction effect of xanthohumol-HP-β-CD were evaluated by single factor experiments in grinding methods. And the inclusion process was further optimized by the methods randomcentroid optimization. The results suggested that the inclusion rate of xanthohumol-HP-β-CD clathrate is up to 99% when the host-guest molecular ratio is 50:1, the adduction temprature is 42℃, and the triturate time is 37 min. Xanthohumol-HP-β-CD was analysed and identified by ultraviolet spectrum, biological microscopy, scanning electron microscopy and infrared spectroscopy. The result shows that xanthohumol and HP-β-CD formed inclusion complexand its solubility can be up to 7.5 mg/mL in water. Under this concentration, the content of xanthohumol is 150 μg/m L in the system, and the water-solubility of xanthohumol is significantly improved. The synergism antioxidant activity of xanthohumol mixed with three acidulants, namely citric acid, sodium citrate and vitamin C, were researched by using two different assays, and Synergistic Effect(SE) as evaluation criteria. The result suggested that there is a synergistic antioxidant activity when the concentration of xanthohumol is over 1.5 mg/mL in the β-carotene-linoleate mode system, but the synergism is weaker than DPPH free radical system, which has synergism antioxidant activity when the concentration of xanthohumol is range from 0.5 to 3.0 mg/mL, while the SE had no obvious changed with the increased concentration of xanthohumol. The other study herein was conduced to determine the effects associated with the stability of xanthohumol-HP-β-CD in different preservatives and different pH condition. Firstly, It was found that the stability of xanthohumol-HP-β-CD in six kinds of preservatives is better than the control group below 80 ℃. However, when the temperature mounted into 90 ℃, BHT, vitamin C, glucose and NaCl could reduce the loss of xanthohumol-HP-β-CD, and there was no obvious effect in citric acid and sucrose. Secondly, the stability of xanthohumol-HP-β-CD in preservatives to hight was investigated, the content of xanthohumol decreased rapidly when placed in bright light and UV light, it should be keep away from light. NaCl solution with different concentration could reduce the decrease of xanthohumol-HP-β-CD, especially 10% NaCl solution. The last, the stability of xanthohumol-HP-β-CD in different pH condition was investigated, the result shows that pH conditions has significant influence to the content of xanthohumol, with the extension of time, the content of xanthohumol gradually reduced, at the same time, higher concentration mean, counterintuitively, a slow change of xanthohumol. The weak base environment is the best condition.