Extraction And Purification Of Hesperidin In Citrus Processing

Hesperidin, a bioflavonoid, is an abundant by-product from citrus processing wastes with significant pharmacological effect, including its effects on the vascular system, anti-inflammatory, antimicrobial, antioxidant and anti-fertility activity etc. Mandrin peel is a large waste in citrus processing industry in China. We are trying to find a relative more effective ,more economical methods. Our study is about extraction and purification of hesperidin from Citrus Poonensis (one of the most important mandrin in Southern China )peel .The main results were described as follows:1,Hesperidin can be completely separated from other substances by self-made thin-layer chromatography (TLC)on silica gel (including NaOH )and with the elvant of acetidin/methanol/water (10/3/2).With the help of uviol lamp in 283nm, a satisfactory result could be gotten.2,Another method developed for the determination of hesperidin depends on forming its complexes with certain metals. Aluminum nitrate (III)-hesperidin complexes have been formed in water-methanol systems and the spectrophotometric determination of the compound is found to be precise in 283nm.3, The mobile phase consisted of methyl alcohol/water/acetic acid(180:250:18,v/v/v)and was filtered through a 0.45μm nylon filter and degassed by ultrasonic treatment for 30min before use.The HPLC system was operated isocratically at a flow rate of 1ml/min at room temperature and detector was set at 283nm. The precision was good.4,Extraction flavanoids using organic solvent is very common .We select methanol(AR grade) as the extracting liquid, under 80℃, and extract 4h each to extract hesperidin. After extraction, make sure that no hesperidin was left.Then use methanol(HPLC grade) to dissolve..Effect of maturity and grounding level upon the extract yield of hesperidin were studied . Variance analysis showed that siganificant difference was occurant inCitrus Poonensis picked from Oct.31 to Dec.l(sig=0.020 by LSD analysis) .Variance analysis also showed that grounding had no significant difference upon yield between 0.45~2mm,but significant from 2-4mm, It suggests that the extract yield decreases as maturity and the best grounding is smaller than 2mm.5^ Ethanol is more favor to our environment .We select Eethanol(AR grade) as the extracting liquor, under 80 °C, and extract 6h each to extract hesperidin in Citrus Poonensis peel. After extraction, use methanol(HPLC grade) to dissolve.lt suggests that the optimum procedure condition is: adding 25ml ethanol per lg hesperidin ,in 80°C for 6 hours, extraction efficiency can reach 78 %? the yield of dry extracts is about 5 %.6n 2g of crude hesperidin was stirred mechanically with water over a period of 45min,while increments of calcium hydroxide were added to maintain the pH at approximately 11.5.Filting, adding acid to maintain the pH at approximately 3 and heated to 65 °C .It was allowed to stand ovemight.The crude hesperidin was filtered,dried,weighed and the purification can reach 97.46% <>7^ High-speed countercurrent chromatography (HSCCC) was used for the prefractionation of flavonoids .The two-phase solvent system consisted of chloroform/methanol/water(volume ratio is 4:3:2) was the best one. The upper phase was mobile phase and the lower phase was stationary phase ,flow rate:2. 0 ml/min, revolution speed: 800 r/min0 After purification,the purity can reach 98%.In a word, using ethanol is an efficient technique to extract hespridin from Citrus Poonensis peel. Recrystallization,alkali/acid and HSCCC are also good ways to purify hespridin.