Research On The Extraction,Stability And Antioxidant Activity Of Anthocyanins From Lycium Ruthenicum

Lycium ruthenicum Murr.?Black goji berries?,is a wild plant endemic to northwestern China,dried fruit of which is often used as Traditional Chinese Medicine.As an edible and medical plant,it is also often used to make tea,wine and other daily food.In recent years,many studies have reported that the fruits of L.ruthenicum showed various biological activities,such as anti-fatigue,anti-diabetes and lowering blood lipids.The fruits of L.ruthenicum are known as a rich source of anthocyanins,a series of natural water-soluble pigments with many biological activities,which could be an ideal resource for functional natural pigments.However,there are few studies for anthocyanins in L.ruthenicum.In order to further develop and utilize the L.ruthenicum,it is necessary to study the anthocyanins in it.This paper mainly includes the following four parts:?1?We reviewed the present status of research on L.ruthenicum at home and abroad in recent years,which provided a theoretical basis for the determination of the research contents and experimental design of this study.?2?In order to improve the extraction efficiency of anthocyanins from L.ruthenicum and reduce the loss of total anthocyanins in the extraction process,the ultrasonic-assisted extraction method was used in this study.Firstly,we investigated the influence of ethanol volume fraction,liquid-solid ratio,extraction time and temperature on the anthocyanin extraction by the single factor experiment,and then the RSM?response surface method?is used to design and predict the best extraction conditions of anthocyanins.Evently,we got the best extraction conditions.In order to facilitate practical operation,a slight modification is made:65%acid ethanol is used as extraction solvent,the ratio of liquid to material is 40:1,the extraction time is 50 min,and the temperature is 55?C.According to the conditions,we made experiment and got anthocyanin extraction yield of 3.603 mg/g,and it is close to predictive value?3.636mg/g?of the RSM,which shows extraction conditions of anthocyanin optimized by RSM is accurate and reliable.?3?DPPH free radical scavenging experiment,ABTS free radical scavenging experiment,and ferric reducing ability of plasma determination were used to evaluate the antioxidant activity of anthocyanins extract of L.ruthenicum.Synthetic antioxidant BHT was used as positive control,and the results showed that the anthocyanins in L.ruthenicum with much stronger DPPH free radical scavenging ability than BHT,and similar antioxidant ability with BHT in the ABTS ultra oxygen anion removal experiment and iron used in the determination of reducing power.Accordingly,the anthocyanins in L.ruthenicum had good oxidation resistance,which may help to capture and counteract free radicals in the human body,so as to reduce the free radicals oxidative damage to the human body.?4?A rapid and efficient DPPH–HPLC–DAD–MS/MS method was used to screen antioxidant active anthocyanins in L.ruthenicum.According to the reduction ratio of their chromatographic peak area,we could judge the strength of their antioxidant activity.In addition,the structures of anthocyanins in L.ruthenicum were determined by Mass spectrometry as petunidin-3-O-rutinoside?glucosyl-p-coumaroyl?-5-O-glucoside and petunidin-3-O-rutinoside?p-coumaroyl?-5-O-glucoside.It is shown that the peak area of anthocyanins is reduced after adding DPPH,indicating that they both have antioxidant activity.Also,the decrement of their peak area ratio have little difference,suggesting their antioxidant capacity are similar,due to the same anthocyanidin?both are petunidin?of the two anthocyanin,and the similar combination of sugar?only a glucosyl is different?.?5?Finally,we studied the stability of anthocyanin in L.ruthenicum,and investigated the influence of different factors on it.The results showed that the pH value had a great influence on the stability of anthocyanin,which is stable only under acidic conditions,whereas anthocyanins would be degraded or converted into other compounds in neutral or alkaline conditions.Temperature could also affect the stability of anthocyanin,but the impact is relatively small.At room temperature 25?,its degradation half-life could be 47 days and at-4?it can stay stable for a long time.The stability of anthocyanins was also influenced by strong light irradiation,whereas the weak light irradiation and UV lights would not obviously affect the stability of anthocyanin.Most of the common metal ions,such as Na⁺,Ca²⁺,Mg²⁺,Ba²⁺,Mn²⁺,Cu²⁺,Pb²⁺,Al³⁺,have no obvious effect on he stability of anthocyanin.But the presence of Fe²⁺and Fe³⁺could affect the stability of anthocyanin and makes the color lighter.In combination,the anthocyanins in L.ruthenicum have a certain tolerance to temperature,light,general metal ions,but easily affected by the pH value.Therefore,dark environment,low temperature,low pH value and avoiding the use of iron containers as the storage condition for anthocyanins were suggested to maintain their stability for a long time.