| Jujube is a dual-purpose plant resources of traditional Chinese medicine and food. The fruit of jujube contains a variety of nutrients, in which the content of saccharides is the highest. As we all know, fructose, glucose and sucrose are the predominant saccharides in jujube. Besides, recent studies has focused on oligosaccharides. First of all, the LC-MS method was used for the preliminarily qualitative analysis of two kinds of higher content oligosaccharides, they are kestose and nistose trihydrate, respectively. Then a Ca-column HPLC method was developed for simultaneous analysis of five main saccharides. And the method was subsequently applied to determine the variations of five saccharides during the growth of two jujube varieties and 32 cultivars from the same growing environment. Finally, combined with the response surface method, the extraction and ethanol precipitation process of jujube oligosaccharides was optimized. The main results were as follows:(1) The value retention time and MS analysis by using HPLC-MS identified the existence of kestose and nistose trihydrate. A method based on HPLC was established for simultaneous determination of 5 saccharides in jujube, such as kestose, nistose trihydrate, fructose, glucose and sucrose. The separation was achieved on a Ca-column and the detection was mesured on the differential refractive index detector. The chromatographic conditions were as follows: column: Hyper Rez XP Carbohydrate Ca2+(300 mm x 7.7 mm, 8 μm); column temperature: 85℃; flow phase: pure water; flow rate: 0.6 m L/min; waters 410 differential refractive detector, detector temperature: 40℃; injection volume: 10 μL; And the external calibration curves were used in quantification. The 5 saccharides in natural air dried and fresh jujube were separated well under these conditions. The relative standard deviations(RSD) of repeated measurement were between 0.53% ~ 0.83%. And the standard addition recovery rate was between 95.0% ? 110.0%. What is more, between the linearity range of 0.30 mg/m L ~ 10.0 mg/m L, the correlation coeffeicent(r) was 0.9996 ? 0.9999. The detection limits were between 0.20 mg/m L ~ 0.25 mg/m L. These 5 saccharides were separated and determined well in 25 min.(2) By this method, we analyzed the content of oligosaccharides in Fuping jujube and Dongzao jujube during the various growing stage and 32 different cultivars under the same growth environment. The results showed that the total content of 5 saccharides of jujube was gradually increased with the growth development, the highest content of five saccharides in Fuping jujube and Dongzao jujube was found in full ripening stage, they were 351 mg/g F.W and 412 mg/g F.W, respectively. The content of 5 saccharides in this period were higher than other different periods. However, the content of oligosaccharides reached the lowest in the ripening stage. The highest content was 28.5 mg/g F.W in Fuping jujube while in white-mature period and 24.4 mg/g F.W in Dongzao jujube while in young fruit stage, respectively. At the same time, we can conclude that there were much differencre on the content of oligosaccharides in jujube in the same period among different cultivars. In terms of oligosaccharides content in full ripening stage, the highest oligosaccharides content was 14.8 mg/g F.W in Fuping jujube. Exploring the relationship among the oligosaccharides, monosaccharides and disaccharides content in jujube. It was found that there was a positive correlation between the oligosaccharides and sucrose content, while there was not very significant correlation between glucose and fructose content.(3) The extraction process of jujube oligosaccharides was optimized through the single factor and response surface method. The optimum extraction conditions was as follows: the extraction temperature was 70℃, the extraction time was 4 h, the ethanol concentration of 70%(V/V), and the ratio of liquid to solid was 25:1. Under these conditions, the oligosaccharides extraction yield of jujube syrup was 1.17%; The optimum ethanol precipitation separation conditions was described as follows: the temperature was 40℃, the ethanol precipitation time was 2 h, the ethanol concentration was 80%(V/V), and the ratio of liquid to solid was 4:1. Under these conditions, the oligosaccharides content can be increased to 5.10% in the precipitation of jujube mixed oligosaccharide syrup. |
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