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黑莓清汁的制备
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摘要
黑莓是新兴的浆果类水果,色泽艳丽,风味醇美,具有丰富的营养价值,被誉为“紫色生命果”。目前我国已经成为亚洲最大的黑莓种植产地,2006年总产量超过了10,000吨。由于黑莓收获期短,容易腐烂,除少部分直接鲜食外,极大部分黑莓都用于加工。黑莓清汁气味浓郁,可用于各种食品与饮料中,具有广阔的发展前景。但是现有的黑莓清汁产品很容易变色和产生沉淀,产品质量差,严重影响了黑莓清汁在市场上的推广。因此,深入系统地进行黑莓清汁的加工技术和黑莓清汁品质的理论研究是十分必要的,具有重要的现实和理论意义。
     本文研究了酶法制备黑莓汁、羧甲基壳聚糖澄清黑莓汁以及黑莓清汁的贮存稳定性,主要研究内容如下:
     研究了蒸汽热烫工艺对引起黑莓酶促褐变的多酚氧化酶(PPO)的钝化作用。结果发现黑莓中的多酚氧化酶是一种不耐热的酶,采用蒸汽热烫2 min,黑莓中心温度达到52℃时,就可以使多酚氧化酶完全失活,从而减轻黑莓汁的褐变。热烫灭酶还促进了花色苷和多酚类物质的溶出,提高了黑莓汁的营养价值。
     通过建立模拟体系探讨了黑莓PPO降解花色苷的机制。在花色苷和PPO的模拟体系中,黑莓中的PPO可以直接催化降解黑莓花色苷,但是黑莓花色苷并不是PPO的良好作用底物,只有当其它多酚类物质在PPO的作用下生成醌类物质时,醌类化合物与花色苷进一步发生反应,使黑莓花色苷的降解速度明显加快。
     通过研究不同果胶酶制剂对黑莓汁品质的影响,发现不同的商业果胶酶制剂中含有的果胶酶、纤维素酶及花色苷酶的酶活各不相同,因而制得的黑莓汁的澄清度、花色苷含量和总酚含量也各不相同。在黑莓果浆中添加具有复合酶活性(果胶酶活和纤维素酶活)的果胶酶制剂Klerzyme150制备的黑莓汁具有较高的澄清度和较高的花色苷含量。通过均匀设计实验优化得到酶法制备黑莓汁的最佳工艺条件为:Klerzyme 150添加量0.063%(v/w),44℃酶解反应110 min。
     将壳聚糖进行羧甲基改性,得到具有不同取代度的羧甲基壳聚糖,通过红外光谱、热重分析、X射线衍射、核磁共振等多种仪器手段,证实了改性壳聚糖为N,O-羧甲基壳聚糖。比较了不同取代度的羧甲基壳聚糖对黑莓汁澄清度的影响,实验发现,取代度为0.86的羧甲基壳聚糖澄清黑莓汁的效果最好,其添加量为0.02%(w/w)时,室温下静置30min,澄清后的黑莓清汁浊度达到0NTU,4℃贮存6个月后无二次沉淀,而壳聚糖澄清的黑莓清汁的浊度不能达到0NTU,贮存160天产生二次沉淀。
     分别用壳聚糖和取代度为0.86的羧甲基壳聚糖澄清黑莓汁,对澄清前后黑莓汁中的化学成分变化和色泽的研究结果表明,壳聚糖和羧甲基壳聚糖澄清后黑莓清汁的可溶性固形物含量都没有显著变化,但澄清后的黑莓清汁的褐变减少了,黑莓清汁中的蛋白质含量比澄清前分别降低了20.5%和8.2%,总酚含量分别降低了11.64%和9.32%。因此羧甲基壳聚糖澄清黑莓汁的效果优于壳聚糖。
     通过红外光谱、热重、X射线衍射和Zeta电位仪等多种仪器手段,探讨了羧甲基壳聚糖澄清黑莓汁的机理。结果表明,由于羧甲基壳聚糖分子链含有-COOH、-NH2、-OH等基团,它们能够通过氢键吸附黑莓汁中的蛋白质、多酚类物质形成小的絮凝体,然后羧甲基壳聚糖的分子链在这些小的絮凝体之间通过架桥形成大絮凝体而起到澄清作用。由于羧甲基壳聚糖分子中具有较高的正电荷密度,依靠分子内正电荷的相互排斥作用,羧甲基壳聚糖分子内的主链得到最大限度的伸展,从而大大增强了吸附架桥能力。
     研究了黑莓清汁在不同温度贮存时花色苷的变化。黑莓清汁中花色苷在贮存中很容易发生降解,降解规律符合一级动力学。贮存期间,花色苷的降解及其与多酚的聚合反应,使得黑莓清汁色度下降,聚合色上升,褐变增加。低温有助于黑莓清汁中花色苷的保留和色泽稳定性。
     通过对贮存6个月的黑莓清汁二次沉淀的主要成分分析,初步探讨了引起黑莓清汁产生二次沉淀的主要因素。黑莓清汁二次沉淀物中含有15.67%的蛋白质、13.19%的总糖、50.39%的多酚和1.73%的灰分。黑莓清汁混浊物的主要成分是单宁或单宁-蛋白质。对黑莓清汁二次沉淀进行固体核磁共振分析,结果表明其中含有缩合单宁,它是由花色苷与儿茶素或者表儿茶素发生缩合反应形成的,缩合单宁进一步聚合以及与蛋白质相互作用生成了二次沉淀。
     通过HPLC-DAD-ESI-MS检测了黑莓中的多酚类物质。黑莓中的花色苷都是矢车菊类花色苷,以矢车菊3-O-葡萄糖苷为主,占到总花色苷的76.92%,其它花色苷包括矢车菊3-O-阿拉伯糖苷(3.29%)、矢车菊3-O-丙二酸酰葡萄糖花色苷(2.86%)和矢车菊3-O-草酸酐酰葡萄糖花色苷(14.46%)。黑莓果肉中的黄酮醇类多酚含量比种子高,黑莓种子中的苯甲酸类、黄烷醇类(儿茶素、表儿茶素)多酚占有较高的比例。由于在贮存期间,黑莓清汁中的儿茶素、表儿茶素等黄烷醇类多酚同花色苷发生缩合反应生成二次沉淀,因此在酶法制备黑莓清汁时除去黑莓种子再进行酶解、澄清,以减少黑莓清汁中的儿茶素和表儿茶素含量,可以防止和减轻黑莓清汁中二次沉淀的产生。
     对黑莓浓缩汁的加工工艺及其流变性质进行了研究。静态流变性质测定结果表明,黑莓浓缩汁是牛顿型流体。温度和可溶性固形物对黑莓浓缩汁粘度的影响可以用一定的数学方程来描述。
     分析了加工对黑莓清汁和黑莓浓缩汁挥发性成分的影响。经SPME富集,GC-MS分离与检测,在黑莓清汁中检测出65种挥发性化合物,其中醇类和醛类物质是主要的挥发性组分。从黑莓浓缩汁中鉴定出47种化合物,大部分来源于黑莓清汁,主要的风味物质是醛类化合物和醇类化合物。热加工引起的脂肪水解、脂肪酸氧化、Maillard反应,使黑莓清汁中醇类化合物、酸类化合物及羰基化合物含量增加。真空浓缩使黑莓浓缩汁中的挥发性风味物质的种类减少。
Blackberry(Rubus fruticosus L.) is a burgeoning berry fruit, which is highly flavored and colored and is called“purple life fruit”for its abundant nutrition. The yield of blackberry in China reached 10,000 tons in 2006, and China has become the first largest producer of blackberry in Asia. Blackberry is primarily a table fruit, but most of blackberries are processed into many products due to its badly storage performance. Clear blackberry juice, with highly acceptable aroma, can be used in various foods and drinks and can be well developed. However, clear blackberry juice has not been commercially successful due to its excessive haze and sediment formation with color loss. In order to improve the qualities of clear blackberry juice, it is essential to study systemically on the processing and qualities of clear juices.
     The objective of this dissertation is to develop an enzymatic technology for processing blackberry juice, prepare carboxymethyle chitosan to clarify blackberry juice and investigate the stability of clear blackberry juice. The main results are as follows:
     The effect of steam blanching on the polyphenol oxidase (PPO) was studied and the results showed that PPO from blackberry pulp had a relatively low thermal stability. The optimum parameters of steam blanching process were confirmed with blanching for 2 min to inactivate PPO when the central temperature of blackberry was 52℃. Moreover, steam blanching could not only inactivate PPO, but also induce a significant recovery increase of anthocyanins and polyphenols, in relation to the higher quality of blackberry juice.
     The mechanism for the degradation of anthocyanins by PPO was investigated by establishing a model system. The loss of the colour in this model system containing PPO and blackberry anthocyanins indicated that PPO could act directly on these pigments. However they remain fairly poor substrates of PPO. The rate of anthocyanin degradation was stimulated by the addition of non anthocyanin polyphenols. Anthocyanins were oxidised via the enzymatically generated o-quinone by coupled oxidation mechanism.
     The effects of different commercial pectinolytic enzyme preparations on blackberry juice quality were determined. The enzyme preparations affected the clarity and contents of anthocyanins and polyphenols in the juice due to different activity profiles of these enzyme preparations. The commercial pectinase named Klerzyme 150, which contained almost no anthocyaninase activity, had a desirable effect on the yield and quality of blackberry juice. Studies on the uniform design for Klerzyme 150 enzyme revealed that the optimum conditions were 0.063 % (v/w), 44 oC and 110 min for enzyme dosage, reaction temperature and reaction time, respectively.
     Chitoan was modified to carboxymethyl chitosan, which had different degree of substitution. The chemical structure was characterized by the combination investigation of FTIR, XRD, TG and NMR and the compound were identified as N,O-carboxymethyl chitosan. Carboxymethyl chitosan was used to clarify the blackberry juice and the results showed that the clarity of blackberry juice was related to the degree of substitution. When the degree of substitution was 0.86, carboxymethyl chitosan could clarify blackberry juice greatly. The characteristics of clear blackberry juices clarified with chitosan and carboxymethyl chitosan were compared. It indicated that the juice clarified with carboxymethyl chitosan had much less potein and polyphenols.
     Clarification mechanism of blackberry juice by carboxymethyl chitosan was investigated. There existed interaction of hydrogen bond between -COOH、-NH2、-OH of carboxymethyl chitosan and blackberry protein and blackberry polyphenol, which was confirmed by FTIR, XRD, TG and Zeta potential. Carboxymethyl chitosan could absorb protein and polyphenol in blackberry to produce primary ?ocs and then bridge among these primary ?ocs to form bigger flocs and thus clarify blackberry juice.
     The results of storage tests at different storage temperature indicated that anthocyanins were prone to degradation. It was found that the thermal degradation of anthocyanins followed first-order reaction kinetics. During storage, the browning index and polymeric color increased, while the color density decreased. Storage at low temperature was good for the color stability of blackberry juices.
     According to the analysis of the main components of sediment of clear blackberry juice after 6 months storage, the main aspects influencing sedimentation of blackberry juice were discussed. There were 50.39% of polyphnols, 15.67% of protein, 13.19% of carbohydrates, and 1.73% of ash. The haze of clear blackberry juice was mainly the type of tannin and protein-tannin haze. The second sediment contained condensed tannin when analyzed the composition with NMR. Anthocyanin and catechin could condense to produce tannin, which could increase degree of polymer and/or interact with protein to produce sediment.
     The polyphenols in blackberry fruit were isolated and characterized by high performance liquid chromatography (HPLC) with UV-visible detection, electrospray ionization mass spectroscopy (ESI-MS). Four defined anthocyanin pigments were detected, all of them containing cyanidin (Cy) as aglycone. Cy 3-O-glucoside was the major pigment constituting 76.92% of the anthocyanins in blackberry fruit followed by Cy 3-O-dioxalylglucose (14.46%)、Cy 3-arabinoside (3.29%)、and Cy 3-malonylglucos(e2.86%). Flavonols (primarily quercetin glycosides) were the major flavonols in berries pulp, whereas procyanidins (catechin and epicatechin) and benzoic acid predominated in seeds.
     Clear blackberry juice concentrates and their rheological properties were studied. The concentrates were predominantly Newtonian over the range of temperature and solids content studied. The effect of temperature and solids content on the viscosity of clear blackberry juice concentrates could be characterized by a mathematical model.
     The volatile compounds of blackberry juice during processing were concentrated by SPME, separated and identified by GC-MS. There were 65 kinds of compounds in volatile fraction of blackberry juice, and the main components were alcohols and aldehydes. The kinds of volatile compounds were decreased by 27% and 2-Hexenal became the most compounds in blackberry juice concentrates. Because of fat hydrolysis, fatty acids oxidation and Maillard reaction, furan and some alcohols produced during juice processing were also detected in original and blackberry juice concentrates.
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