CN1116419C - Enzymolysis process for preparing functional oligoxylose - Google Patents

Enzymolysis process for preparing functional oligoxylose Download PDF

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CN1116419C
CN1116419C CN 99126547 CN99126547A CN1116419C CN 1116419 C CN1116419 C CN 1116419C CN 99126547 CN99126547 CN 99126547 CN 99126547 A CN99126547 A CN 99126547A CN 1116419 C CN1116419 C CN 1116419C
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enzyme
xos
functional
preparation
water
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CN1266633A (en )
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蔡敬民
吴克
潘仁瑞
张洁
刘斌
杨本宏
居萌
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合肥联合大学
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本发明公开一种酶法制备可作功能性食品及食品添加剂的低聚木糖的方法,它包括以下步骤:用玉米芯、玉米秸杆、稻草等作为原料经筛选、干燥、粉碎、过筛制得原料,上述原料用水浸湿、脱水,用碱在室温及高温时搅拌浸提,收集滤液,用酸中和至微酸性,得半纤维素溶液中加入酶粉或酶制剂,酶的水解液再经脱色、离子交换、浓缩后得低聚二木糖为主的混合物或其结晶,功能性低聚木糖在食品、饲料、保健以及植物保护方面都有广阔的应用前景。 The present invention discloses a method for enzymatic preparation Xylo functional foods and food additives can be used, comprising the steps of: corn cobs, corn stover, rice straw as a raw material by screening, drying, crushing, sieving preparing a material, the raw material soaked with water, dehydrated, alkali leaching was stirred at room temperature and high temperature, the filtrate was collected, and neutralized with an acid to slightly acidic, or enzyme powder obtained hemicellulase enzyme preparation was added, enzymatic hydrolysis of then decolored liquid, ion exchange, and concentrated to give a mixture of two oligomeric xylose-based or a crystalline, XOS functional food, feed, health and plant protection has broad application prospects.

Description

酶法制备功能性低聚木糖的生产工艺 Enzymatic Preparation of functional XOS production process

技术领域 FIELD

本发明涉及一种功能性低聚糖的制备方法,具体讲是酶法制备功能性低聚木糖的生产工艺。 The present invention relates to a method for preparing a functional oligosaccharide, specifically prepared enzymatically functional XOS production process.

然而当前最受人注意的还是低聚木糖的双歧因子作用。 Bifidus factor effect, however most people pay attention to current or Xylo. 众所周知,双歧乳杆菌是一种人体肠道正常菌,婴儿出生三四日即出现双歧杆菌,并且占绝对优势。 As we all know, Lactobacillus bifidus is a normal human intestinal bacteria, the baby was born three 24 appears bifidobacteria, and dominant. 随着年龄增大双歧杆菌逐渐减少,而产气荚膜梭杆菌、大肠杆菌等腐败细菌大量增加。 With the increasing age of bifidobacteria gradually reduced, and Clostridium perfringens, E. coli and other bacteria substantial increase in corruption. 到了老年,肠道充斥腐败细菌,双歧杆菌几乎消失。 In old age, full of intestinal bacteria of corruption, Bifidobacterium almost disappeared. 腐败细菌在肠道中分解食物成分,产生氨气、胺类、硫化氢、粪臭素、吲哚、酚类以及亚硝胺等有毒变异源,人体长期吸收这些毒素,会促进衰老,诱发癌症,引起动脉硬化、肝脏障碍等疾病。 Spoilage bacteria in the gut break down food ingredients, toxic sources of variation ammonia, amines, hydrogen sulfide, skatole, indole, phenol and nitrosamines, long body absorb these toxins, promote aging, cause cancer, cause arteriosclerosis disease, liver disorders. 而双歧杆菌则对保卫人体健康起着重要的作用:①抑制肠道有害微生物之繁殖,分解碳水化合物产生有机酸,维持肠道菌群平衡,促进肠道蠕动,有利于防止便秘:②不腐败蛋白质,不生成有害物质,促进蛋白质消化吸收;③合成B族维生素,并促进对某些无机盐利用;④产生某些物质可增强提高机体的免疫力;⑤分解致癌物质,预防和抑制肿瘤的发生,还可防止体内胆固醇的积蓄。 The bifidobacteria is plays an important role in the defense of human health: ① inhibit the growth of harmful microorganisms in the gut break down carbohydrates produce organic acids, maintain intestinal flora balance, promote intestinal peristalsis, help prevent constipation: ② not corruption protein, does not generate harmful substances, promote protein digestion and absorption; ③ synthesis of B vitamins, and to facilitate the use of certain inorganic salts; ④ enhance the production of certain substances can improve the body's immune system; ⑤ decomposition carcinogens, prevention and suppression of tumors It occurs, but also to prevent the accumulation of cholesterol.

为实现上述目的,本发明采用的技术方案:一种酶法制备功能性低聚木糖的生产工艺,它包括以下步骤:a.将玉米芯、秸杆、甘蔗渣、麦秸粉、稻草等进行干燥、粉碎、过筛制得原料;b.上述原料用清水浸湿、脱水,再用碱室温下搅拌浸提15~30小时,70~90℃时搅拌浸提1~3小时,去除固形物,收集滤液,并用酸缓冲液中和至微酸性,得到半纤维素溶液;c.在上述半纤维素溶液中加入酶粉或纯化后的酶制剂水解半纤维素,酶的水解液再经脱色、离子交换、浓缩等过程得低聚木二糖为主的混合物。 To achieve the above object, the present invention adopts the technical scheme: preparing a functional enzyme XOS production process, comprising the steps of:. A cobs, straw, bagasse, wheat straw, rice straw, etc. dried, pulverized, sieved to prepare a material;. b of the raw material soaked with water, dehydrated, and then stirred at room temperature alkali leaching 15 to 30 hours, stirring for 1 to 3 hours leaching time 70 ~ 90 ℃, solids removed the filtrate was collected, and washed with an acid buffer and to slightly acidic, hemicellulose solution was obtained;. c hemicellulose hydrolyzing enzyme preparation is added after the above purified enzyme powder or solution, hemicellulose, enzymatic hydrolyzate then decolored , ion exchange, concentration and the like processes to give oligomerization xylobiose based mixtures.

d.上述的酶制剂主要是木聚糖酶(EC3.2.1.8),它可以由微生物分别通过固态或液态发酵方法制得,经过发酵产生的木聚糖酶可以制备成部分纯化的酶制剂或经sephadex G-100和DEAE-sephadex-A50纯化后的纯酶。 d. above enzyme preparations are mainly xylanase (EC 3.2.1.8), which can be solid or liquid, respectively, through fermentation process by microorganisms obtained after fermentation of the xylanase can be prepared as part of a purified enzyme preparation of or with sephadex G-100, and after DEAE-sephadex-A50 pure enzyme purification.

上述生产工艺也可用如图1所示的流程表示;用玉米芯、玉米秸杆、稻草等作为原料经筛选、干燥、粉碎、过筛制得原料,上述原料用水浸湿、脱水,用碱在室温及高温下搅拌浸提,去除固形物,收集滤液,用酸中和至微酸性,得半纤维素溶液,在制得的溶液中加入酶粉或酶制剂水解半纤维素,酶的水解液再经脱色、离子交换、浓缩后得低聚二木糖为主的混合物。 The aforementioned production process can also be represented by the flowchart shown in FIG. 1; corn cobs, corn stover, rice straw as a raw material by screening, drying, crushing, and sieved to obtain a raw material, the raw material soaked in water, dehydrated with a base stirred at room temperature and high temperature extraction, removal of solids, the filtrate was collected, and neutralized with an acid to slightly acidic, hemicellulose solution obtained, was added hydrolase enzyme powder or formulations of hemicellulose in the resulting solution, enzymatic hydrolyzate then by bleaching, ion exchange, to give two oligomeric xylose based mixture was concentrated.

在健康人体消化道中的细菌有100多种,其数量达1000亿以上。 There are more than 100 kinds of bacteria in the digestive tract of healthy human body, their number reached 100 billion or more. 其中双歧杆菌在肠道中占有优势,检查排便中双歧杆菌多少,便可判断人体健康如何。 Which bifidobacteria in the gut possession advantage, the number of bifidobacteria in stool examination, can determine how human health. 长寿老人肠道中菌群结构接近青年人,这已为国内外学者所证实。 Long-lived people in the intestinal flora structure close to the young people, which has been confirmed by scholars. 因此若能调整肠道菌群结构,使双歧杆菌处于优势,无疑对人的健康和抗衰老是有益的,食用双歧因子是一种有力的辅助手段,低聚木糖已证明具有良好的双歧因子功能。 Therefore, if we adjust the structure of the intestinal flora, the bifidobacteria in a dominant position, no doubt to human health and anti always useful, edible bifidus factor is a powerful adjunct to, XOS has proved to have good bifidus factor function. 目前,国内还未检索到低聚木糖的同类产品的专利。 At present, China has not been retrieved to similar products patented Xylo. 本发明中的酶制剂由黑曲酶生产,生产原料为可食用的农业有机废料,对人体安全无毒,所生产的功能性低聚木糖可在食品、饲料,保健以及植物保护方面都有广阔的前景。 Enzyme preparation of the present invention is the Aspergillus enzyme production, raw materials for the production of edible organic agricultural waste, safe non-toxic to humans, the production of functional XOS may have food, feed, health and plant protection Broad prospects.

原料加工:用以生产功能性低聚木糖的原料主要是来自农业有机废料,如玉米芯、玉米秸杆、甘蔗渣、麦秸粉、麸皮、稻草、花生壳、高梁秸秆等,这些原料都富含半纤维素,而且对人体无毒,由它们制得功能性低聚木糖安全可靠。 Raw materials: raw materials for the production of functional XOS mainly organic waste from agriculture such as corn cobs, corn stover, bagasse, wheat straw, wheat bran, rice straw, peanut shells, straw, sorghum, these materials are hemicellulose rich, but also toxic to humans, they are prepared by a safe and reliable functionality of XOS. 玉米芯、秸杆等原料首先应筛选去除霉变成份,并干燥保存,再经分级粉碎后,过360目筛备用。 Corn cobs, straw and other raw materials should be first screened ingredients mildew removal, save and dried, and then pulverized and classified, through 360 mesh sieve for use.

半纤维素的制备:将上述原料用清水充分浸湿,溶胀后离心去除水分,再以10∶1(V/W)的固液比,加入碱,碱浓度控制得3~10%,室温下及高温时浸提,即室温下,搅拌浸提15~30小时,温度70~90℃时,搅拌浸提1~3小时,之后离心去除固形物,收集滤液,并用磷酸缓冲液中和至微酸性。 Hemicellulose prepared: The above starting materials were thoroughly wetted with water, the water was removed by centrifugation after swelling, and then to 10:1 (V / W) of the solid-liquid ratio, addition of a base, the alkali concentration is controlled to be 3 to 10%, at room temperature leaching at high temperatures, i.e. at room temperature and stirred for 15 to 30 hours leaching, when the temperature is 70 ~ 90 ℃, extraction was stirred for 1-3 hours, then centrifuged to remove solids, the filtrate was collected, and washed with phosphate buffer to the micro and acidity.

酶制剂的制备:所述的酶制剂主要是木聚糖酶(EC3.2.1.8),它可以由微生物分别通过固态或液态发酵方法来生产制得,经过发酵产生的木聚糖酶可以制备成部分纯化的酶制剂如硫酸铵酶粉、乙醇沉淀酶粉、丙酮酶粉或经sephadexG-100和DEAC-sephadex-A50纯化后的纯酶等。 Preparation of enzyme preparation: said enzyme preparation is mainly xylanase (EC 3.2.1.8), which can respectively be produced by solid or liquid prepared by a microbial fermentation process, after fermentation xylanase may be prepared into a partially purified enzyme powder enzyme preparations such as ammonium sulfate, ethanol precipitation enzyme powder, acetone or enzyme powder by sephadexG-100 and the DEAC-sephadex-A50 pure enzyme purification. 这些酶制剂都可以用于生产功能性低聚木糖。 These enzymes can be used in the production of functional XOS.

酶解反应:中和后的滤液为以半纤维素为主的溶液。 Enzymatic hydrolysis: filtrate after neutralization to hemicellulose-based solution. 上述酶粉或纯化后的酶制剂加入酶按1∶0.2~0.7(g/Iu)水解半纤维素,酶解反应条件为:温度35~55℃,PH4~7,反应时间控制在2小时内,以产物聚合度为依据,此时产物聚合度为2~10,反应完毕后升温至100℃,使酶活丧失。 After the enzyme or purified enzyme powder described above by adding an enzyme 1:0.2 ~ 0.7 (g / Iu) hydrolyzing hemicellulose, hydrolysis reaction conditions were: temperature 35 ~ 55 ℃, PH4 ~ 7, to control the reaction time within 2 hours to the product based on the degree of polymerization, then the polymerization product of from 2 to 10, after the completion of the reaction was heated to 100 deg.] C, the loss of enzyme activity.

低聚木糖的制备:所述的酶的水解液再经脱色,离子交换、浓缩等即可得低聚木糖为主的混合物,其中含少量单糖;经浓缩后的低聚木糖在低温<8℃下结晶,得低聚木糖结晶。 Preparation of XOS: the enzyme hydrolyzate was decolorized and then, ion-exchange, concentration and the like to give a mixture of xylo-based, which contain small amounts of monosaccharides; xylooligosaccharides was concentrated after low temperature <8 ℃ at crystallization, crystallization XOS. 后者为低聚木糖的精制产品。 Xylo latter refined products.

分组实验数据: Packet experimental data:

产品的分析鉴定:本发明所制得功能性低聚木糖可以经HPLC以乙晴和水按一定比例作为流动相检测,结果产品以木二糖为主;也可用TLC法,以硅胶G-250为载体,以正丁醇∶吡啶∶水=6∶4∶3展层,再用苯胺、二苯胺显色,也可分析产品组分。 Product analysis identified: XOS functionality prepared by the present invention may acetate Qinghe HPLC to certain proportion of water as mobile phase, and the results xylobiose based products; also be used by TLC, silica gel G- 250 is a vector, n-butanol: pyridine: water = 6:4:3 developing layer, and then, diphenylamine color, component products may also be analyzed. 这两种方法快速方便。 Both methods quickly and easily.

另外,将收集原料经碱处理后的固型物,与一绿醋酸反应可制备CMC。 Further, the collected solid type material after base-treated material can be prepared by reaction of a CMC and acid green.

Claims (4)

  1. 1.一种酶法制备功能性低聚木糖的生产工艺,它包括以下步骤:a.将玉米芯、秸杆、甘蔗渣、麦秸粉或稻草 进行干燥、粉碎、过筛制得原料;b.上述原料用清水浸湿、脱水,在室温下再用碱搅拌浸提15~30小时,70~90℃时搅拌浸提1~3小时,去除固形物,收集滤液,并用酸缓冲液中和至微酸性,得到半纤维素溶液;c.在上述半纤维素溶液中加入酶粉或纯化后的酶制剂水解半纤维素,酶的水解液再经脱色、离子交换、浓缩等过程得低聚木二糖为主的混合物。 An enzyme preparation of functional XOS production process, comprising the steps of: a cobs, straw, bagasse, wheat straw or straw was dried, pulverized, sieved to prepare a material; b. the above material soaked with water, dehydrated, and then stirred at room temperature for 15 to 30 hours after leaching with alkali, stirred for 70 ~ 90 ℃ leaching when 1 to 3 hours, removing the solids, the filtrate was collected, and washed with an acid buffer and to slightly acidic, hemicellulose solution was obtained;. c hydrolase formulation after addition of the above purified enzyme powder or solution, hemicellulose hemicellulose, and then the enzyme hydrolyzate bleaching, ion exchange, concentration and the like processes to give oligomerization a mixture of wood-based disaccharide. d.上述的酶制剂主要是木聚糖酶(EC3.2.1.8),它可以由微生物分别通过固态或液态发酵方法制得,经过发酵产生的木聚糖酶可以制备成部分纯化的酶制剂或经sephadex G-100和DEAE-sephadex-A50纯化后的纯酶。 d. above enzyme preparations are mainly xylanase (EC 3.2.1.8), which can be solid or liquid, respectively, through fermentation process by microorganisms obtained after fermentation of the xylanase can be prepared as part of a purified enzyme preparation of or with sephadex G-100, and after DEAE-sephadex-A50 pure enzyme purification.
  2. 2.根据权利要求1所述的酶法制备功能性低聚木糖的生产工艺,其特征在于:玉米芯或秸杆原料首先应筛选去除霉变成份,并干燥保存,用清水充分浸湿,溶胀后离心去除水分,再以10∶1(V/W)的固液比,加入碱,碱浓度控制在3~10%,室温下及高温时浸提,之后,离心去除固形物,收集滤液,并用磷酸缓冲液中和。 Enzymatic preparation according to claim functional XOS production process according to claim 1, wherein: the core or corn stover feedstock should first screening component removing mildew, and stored dry, fully soaked with water, after centrifugation to remove the water-swellable, then 10:1 (V / W) of the solid-liquid ratio, addition of a base, the base concentration at 3 to 10%, at room temperature and high temperature leaching, after centrifugation to remove solids, the filtrate was collected , and neutralized with phosphate buffer.
  3. 3.根据权利要求1所述的酶法制备功能性低聚木糖的生产工艺,其特征在于:上述酶粉或纯化后的酶制剂加入酶按1∶0.2~0.7(g/Iu)水解半纤维素,酶解反应条件为:温度35~55℃,PH4~7,反应时间控制在2小时内,以产物聚合度为依据,此时产物聚合度为2~10,反应完毕后升温至100℃,使酶活丧失。 The enzyme preparation of functional XOS production process according to claim 1, wherein: the enzyme or purified enzyme powder described above by adding an enzyme 1:0.2 ~ 0.7 (g / Iu) Hydrolysis half cellulose, hydrolysis reaction conditions were: temperature 35 ~ 55 ℃, PH4 ~ 7, to control the reaction time within 2 hours, according to degree of polymerization of the product, then the product of polymerization of from 2 to 10, after the completion of the reaction was warmed to 100 ℃, loss of enzyme activity.
  4. 4.根据权利要求1所述的酶法制备功能性低聚木糖的生产工艺,其特征在于:所述的酶的水解液再经脱色、离子交换、浓缩等即可得低聚木糖为主的混合物,其中含少量单糖,经浓缩后的低聚木糖在低温<8℃下结晶,得低聚木糖结晶。 The enzyme preparation of functional XOS production process according to claim 1, wherein: the enzyme hydrolyzate was decolorized and then, ion-exchange, concentration and the like can be obtained as xylooligosaccharide the main mixture, which contains a small amount of a monosaccharide, after xylooligosaccharides was concentrated under a low temperature <8 ℃ crystallization, crystallization XOS.
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CN101381753B (en) 2008-10-28 2012-05-30 上海师范大学 Method for preparing rice husk xylo-oligosaccharides
CN101843329B (en) 2009-12-30 2013-01-02 吉林大学 Method for preparing dietary fiber by adopting corn bran hydrolyzed by multifunctional enzyme
CN101748157B (en) 2010-01-08 2012-07-25 安徽丰原发酵技术工程研究有限公司 Corn cob acid hydrolysis residue and straw alkali cooking residue mixing enzymolysis method
CN101781669B (en) * 2010-03-22 2013-07-17 苏州工业园区尚融科技有限公司 Method for preparing high-purity xylo-oligosaccharide by adopting straws
CN102399842B (en) * 2011-11-15 2014-10-29 上海师范大学 The method of preparing extracts from straw Xylo
CN103467627A (en) * 2013-09-14 2013-12-25 大连工业大学 Preparation method for corn stalk hemicellulose
CN103614435B (en) * 2013-12-02 2016-05-04 广西壮族自治区农业科学院农产品加工研究所 To a process for the preparation of sugar cane bagasse Xylooligosaccharides
CN103757072A (en) * 2014-01-10 2014-04-30 杭州师范大学 Method for preparing xylan and xylo-oligosaccharide from rice straw
CN104357516A (en) * 2014-11-07 2015-02-18 宜宾雅泰生物科技有限公司 Process for producing food-grade xylo-oligosaccharide from viscose fiber squeezed alkali liquor
CN104928331A (en) * 2015-07-08 2015-09-23 青岛嘉瑞生物技术有限公司 Technology for preparing functional xylo-oligosaccharide by comprehensively utilizing wheat straw

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