WO2013143503A1 - 甘薯渣酶法水解糖工艺 - Google Patents

甘薯渣酶法水解糖工艺 Download PDF

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WO2013143503A1
WO2013143503A1 PCT/CN2013/073545 CN2013073545W WO2013143503A1 WO 2013143503 A1 WO2013143503 A1 WO 2013143503A1 CN 2013073545 W CN2013073545 W CN 2013073545W WO 2013143503 A1 WO2013143503 A1 WO 2013143503A1
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per gram
sweet potato
enzyme
sugar
cellulase
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PCT/CN2013/073545
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English (en)
French (fr)
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吴允山
严共鸿
易勇
寇正恩
吴震
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山东宏河圣齐生物工程有限公司
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Priority to EP13770343.5A priority Critical patent/EP2832861B1/en
Priority to ES13770343.5T priority patent/ES2686539T3/es
Priority to PL13770343T priority patent/PL2832861T3/pl
Priority to US14/385,817 priority patent/US9506096B2/en
Publication of WO2013143503A1 publication Critical patent/WO2013143503A1/zh

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P19/00—Preparation of compounds containing saccharide radicals
    • C12P19/02—Monosaccharides
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L29/00—Foods or foodstuffs containing additives; Preparation or treatment thereof
    • A23L29/30—Foods or foodstuffs containing additives; Preparation or treatment thereof containing carbohydrate syrups; containing sugars; containing sugar alcohols, e.g. xylitol; containing starch hydrolysates, e.g. dextrin
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L29/00—Foods or foodstuffs containing additives; Preparation or treatment thereof
    • A23L29/30—Foods or foodstuffs containing additives; Preparation or treatment thereof containing carbohydrate syrups; containing sugars; containing sugar alcohols, e.g. xylitol; containing starch hydrolysates, e.g. dextrin
    • A23L29/35—Degradation products of starch, e.g. hydrolysates, dextrins; Enzymatically modified starches
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P19/00—Preparation of compounds containing saccharide radicals
    • C12P19/14—Preparation of compounds containing saccharide radicals produced by the action of a carbohydrase (EC 3.2.x), e.g. by alpha-amylase, e.g. by cellulase, hemicellulase
    • C—CHEMISTRY; METALLURGY
    • C13—SUGAR INDUSTRY
    • C13K—SACCHARIDES OBTAINED FROM NATURAL SOURCES OR BY HYDROLYSIS OF NATURALLY OCCURRING DISACCHARIDES, OLIGOSACCHARIDES OR POLYSACCHARIDES
    • C13K1/00—Glucose; Glucose-containing syrups
    • C13K1/06—Glucose; Glucose-containing syrups obtained by saccharification of starch or raw materials containing starch
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
    • Y02E50/00—Technologies for the production of fuel of non-fossil origin
    • Y02E50/10—Biofuels, e.g. bio-diesel

Definitions

  • the present invention relates to the field of biology, and in particular to a method for producing sugar by using a biomass sweet potato slag for microbial fermentation as a sugar source. Background technique
  • Sweet potato is one of the important food crops in China. According to statistics, China's sweet potato cultivation and processing ranks first in the world, with a total output of 85.2 million tons, of which 55% is about 46.86 million tons converted into industrial raw materials. Sweet potato residue is a waste of large amount of waste produced in the processing of starch in fresh sweet potato. The huge biomass regeneration resources are still to be developed and utilized.
  • Fresh sweet potatoes generally contain 30% dry matter and 70% moisture.
  • Sweet potato processing produces a large amount of wastewater containing cell liquid, which is sweet potato starch wastewater, with various nutrients such as soluble carbohydrates, proteins, vitamins and trace elements.
  • the sweet potato starch processed potato residue was analyzed, and the main chemical components were water, starch, crude protein, fiber, fat and the like.
  • the waste sweet potato slag accounts for about 10% of the raw materials.
  • fresh sweet potato slag usually contains about 90% of the cell liquid wastewater, and a large amount of wet sweet potato slag is accumulated.
  • Effective development and utilization, and because of the high water holding capacity and swelling of potato dregs, the new potato residue with multiple nutrients of sugar and nitrogen has more than 90% water content, and its chemical oxygen demand COD > 15000mg/L, easy to store. It is rancid by fermentation of miscellaneous bacteria, which seriously pollutes the environment and causes huge waste of biomass regeneration resources.
  • Sweet potato slag dry meter generally containing more than 50% starch, 22 ⁇ 26% fiber, the main components of fiber are cellulose, hemicellulose, lignin and pectin, etc., is thousands of glucose glycosyl
  • the dense structure of carbohydrates is difficult to be degraded by the cellulase secreted by the commercially available Trichoderma koningii; the sweet potato is separated by the masher and the starch produced by the sieving equipment, and the residual starch in the potato residue is liquefied by ⁇ -amylase. Glycosylation with glucoamylase, its degradation is not complete; other high molecular polysaccharides are more difficult to degrade, and the hydrolysis rate is low.
  • the object of the present invention is to provide a sugar-making method for multi-enzyme combined hydrolysis to destroy cell wall, translating monomer and enzymatic multi-component biodegradation reaction of several polysaccharide components of potato residue, thereby improving glucose conversion rate and ensuring product The purity of the sugar.
  • the invention provides a method for preparing sugar by using sweet potato residue, comprising the following steps: Step 1: taking sweet potato residue to smash or grinding wet slag, adding sweet potato starch wastewater to adjust pulp; adjusting slurry ⁇ 4.0-6.0, at 20 ⁇ 70 °C, adding cellulase and adding ⁇ -glucanase for 2 ⁇ 10h;
  • Step 2 at 25 ⁇ 60 ° C, add xylanase and pectinase at pH 3.5 ⁇ 6.0 for 2 ⁇ 10h; temperature 30 ⁇ 50°CpH2.5 ⁇ 6.0, add acid protease to hydrolyze for l ⁇ 8h; 110 ⁇ 120 °C, 30min to carry out the enzyme treatment; adjust the pH 5.5 ⁇ 8.0, add high temperature ⁇ -amylase or medium temperature ⁇ -amylase hydrolysis l ⁇ 2h; cool down 40 ⁇ 65 °C, adjust the pH 3.0 ⁇ 5.5, Glycation enzyme hydrolysis 10 ⁇ 20h;
  • Step 3 The solid solution is separated from the liquid, and the liquid is concentrated to obtain a liquid sugar product, and the composition thereof is basically glucose.
  • the pulverization in step 1 is a sieve diameter of 830 ⁇ n! ⁇ 150 ⁇ m.
  • the pulping in step 1 is mixed according to the mass ratio of the liquid to 1: 4 ⁇ 6.
  • step 1 adds 70 to 200 U of cellulase per gram of material.
  • step 1 adds 4.5 to 13.51 ⁇ -glucanase per gram of material.
  • step 2 adds 14.5 to 29 U of xylanase per gram of material, and adds 9 to 30 U of pectinase per gram of material.
  • step 2 adds 10 to 15 U of acid protease per gram of material.
  • step 2 is added with a temperature-resistant alpha-amylase of 12 to 20 U per gram.
  • step 2 adds 100 to 300 U of saccharification enzyme per gram of material.
  • step 2 after step 3 further comprises the following steps: adjusting pH 5.5 ⁇ 8.0, adding high temperature alpha-amylase for secondary liquefaction, 45 min; cooling to 40 ⁇ 65 ° C, adjusting pH 3.0 ⁇ 5.5, Saccharification enzyme and cellulase are used for saccharification; preferably, 100 U of saccharification enzyme per gram of material is added, and 70 U of cellulase is added per gram of material.
  • the conversion rate of sweet potato slag to glucose is more than 65%, and the conversion rate of starch to reducing sugar in sweet potato slag can reach 110%, which is higher than the conversion rate of traditional double enzyme method.
  • the enzymatic hydrolysis conversion rate refers to the mass fraction of the dry weight of the tested potato slag by the total amount of the reducing sugar released by the enzymatic hydrolysis of the potato dregs, indicating various polysaccharides in the potato dregs.
  • the mechanism of the combined enzymatic hydrolysis of the present invention is: synergistic action of cellulase, ⁇ -glucanase, pectinase and xylase, destroying the cell wall, disassembling the attached lignin and pectin (protecting the cellulose matrix) And the glycoprotein in the binding state, enzymatically degrading the multi-component high molecular polysaccharide in the sweet potato residue into a biochemical reaction of the monosaccharide, thereby improving the conversion rate.
  • the sweet potato slag contains 50% starch (dry basis) or more, and the loosely structured hemicellulose and pectin polysaccharide can be completely or partially enzymatically converted into sugar by hydrolase, and the sample prepared by the method of the invention is subjected to Dionex company.
  • HPAEC analysis select PA10 analytical column, using 18 mM NaOH as buffer, flow rate: lml/min, each sample collection time is 40min, identification is basically glucose, is a high-quality sugar material for microbial fermentation, can be used in industrial fermentation industry .
  • the liquid sugar component prepared by the process is basically identified as glucose by the Chinese Academy of Sciences, instead of other reducing monosaccharides, indicating that the preparation process of the patent can basically achieve complete enzymatic hydrolysis.
  • the method of the invention has mild process conditions, simple process equipment, strong specificity, improved total reducing sugar conversion rate, pure product quality, and can solve the serious environmental pollution problem of sweet potato slag, and has good industrial application prospect.
  • the invention discloses a sugar potato slag sugar-making method, which can be used by those skilled in the art. Capacity, appropriate improvement of process parameters. It is to be understood that all such alternatives and modifications are obvious to those skilled in the art and are considered to be included in the present invention.
  • the method and the application of the present invention have been described by the preferred embodiments, and it is obvious that the method and application described herein may be modified or appropriately modified and combined without departing from the scope of the present invention. The technique of the present invention is applied.
  • the liquid is subjected to solid-liquid separation, and the liquid is concentrated to obtain a liquid sugar product.
  • the sweet potato slag dry matter conversion rate to glucose was 65.6%.
  • the conversion rate of "starch” to monosaccharide in sweet potato slag was 108%.
  • the starch content of sweet potato residue is determined according to the "National Standard of the People's Republic of China GB/T 5514-2008" grain and oil test starch content in the grain oil, the method is as follows: Quantitatively weigh the measured "starch” content of sweet potato residue, hydrolyzed mash , Washing and centrifuging, measuring the total amount of reducing sugar in the clear solution, and calculating the conversion rate according to the following formula:
  • the temperature was lowered to 42 °C, pH 4.0, and xylanase and pectinase (1.45 U per gram of xylanase plus 9 U of pectinase) were hydrolyzed for 4 h.
  • the acid protease (12 U per gram of enzyme) was hydrolyzed for 2 h at a temperature of 50 ° C and pH 4.0. The temperature was raised at 115 ° C to kill the enzyme. Adjust the pH to 6.5 and add 5% high temperature alpha-amylase (15 U per gram of enzyme) to liquefy for 45 min. The temperature was lowered to 60 ° C and the pH was 4.5, and the saccharification enzyme (100 U per gram of enzyme added) was hydrolyzed for 15 h. In the enzymatic decanter batch, pH 6.5 was directly adjusted, and a high temperature ⁇ -amylase (12 U per gram of enzyme) was added for secondary liquefaction for 45 minutes. Cool down to 60 °C, adjust pH 4.5, add glucoamylase (100 U per gram of enzyme) and cellulase (100 U per gram of enzyme) for saccharification.
  • the temperature was lowered to 50 ° C, pH 4.6, and xylanase and pectinase (20 U per gram plus xylanase plus 30 U of pectinase) were hydrolyzed for 6 h.
  • the acid protease (15 U per gram of enzyme added) was hydrolyzed for 2 h at a temperature of 50 ° C and pH 4.0.
  • the enzyme was inactivated by heating at 120 °C.
  • the pH was adjusted to 6.5, and the high temperature alpha-amylase (20 U per gram of enzyme added) was liquefied for 50 min.
  • the temperature was lowered to 60 ° C to adjust the pH to 4.5, and the saccharification enzyme (300 U per gram of enzyme) was hydrolyzed for 20 h.
  • pH 6.5 was directly adjusted, and high temperature ⁇ -amylase (12 U per gram of enzyme) was added for secondary liquefaction for 45 min.
  • Example 4 Analysis of the liquid sugar product obtained in any of Examples 1-3
  • the collection time for each sample was 40 min.
  • Code 3# 2# Concentrate of liquefied feed liquid centrate.
  • a saccharification enzyme i.e., a liquid sugar product obtained in any of Examples 1-3
  • 1,2,3,4# samples are all diluted 100 times, 5# samples are translated 200 times, tested by the Sugar Bioengineering Center of the Institute of Microbiology, Chinese Academy of Sciences: using the HPAEC analysis of Dionex, the PA10 analytical column was selected. Tested using rhamnose, arabinose, galactose, glucose, xylose and mannose standards.
  • the 1# sample has two peaks: 8.667 and 11.917, and the glucose content is 10.5 mg/ml. The amount of arabinose is small.
  • the main peak time of the 2# sample was 11.750 and the content was 172 mg/ml.

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Abstract

本发明涉及生物领域，具体涉及了以供微生物发酵用的生物质甘薯渣为糖源的制糖方法。本发明所述方法制得的液糖产品，其成分基本为葡萄糖。本发明所述方法工艺简单，专一性强，所得产品品质好，收率高，能解决甘薯渣严重污染环境问题，具有良好的工业应用前景。

Description

甘薯渣酶法水解糖工艺
本申请要求于 2012 年 03 月 30 日提交中国专利局、 申请号为 201210090800.8、 发明名称为 "甘薯渣酶法水解糖工艺" 的中国专利申请 的优先权， 其全部内容通过引用结合在本申请中。 技术领域
本发明涉及生物领域，具体涉及以供微生物发酵用的生物质甘薯渣为 糖源的制糖方法。 背景技术
甘薯是我国重要的粮食作物之一，据统计，我国甘薯种植与加工均居 世界第一， 总产量达 8520万吨， 其中 55%约 4686万吨转成工业原料。 甘薯渣是鲜甘薯加工淀粉过程中产生的大量渣滓而作废物处理，巨大的生 物质再生资源尚待人们开发利用。
鲜甘薯一般含干物质 30%, 水分 70%。 甘薯加工产生大量的含细胞 液废水即为甘薯淀粉废水， 有各种营养有机物， 如可溶性的碳水化合物、 蛋白质、 维生素和微量元素。 对甘薯淀粉加工的薯渣进行分析， 主要化学 成分为水分、 淀粉、 粗蛋白、 纤维、 脂肪等。
用甘薯精细加工生产淀粉、粉条、方便面粉丝等食品产出废甘薯渣约 占原料 10%左右， 刚下生产线时鲜甘薯渣通常含细胞液废水 90%左右， 数量巨大的湿甘薯渣堆积，未能有效的开发利用，且由于薯渣纤维高持水 力和溶胀性， 有糖、 氮多种营养成分的新薯渣， 含水 90%以上， 其废水 化学耗氧量 COD > 15000mg/L, 存放又易受杂菌发酵而酸败， 严重污染 环境， 造成生物质再生资源的巨大浪费。
近年来甘薯渣开发热点是用酸法、 酶法、 筛法去掉甘薯渣大部分的淀 粉、蛋白和脂肪，提取膳食纤维和果胶、制成产品，但产品市场需求不旺， 尚未形成大型产业化规模。
甘薯渣折干计， 一般含淀粉 50%以上， 纤维 22 ~ 26%, 纤维主要构 成成分是纤维素、半纤维素、木质素和果胶等， 是数以千计葡萄糖糖基致 密结构的碳水化合物，难以为市售的康氏木霉分泌的纤维素酶所降解；甘 薯经锉磨机细碎和筛理设备产生的淀粉分离出来， 薯渣中残留淀粉经 α- 淀粉酶液化和糖化酶糖化， 其降解不完全； 其他高分子多糖更难降解， 水 解率低。 因此， 寻找一种能够高效利用甘薯渣转化为葡萄糖的方法、 开辟 新糖源并实行产业化是当前开展生物质甘薯渣废弃资源综合利用的一项 重要任务， 这种非粮糖源具有显著的环保效益、 社会效益和经济效益。 发明内容
本发明目的是提供多酶联合水解破坏薯渣几种多糖组分的细胞壁、 译放单体，酶促多组分生化降解反应的一种制糖方法，该方法提高葡萄糖 的转化率， 保证产品糖的纯度。
本发明提供的一种利用甘薯渣制糖的方法， 包含以下步骤： 步骤 1 :取甘薯渣粉碎或磋磨湿渣，加甘薯淀粉废水调浆；调浆料 ρΗ 4.0-6.0, 在 20〜70°C , 加纤维素酶和加 β-葡聚糖酶水解 2〜10h;
步骤 2: 在 25〜60°C , 在 pH3.5〜6.0加木聚糖酶和果胶酶水解 2〜10h; 温度 30〜50°CpH2.5〜6.0,加酸性蛋白酶水解 l〜8h;升温 110〜120°C , 30min 进行灭酶处理； 调 pH 5.5〜8.0, 加耐高温 α-淀粉酶或中温 α-淀粉酶水解 l〜2h; 降温 40〜65°C , 调 pH3.0〜5.5 , 加糖化酶水解 10〜20h;
步骤 3: 对料液进行固液分离， 液体浓缩即得液糖产品， 其成分基本 为葡萄糖。
作为优选， 步骤 1所述粉碎为筛径 830 μ n！〜 150 μ m。
作为优选， 步骤 1所述调浆为按料液质量比 1 : 4〜6混合。
作为优选， 步骤 1每克料加 70〜200U纤维素酶。
作为优选， 步骤 1每克料加 4.5〜13.51Ιβ-葡聚糖酶。
作为优选， 步骤 2每克料加 14.5〜29U木聚糖酶， 每克料加 9〜30U果 胶酶。
作为优选， 步骤 2每克料加酸性蛋白酶 10〜15U。
作为优选， 步骤 2每克料加耐高温 α-淀粉酶 12〜20U。
作为优选， 步骤 2每克料加糖化酶 100〜300U。 作为优选， 步骤 2之后步骤 3之前还包括以下步骤： 调 pH5.5〜8.0, 加耐高温 α-淀粉酶进行二次液化， 45min;降温至 40〜65°C ,调 pH3.0〜5.5, 加糖化酶和纤维素酶进行糖化； 作为优选， 每克料加糖化酶 100U, 每克 料加纤维素酶 70U。
采用本发明所述方法， 用国产市售的纤维素酶、 β-葡聚糖酶、 木聚糖 酶和果胶酶、 酸性蛋白酶、 耐高温 α-淀粉酶、 糖化酶对甘薯渣酶法制糖， 以不同的料液比进行比较， 甘薯渣干物质对葡萄糖转化率达 65%以上， 相对于甘薯渣中淀粉对还原糖的转化率可达 110%, 较传统双酶法的转化 率有较大的提高。所述酶水解转化率是指对薯渣经酶法水解译放出的还原 糖总量按葡萄糖计占试料薯渣干物质量的质量分数,表明薯渣中各种多糖
(淀粉及果胶、 半纤维素和纤维素等） 水解的程度。
实验表明， 薯渣中各种多糖组分用国产酶制剂水解技术难度大， 特 别是纤维素和木质素降解制糖转化率低。 本发明采用联合酶解的机理是: 纤维素酶、 β-葡聚糖酶、 果胶酶和木糖酶的协同作用， 破坏细胞壁， 拆开 连接的木质素和果胶 (保护纤维素基质 )及结合态的糖蛋白， 酶促甘薯渣 中多组分高分子多糖降解成单糖的生化反应， 提高转化率。
甘薯渣中含淀粉 50% (干基） 以上， 结构疏松的半纤维素和果胶多 糖均可由水解酶类完全或部分酶解转化为糖，本发明所述方法制得的样品 经 Dionex公司的 HPAEC分析， 选取 PA10分析柱， 以 18mM的 NaOH 为緩冲液， 流速： lml/min, 每个样收集时间为 40min, 鉴定基本为葡萄 糖， 是微生物发酵用的优质糖料， 可用于工业发酵行业。
用本工艺制备的液糖组分， 经中科院科学鉴定基本为葡萄糖， 而不 是其他还原性单糖， 说明本专利的制备工艺可基本达到酶解完全。
本发明所述方法工艺条件温和， 工艺设备简单， 专一性强， 提高了 总还原糖转化率， 产品品质纯， 且能解决甘薯渣严重污染环境问题， 具有 良好的工业应用前景。 具体实施方式
本发明公开了一种甘薯渣制糖方法，本领域技术人员可以借鉴本文内 容， 适当改进工艺参数实现。特别需要指出的是， 所有类似的替换和改动 对本领域技术人员来说是显而易见的，它们都被视为包括在本发明。本发 明的方法及应用已经通过较佳实施例进行了描述,相关人员明显能在不脱 离本发明内容、精神和范围内对本文所述的方法和应用进行改动或适当变 更与组合， 来实现和应用本发明技术。
为了使本领域的技术人员更好地理解本发明的技术方案， 下面结合 具体实施例对本发明作进一步的详细说明。 实施例 1 :
取山东某地甘薯渣（水份 12.7%, 淀粉 56.19%, 粗蛋白 3.69% ), 粉 碎至 250 μ ιη筛径， 在 30L罐中按料液比 1 : 5加甘薯淀粉废水调浆。 浆 料调 ρΗ至 5.5, 升温至 55°C , 加纤维素酶 (每克料加酶 100U )和加 β- 葡聚糖酶（每克料加酶 10U )水解 4h。 降温至 45°C , pH4.5, 加木聚糖酶 和果胶酶（每克料加木聚糖酶 29U, 加果胶酶 20U ) 水解 4h。
在温度 50°C、 pH4.0, 加酸性蛋白酶 (每克料加酶 10U )水解 2h。 升 温 110°C灭酶。 调 pH至 6.5 , 加耐高温 α-淀粉酶 (每克料加酶 12U )液 化 45min。 降温至 60°C调 pH为 4.5, 加糖化酶（每克料加酶 200U )水解 10h。在该酶解罐批中直接调 pH6.5,加耐高温 α-淀粉酶（每克料加酶 12U ) 进行二次液化， 60min。 降温至 60°C , 调 pH4.5, 加糖化酶（每克料加 酶 100U )和纤维素酶（每克料加酶 100U )进行糖化。
糖化结束后对料液进行固液分离，液体浓缩即得液糖产品。甘薯渣干 物质对葡萄糖转化率为 65.6%。 相对于甘薯渣中 "淀粉"对单糖的转化率 为 108%。
甘薯渣淀粉含量计算按 "中华人民共和国国家标准 GB/T 5514— 2008"粮油检验粮食油料中淀粉含量测定， 方法如下： 定量称取已测 "淀 粉"含量的甘薯渣， 水解后的糖化醪离心， 洗涤离心， 测计清液还原糖总 量， 按下式计算转化率：
甘薯渣质量 (折干 )对糖的转化率（％ ) =【清液还原糖总量 (折干 )
÷ 甘薯渣质量（折干）】 X loo 实施例 2:
取山东某地甘薯渣（水份 12.7%, 淀粉 56.19%, 粗蛋白 3.69% ), 粉 碎至 380 μ m筛径， 在 30L罐中按料液比 1 : 4.5加甘薯淀粉废水调浆。 浆料调 pH至 5.0, 升温至 50°C , 加纤维素酶 (每克料加酶 70U )和加 β- 葡聚糖酶（每克料加酶 10U )水解 4h。 降温至 42 °C , pH4.0, 加木聚糖酶 和果胶酶（每克料加木聚糖酶 14.5U, 加果胶酶 9U )水解 4h。
在温度 50°CpH4.0, 加酸性蛋白酶（每克料加酶 12U )水解 2h。 升温 115°C灭酶。 调 pH至 6.5 , 加耐高温 α-淀粉酶 (每克料加酶 15U )液化 45min。降温至 60°C调 pH为 4.5 ,加糖化酶（每克料加酶 100U )水解 15h。 在该酶解罐批中直接调 pH6.5, 加耐高温 α-淀粉酶 (每克料加酶 12U )进 行二次液化， 45min。降温至 60 °C ,调 pH4.5,加糖化酶（每克料加酶 100U ) 和纤维素酶（每克料加酶 100U )进行糖化。
糖化结束后对料液进行固液分离，液体浓缩即得液糖产品。甘薯渣干 物质对葡萄糖转化率为 65.2%。 相对于甘薯渣中 "淀粉"对单糖的转化率 为 106%。 实施例 3:
取山东某地甘薯渣（水份 12.7%, 淀粉含量 > 62%,粗蛋白 3.69% ), 粉碎至 230 μ m筛径， 在 30L罐中按料液比 1： 4加甘薯淀粉废水调浆。 浆料调 pH至 6.0, 升温至 60°C , 加纤维素酶 (每克料加酶 200U )和加 β- 葡聚糖酶（每克料加酶 13.5U )水解 4h。 降温至 50°C , pH4.6, 加木聚糖 酶和果胶酶（每克料加木聚糖酶 20U, 加果胶酶 30U )水解 6h。
在温度 50°CpH4.0, 加酸性蛋白酶（每克料加酶 15U )水解 2h。 升温 120°C灭酶。 调 pH至 6.5, 加耐高温 α-淀粉酶 (每克料加酶 20U )液化 50min。降温至 60°C调 pH为 4.5 ,加糖化酶（每克料加酶 300U )水解 20h。 在该酶解罐批中直接调 pH6.5, 加耐高温 α-淀粉酶 (每克料加酶 12U )进 行二次液化， 45min。降温至 60 °C ,调 pH4.5,加糖化酶（每克料加酶 100U ) 和纤维素酶（每克料加酶 100U )进行糖化。 糖化结束后对料液进行固液分离，液体浓缩即得液糖产品。甘薯渣干 物质对葡萄糖转化率为 68.4%。 相对于甘薯中 "淀粉"对单糖的转化率为 110%。 实施例 4: 取实施例 1-3任一项所得液糖产品分析
样品处理：
12000rpm, lOmin. 取上清。 用 0.45um的 Pal膜过滤。
样品分析： Dionex公司的 HPAEC分析 , 选取 PA 10分析柱。
緩冲液： 18mM的 NaOH
流速： lml/min
每个样收集时间为 40min。
实验结果：
选取样品中可能含有的单糖做标准品， 在 PA10中分析。 表 1中表示 的单一糖的保留时间。 表 1 单一糖的保留时间
Figure imgf000007_0001
经验证木糖 xylose和甘露糖 mannose在此条件下不能分开，所以 13.5 分钟的保留时间是木糖 xylose和甘露糖 mannose的混合物。 液糖组分分析
取样： 在同一罐批中按时间顺序和加不同酶水解后取样送检。
代号 1#: 0〜10h, 加纤维素酶和 β -葡聚糖酶（4h ); 继之加木聚糖 酶和果胶酶（4h ); 又加酸性蛋白酶（2h )后取样、 离心分离得离心清液。
代号 2#: 101！〜 llh (接 1#工艺 )继加 α-淀粉酶水解的离心液。
代号 3#: 2#液化料液离心液的浓缩液。 代号 4#: 111！〜 26h (接 2#工艺）继加糖化酶水解的糖化料液的离心 液 (即实施例 1-3任一项所得液糖产品 )
代号 5# 4#糖化液离心液的浓缩液
1,2,3,4#样品均稀译 100倍， 5#样品稀译 200倍， 经中科院微生物所 糖生物工程中心检测：用 Dionex公司的 HPAEC分析 ,选取 PA10分析柱。 使用鼠李糖、阿拉伯糖、半乳糖、葡萄糖、木糖和甘露糖标准品进行测试。
1#样品有两个峰: 8.667和 11.917, 葡萄糖含量是 10.5mg/ml。 阿拉 伯糖的含量很少。
2#样品的主峰时间是 11.750 , 含量是 172mg/ml。
3#样品： 11.417, 糖的含量为 548mg/ml。
4#样品： 11.583。 糖的含量为 184mg/ml, 且未检测到二糖、 多糖。 5#样品： 11.5。 糖的含量为 511mg/ml, 且未检测到二糖、 多糖。 初步结果分析： 样品中组分大部分是葡萄糖， 即除了 "液化工艺"前的 1#样中含有微量鼠李糖和葡萄糖外， 其他工艺重点的四个样全是葡萄糖， 亦即糖化料液离心液全是葡萄糖。 综合考虑， 实施例 1-3任一项所得液糖 产品样品中组分基本全部是葡萄糖。 以上对本发明所提供的甘薯渣酶法水解糖工艺进行了详细介绍。本文 明只是用于帮助理解本发明的方法及其核心思想。应当指出，对于本技术 领域技术人员来说，在不脱离本发明原理的前提下，还可以对本发明进行 若干改进和修饰， 这些改进和修饰也落入本发明权利要求的保护范围内。

Claims

权 利 要 求
1. 一种利用甘薯渣制糖的方法， 其特征在于， 包含以下步骤： 步骤 1 :取甘薯渣粉碎或磋磨湿渣，加甘薯淀粉废水调浆；调浆料 pH 4.0-6.0, 在 20〜70°C , 加纤维素酶和加 β-葡聚糖酶水解 2〜10h;
步骤 2: 在 25〜60°C , pH3.5〜6.0加木聚糖酶和果胶酶水解 2〜10h; 温 度 30〜50°C、 pH2.5〜6.0,加酸性蛋白酶水解 l〜8h;升温 110〜120°C , 30min 进行灭酶处理； 调 pH 5.5〜8.0, 加耐高温 α-淀粉酶或中温 α-淀粉酶水解 l〜2h; 降温 40〜65°C , 调 pH3.0〜5.5 , 加糖化酶水解 10〜20h;
步骤 3: 对料液进行固液分离， 液体浓缩即得液糖产品， 其成分基本 为葡萄糖。
2、根据权利要求 1所述的方法，其特征在于，步骤 1所述粉碎为 830 μ m〜150 μ m陣径。
3、 根据权利要求 1所述的方法， 其特征在于， 步骤 1所述调浆为按 料液质量比 1 : 4〜6混合。
4、根据权利要求 1所述的方法，其特征在于，步骤 1每克料加 70〜200U 纤维素酶。
5、 根据权利要求 1所述的方法， 其特征在于， 步骤 1每克料加 4.5〜13.51Ιβ-葡聚糖酶。
6、 根据权利要求 1所述的方法， 其特征在于， 步骤 2每克料加
14.5〜29U木聚糖酶， 每克料加 9〜30U果胶酶。
7、 根据权利要求 1所述的方法， 其特征在于， 步骤 2每克料加酸性 蛋白酶 10〜15U。
8、 根据权利要求 1所述的方法， 其特征在于， 步骤 2每克料加耐高 温 α-淀粉酶 12〜20U。
9、 根据权利要求 1所述的方法， 其特征在于， 步骤 2每克料加糖化 酶 100〜300U。
10、根据权利要求 1所述的方法， 其特征在于， 步骤 2之后还包括以 下步骤： 调 pH 5.5〜8.0, 加耐高温 α-淀粉酶进行二次液化， 45min; 降温 0〜65°C, 调 pH3.0〜5.5, 加糖化酶和纤维素酶进行糖化。
11、 根据权利要求 10 所述的方法， 其特征在于， 每克料加糖化酶 〜300U, 每克料加纤维素酶 70〜200U。
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CN113881736A (zh) * 2021-11-09 2022-01-04 无锡德林海环保科技股份有限公司 一种芦苇资源化利用工艺
CN113881736B (zh) * 2021-11-09 2024-03-05 无锡德林海环保科技股份有限公司 一种芦苇资源化利用工艺

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