CN113388652B - Preparation method of green efficient nanocellulose - Google Patents
Preparation method of green efficient nanocellulose Download PDFInfo
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Abstract
本发明属于生物质材料技术领域，尤其涉及一种绿色高效纳米纤维素的制备方法。本发明提供的绿色高效纳米纤维素的制备方法，包括以下步骤：将生物质纤维素、纤维素酶水溶液和果胶酶水溶液混合进行酶解处理；所述纤维素酶水溶液和所述果胶酶水溶液的总质量与所述生物质纤维素的质量比为(15～30):1，所述纤维素酶水溶液中纤维素酶的酶活力为30～150U/mL，所述果胶酶水溶液中果胶酶的酶活力为300～1500U/mL，所述酶解处理的时间为15～60min。本发明提供的纳米纤维素的制备方法，采用纤维素酶水溶液和果胶酶水溶液共同处理生物质纤维素，显著提高了纳米纤维素的生产效率和得率且绿色环保。
The invention belongs to the technical field of biomass materials, in particular to a method for preparing green and high-efficiency nano-cellulose. The preparation method of green high-efficiency nano-cellulose provided by the present invention comprises the following steps: mixing biomass cellulose, cellulase aqueous solution and pectinase aqueous solution to carry out enzymolysis treatment; said cellulase aqueous solution and said pectinase The mass ratio of the total mass of the aqueous solution to the biomass cellulose is (15-30):1, the enzyme activity of the cellulase in the aqueous solution of cellulase is 30-150U/mL, and the enzyme activity of the cellulase in the aqueous solution of pectinase is The enzyme activity of the pectinase is 300-1500 U/mL, and the enzymolysis treatment time is 15-60 minutes. The preparation method of the nano-cellulose provided by the invention adopts the aqueous solution of cellulase and the aqueous pectinase to jointly treat the biomass cellulose, which significantly improves the production efficiency and yield of the nano-cellulose and is environmentally friendly.
Description
技术领域technical field
本发明属于生物质材料技术领域，尤其涉及一种绿色高效纳米纤维素的制备方法。The invention belongs to the technical field of biomass materials, in particular to a method for preparing green and high-efficiency nano-cellulose.
背景技术Background technique
目前，纳米纤维素的制备主要采用酸法、机械法、TEMPO氧化、次氯酸钠法或生物法(含生物酶法)。酸法制备纳米纤维素利用的酸主要包括硫酸、磷酸、醋酸等，酸液反应后排放时易造成环境污染；机械法包括高压研磨、球磨、冷冻研磨、超声处理等，但是机械法制备的纳米纤维素产品得率低；TEMPO氧化或次氯酸钠法就是利用TEMPO、NaClO/NaClO2体系在酸性或碱性条件下制备纳米纤维素，与酸法相同，易造成环境污染。At present, the preparation of nanocellulose mainly adopts acid method, mechanical method, TEMPO oxidation, sodium hypochlorite method or biological method (including biological enzyme method). The acids used in the preparation of nanocellulose by the acid method mainly include sulfuric acid, phosphoric acid, acetic acid, etc., which are likely to cause environmental pollution when the acid solution is discharged after the reaction; mechanical methods include high-pressure grinding, ball milling, freeze grinding, ultrasonic treatment, etc. The yield of cellulose products is low; TEMPO oxidation or sodium hypochlorite method is to use TEMPO, NaClO/NaClO 2 system to prepare nanocellulose under acidic or alkaline conditions, which is the same as the acid method, which is easy to cause environmental pollution.
生物法制备纳米纤维素包括利用纤维素酶法、细菌降解生物质材料制备纳米纤维素及利用细菌生产纳米细菌纤维素，虽然生物法制备纳米纤维素得率高且绿色环保，但是酶解时间通常需要7～30天，纳米纤维素的生产效率低。Biological preparation of nanocellulose includes the use of cellulase, bacterial degradation of biomass materials to prepare nanocellulose and the use of bacteria to produce nano-bacterial cellulose. Although the biological preparation of nanocellulose has a high yield and is environmentally friendly, the enzymatic hydrolysis time is generally It takes 7 to 30 days, and the production efficiency of nanocellulose is low.
发明内容Contents of the invention
有鉴于此，本发明提供了一种绿色高效纳米纤维素的制备方法，本发明提供的制备方法显著提高了纳米纤维素的生产效率。In view of this, the present invention provides a method for preparing green and high-efficiency nanocellulose. The preparation method provided by the present invention significantly improves the production efficiency of nanocellulose.
为了实现上述发明目的，本发明提供以下技术方案：In order to achieve the above-mentioned purpose of the invention, the present invention provides the following technical solutions:
本发明提供了一种绿色高效纳米纤维素的制备方法，包括以下步骤：The invention provides a method for preparing green and high-efficiency nanocellulose, comprising the following steps:
将生物质纤维素、纤维素酶水溶液、果胶酶水溶液混合进行酶解处理，得到所述纳米纤维素；mixing biomass cellulose, cellulase aqueous solution, and pectinase aqueous solution to perform enzymolysis treatment to obtain the nanocellulose;
当所述纤维素酶水溶液和所述果胶酶水溶液的总质量与所述生物质纤维素的质量比为(15～30):1，所述纤维素酶水溶液中纤维素酶的酶活力为30～150U/mL，所述果胶酶水溶液中果胶酶的酶活力为300～1500U/mL，所述酶解处理的时间为15～60min。When the mass ratio of the total mass of the cellulase aqueous solution and the pectinase aqueous solution to the biomass cellulose is (15-30): 1, the enzyme activity of cellulase in the cellulase aqueous solution is 30-150 U/mL, the enzyme activity of pectinase in the pectinase aqueous solution is 300-1500 U/mL, and the enzymatic hydrolysis treatment time is 15-60 min.
优选的，所述纤维素酶水溶液和果胶酶水溶液的质量比为(1～4):1。Preferably, the mass ratio of the cellulase aqueous solution to the pectinase aqueous solution is (1-4):1.
优选的，所述生物质纤维素中纤维素的质量含量≥98％。Preferably, the mass content of cellulose in the biomass cellulose is ≥98%.
优选的，所述酶解处理的温度为45～55℃。Preferably, the temperature of the enzymatic hydrolysis treatment is 45-55°C.
优选的，所述酶解处理在搅拌的条件下进行，所述搅拌的转速为800～1200r/min。Preferably, the enzymatic hydrolysis treatment is carried out under the condition of stirring, and the rotating speed of the stirring is 800-1200 r/min.
优选的，所述酶解处理得到纳米纤维素的胶体溶液，所述酶解处理后还包括，将所述纳米纤维素的胶体溶液进行冷冻干燥，得到纳米纤维素固体粉末，所述冷冻干燥的温度为-70～-85℃，所述冷冻干燥的时间为24～48h。Preferably, the enzymolysis treatment obtains a colloidal solution of nanocellulose, and after the enzymolysis treatment, it also includes freeze-drying the colloidal solution of nanocellulose to obtain a solid powder of nanocellulose, and the freeze-dried The temperature is -70~-85°C, and the freeze-drying time is 24~48h.
优选的，所述冷冻干燥为真空冷冻干燥，所述真空冷冻干燥的真空度为0.08～0.1Pa。Preferably, the freeze-drying is vacuum freeze-drying, and the vacuum degree of the vacuum freeze-drying is 0.08-0.1Pa.
优选的，所述纳米纤维的胶体溶液的固含量为0.12～0.46％。Preferably, the solid content of the nanofiber colloid solution is 0.12-0.46%.
本发明提供了一种绿色高效纳米纤维素的制备方法，包括以下步骤：将生物质纤维素、纤维素酶水溶液和果胶酶水溶液混合进行酶解处理，得到所述纳米纤维素；当所述纤维素酶水溶液和所述果胶酶水溶液的总质量与所述生物质纤维素的质量比为(15～30):1，所述纤维素酶水溶液中纤维素酶的酶活力为30～150U/mL，所述果胶酶水溶液中果胶酶的酶活力为300～1500U/mL，所述酶解处理的时间为15～60min。本发明提供的制备方法，相较于单一的纤维素酶进行酶解反应，采用纤维素酶水溶液和果胶酶水溶液共同酶解生物质纤维素，利用纤维素酶水溶液和果胶酶水溶液的协同作用，当所述纤维素酶水溶液和所述果胶酶水溶液的总质量与所述生物质纤维素的质量比为(15～30):1时，且所述纤维素酶水溶液中纤维素酶的酶活力为30～150U/mL，所述果胶酶水溶液中果胶酶的酶活力为300～1500U/mL时，本发明酶解处理的时间仅为15～60min，从而显著提高了纳米纤维素的生产效率。The invention provides a method for preparing green and high-efficiency nanocellulose, comprising the following steps: mixing biomass cellulose, cellulase aqueous solution and pectinase aqueous solution to perform enzymolysis treatment to obtain the nanocellulose; when the The mass ratio of the total mass of the cellulase aqueous solution and the pectinase aqueous solution to the biomass cellulose is (15-30):1, and the enzyme activity of the cellulase in the cellulase aqueous solution is 30-150U /mL, the enzyme activity of pectinase in the pectinase aqueous solution is 300～1500U/mL, and the time of the enzymolysis treatment is 15～60min. In the preparation method provided by the invention, compared with a single cellulase enzymatic hydrolysis reaction, the biomass cellulose is jointly enzymolyzed by the cellulase aqueous solution and the pectinase aqueous solution, and the synergy of the cellulase aqueous solution and the pectinase aqueous solution is utilized. Effect, when the mass ratio of the total mass of the cellulase aqueous solution and the pectinase aqueous solution to the biomass cellulose is (15-30): 1, and the cellulase in the cellulase aqueous solution The enzyme activity of pectinase is 30～150U/mL, when the enzyme activity of pectinase in the pectinase aqueous solution is 300～1500U/mL, the time of enzymolysis treatment of the present invention is only 15～60min, thereby significantly improved nanofiber prime production efficiency.
附图说明Description of drawings
图1为本发明实施例1制备的纳米纤维素的电镜照片；Fig. 1 is the electron micrograph of the nanocellulose prepared by the embodiment of the present invention 1;
图2为本发明实施例2制备的纳米纤维素的电镜照片。Fig. 2 is an electron micrograph of nanocellulose prepared in Example 2 of the present invention.
具体实施方式Detailed ways
本发明提供了一种绿色高效纳米纤维素的制备方法，包括以下步骤：The invention provides a method for preparing green and high-efficiency nanocellulose, comprising the following steps:
将生物质纤维素、纤维素酶水溶液和果胶酶水溶液混合进行酶解处理，得到所述纳米纤维素；mixing biomass cellulose, cellulase aqueous solution and pectinase aqueous solution to perform enzymatic hydrolysis treatment to obtain the nanocellulose;
当所述纤维素酶水溶液和所述果胶酶水溶液的总质量与所述生物质纤维素的质量比为(15～30):1，所述纤维素酶水溶液中纤维素酶的酶活力为30～150U/mL，所述果胶酶水溶液中果胶酶的酶活力为300～1500U/mL，所述酶解处理的时间为15～60min。When the mass ratio of the total mass of the cellulase aqueous solution and the pectinase aqueous solution to the biomass cellulose is (15-30): 1, the enzyme activity of cellulase in the cellulase aqueous solution is 30-150 U/mL, the enzyme activity of pectinase in the pectinase aqueous solution is 300-1500 U/mL, and the enzymatic hydrolysis treatment time is 15-60 min.
在本发明中，所述生物质纤维素优选为木材纤维素、秸秆纤维素、竹材纤维素或叶片纤维素，所述叶片纤维素优选为杨树叶纤维素、槐树叶纤维素或竹叶纤维素。在本发明中，所述生物质纤维素中纤维素的质量含量优选≥98％。In the present invention, the biomass cellulose is preferably wood cellulose, straw cellulose, bamboo cellulose or leaf cellulose, and the leaf cellulose is preferably poplar leaf cellulose, pagoda tree leaf cellulose or bamboo leaf cellulose . In the present invention, the mass content of cellulose in the biomass cellulose is preferably ≥98%.
在本发明中，所述生物质纤维素的制备方法优选包括以下步骤：In the present invention, the preparation method of the biomass cellulose preferably comprises the following steps:
将生物质材料依次进行去除木质素和去除半纤维素，得到所述生物质纤维素。The biomass material is sequentially subjected to lignin removal and hemicellulose removal to obtain the biomass cellulose.
在本发明中，所述生物质材料优选为木材、秸秆、竹材或植物叶片，所述植物叶片优选为杨树叶、槐树叶或竹叶。本发明优选对所述生物质材料进行前处理，在本发明中，所述前处理优选包括：去除所述生物质材料中的油脂；在本发明中，所述去除生物质材料中油脂优选为：将所述生物质材料浸渍(以下称为第一浸渍)于极性溶剂中，去除所述生物质材料中的油脂；在本发明中，所述极性溶剂优选为无水乙醇或乙醇和水的混合溶剂，在本发明中，所述乙醇和水的混合溶剂中乙醇的质量百分含量优选≥95％；本发明对所述极性溶剂的用量没有特殊要求，能够将所述生物质材料完全浸没即可；在本发明中，所述第一浸渍的温度优选为65～70℃，所述第一浸渍的时间优选为6～8h。In the present invention, the biomass material is preferably wood, straw, bamboo or plant leaves, and the plant leaves are preferably poplar leaves, pagoda tree leaves or bamboo leaves. In the present invention, the biomass material is preferably pretreated. In the present invention, the pretreatment preferably includes: removing the grease in the biomass material; in the present invention, the removal of grease in the biomass material is preferably : the biomass material is immersed (hereinafter referred to as the first immersion) in a polar solvent to remove the grease in the biomass material; in the present invention, the polar solvent is preferably dehydrated alcohol or ethanol and A mixed solvent of water, in the present invention, the mass percent content of ethanol in the mixed solvent of ethanol and water is preferably ≥ 95%; It only needs to immerse the material completely; in the present invention, the temperature of the first immersion is preferably 65-70° C., and the time of the first immersion is preferably 6-8 hours.
本发明优选对所述前处理后的生物质材料进行水洗，本发明对所述水洗的具体实施过程没有特殊要求，本发明优选通过水洗去除所述生物质材料表面残留的极性溶剂。In the present invention, the pretreated biomass material is preferably washed with water. The present invention has no special requirements for the specific implementation process of the water washing. In the present invention, the residual polar solvent on the surface of the biomass material is preferably removed by water washing.
本发明将所述生物质材料进行去除木质素；在本发明中，所述去除木质素优选为：将所述生物质材料浸渍于酸性亚氯酸钠溶液中(以下称为第二浸渍)，进行所述去除木质素，本发明对所述酸性亚氯酸钠溶液的用量没有特殊要求，能够将所述生物质材料完全浸没即可。本发明采用酸性亚氯酸钠溶液去除所述生物质材料中的木质素时，所述酸性亚氯酸钠溶液的处理次数优选为3～5次，在本发明中，每一次酸性亚氯酸钠溶液处理的时间优选为1h，在本发明的具体实施例中，所述酸性亚氯酸钠溶液的处理次数为5次，第1次处理时，所述酸性亚氯酸钠溶液的质量百分含量优选为0.67％，第2次处理时，所述酸性亚氯酸钠溶液的质量百分含量优选为1.34％，第3次处理时，所述酸性亚氯酸钠溶液的质量百分含量优选为2.01％，第4次处理时，所述酸性亚氯酸钠溶液的质量百分含量优选为2.68％，第5次处理时，所述酸性亚氯酸钠溶液的质量百分含量优选为3.35％。在本发明中，所述第二浸渍的温度优选为75～80℃。在本发明中，酸性亚氯酸钠溶液的pH值优选为3～4，所述酸性亚氯酸钠的pH值优选通过pH调节剂进行调节，在本发明中，所述pH调节剂优选为冰醋酸。In the present invention, the biomass material is subjected to lignin removal; in the present invention, the lignin removal is preferably: immersing the biomass material in an acidic sodium chlorite solution (hereinafter referred to as the second immersion), For the removal of lignin, the present invention has no special requirements on the amount of the acidic sodium chlorite solution, as long as the biomass material can be completely submerged. When the present invention uses acidic sodium chlorite solution to remove lignin in the biomass material, the number of times of treatment of the acidic sodium chlorite solution is preferably 3 to 5 times. In the present invention, each acidic chlorous acid The time of sodium solution processing is preferably 1h, and in the specific embodiment of the present invention, the processing frequency of described acidic sodium chlorite solution is 5 times, and during the 1st treatment, the mass of described acidic sodium chlorite solution Content is preferably 0.67%, and during the 2nd time processing, the mass percentage composition of described acidic sodium chlorite solution is preferably 1.34%, during the 3rd time processing, the mass percentage composition of described acidic sodium chlorite solution It is preferably 2.01%, and during the 4th treatment, the mass percentage of the acidic sodium chlorite solution is preferably 2.68%, and during the 5th treatment, the mass percentage of the acidic sodium chlorite solution is preferably 3.35%. In the present invention, the temperature of the second immersion is preferably 75-80°C. In the present invention, the pH value of the acidic sodium chlorite solution is preferably 3 to 4, and the pH value of the acidic sodium chlorite is preferably adjusted by a pH regulator. In the present invention, the pH regulator is preferably glacial acetic acid.
本发明优选对所述除去木质素后的生物质材料进行水洗，本发明对所述水洗的具体实施过程没有特殊要求，本发明通过水洗去除所述生物质材料表面残留的酸性亚氯酸钠。In the present invention, the biomass material after lignin removal is preferably washed with water. The present invention has no special requirements for the specific implementation process of the water washing. The present invention removes the residual acidic sodium chlorite on the surface of the biomass material through water washing.
去除所述木质素后，本发明将所述生物质材料进行去除半纤维素；在本发明中，所述去除半纤维素优选为：将所述除去木质素的生物质材料浸渍于强碱溶液中(以下称为第三浸渍)，进行所述去除半纤维素，本发明对所述强碱溶液的用量没有特殊要求，能够将所述生物质材料完全浸没即可；在本发明中，所述强碱溶液优选为碱金属氢氧化物溶液，更优选为氢氧化钠溶液，在本发明中，所述氢氧化钠溶液的质量浓度优选为15～20％，更优选为17.5％；在本发明中，所述第三浸渍的温度优选为20～25℃，所述第三浸渍的时间优选为0.5～2h。After the lignin is removed, the present invention removes the hemicellulose from the biomass material; in the present invention, the hemicellulose removal is preferably: immersing the lignin-removed biomass material in a strong alkaline solution (hereinafter referred to as the third impregnation), carry out the removal of hemicellulose, the present invention has no special requirements on the amount of the strong alkali solution, as long as the biomass material can be completely submerged; in the present invention, the The strong base solution is preferably an alkali metal hydroxide solution, more preferably a sodium hydroxide solution, and in the present invention, the mass concentration of the sodium hydroxide solution is preferably 15 to 20%, more preferably 17.5%; In the invention, the temperature of the third immersion is preferably 20-25°C, and the time of the third immersion is preferably 0.5-2h.
本发明优选对所述去除半纤维素后的生物质材料进行水洗和干燥，本发明对所述水洗的具体实施过程没有特殊要求，本发明优选通过水洗去除所述生物质材料表面残留的强碱溶液；本发明对所述干燥的具体实施过程没有特殊要求；在本发明中，所述干燥后生物质纤维素的含水率优选≤10％。The present invention preferably washes and dries the biomass material after the removal of hemicellulose. The present invention has no special requirements for the specific implementation process of the water washing. The present invention preferably removes the residual strong alkali on the surface of the biomass material by washing. solution; the present invention has no special requirements on the specific implementation process of the drying; in the present invention, the moisture content of the dried biomass cellulose is preferably ≤10%.
本发明对所述纤维素酶和果胶酶的来源没有特殊要求，在本明发明中，所述纤维素酶和果胶酶购买自上海阿拉丁。The present invention has no special requirements on the sources of the cellulase and pectinase, and in the present invention, the cellulase and pectinase are purchased from Shanghai Aladdin.
在本发明的实施例中，所述水优选为去离子水或蒸馏水。In an embodiment of the present invention, the water is preferably deionized water or distilled water.
在本发明中，所述纤维素酶水溶液和果胶酶水溶液的质量比优选为(1～4):1，更优选为(1.5～3.5):1。In the present invention, the mass ratio of the aqueous cellulase solution to the aqueous pectinase solution is preferably (1-4):1, more preferably (1.5-3.5):1.
在本发明中，所述纤维素酶水溶液和果胶酶水溶液的总质量与所述生物质纤维素的质量比优选为(15～30):1，更优选为(20～25):1。In the present invention, the mass ratio of the total mass of the cellulase aqueous solution and the pectinase aqueous solution to the biomass cellulose is preferably (15-30):1, more preferably (20-25):1.
在本发明中，所述纤维素酶水溶液中纤维素酶的酶活力为30～150U/mL，优选为45～120U/mL。In the present invention, the enzyme activity of cellulase in the aqueous solution of cellulase is 30-150 U/mL, preferably 45-120 U/mL.
在本发明中，所述果胶酶水溶液中果胶酶的酶活力为300～1500U/mL，优选为375～1200U/mL。In the present invention, the enzyme activity of pectinase in the pectinase aqueous solution is 300-1500 U/mL, preferably 375-1200 U/mL.
在本发明的具体实施例中，所述纤维素酶水溶液中纤维素酶的酶活力优选为150U/mL时，所述果胶酶水溶液中果胶酶的酶活力优选为1500U/mL。In a specific embodiment of the present invention, when the enzyme activity of cellulase in the aqueous solution of cellulase is preferably 150 U/mL, the enzyme activity of pectinase in the aqueous solution of pectinase is preferably 1500 U/mL.
本发明对所述第一混合的具体实施过程没有特殊要求。The present invention has no special requirements on the specific implementation process of the first mixing.
在本发明中，所述酶解处理的温度优选为45～55℃，更优选为50℃；所述纤维素酶水溶液和果胶酶水溶液的总质量与所述生物质纤维素的质量比为(15～30):1时，所述酶解处理的时间为15～60min，优选为30～45min；所述酶解处理优选在搅拌的条件下进行，所述搅拌的转速优选为800～1200r/min，更优选为850～1000r/min。本发明所述搅拌的具体实施过程没有特殊要求。In the present invention, the temperature of the enzymolysis treatment is preferably 45-55°C, more preferably 50°C; the mass ratio of the total mass of the cellulase aqueous solution and the pectinase aqueous solution to the biomass cellulose is (15～30): 1 hour, the time of described enzymolysis treatment is 15～60min, preferably 30～45min; Described enzymolysis treatment is preferably carried out under the condition of stirring, and the rotating speed of described stirring is preferably 800～1200r /min, more preferably 850 to 1000r/min. The specific implementation process of the stirring described in the present invention has no special requirements.
本发明进行所述酶解处理时，所述纤维素酶和果胶酶能够产生协同作用。When performing the enzymatic hydrolysis treatment in the present invention, the cellulase and pectinase can produce a synergistic effect.
所述酶解处理后，本发明优选将得到的纳米纤维素胶体溶液进行干燥。After the enzymatic treatment, the present invention preferably dries the obtained nanocellulose colloidal solution.
本发明优选对所述酶解处理后的酶解反应液进行后处理，得到所述纳米纤维素的胶体溶液；在本发明中，所述后处理优选包括：依次进行生物酶灭活、固液分离和水洗；在本发明中，所述生物酶灭活优选为：将所述酶解反应液进行高温处理；所述高温处理的温度优选为100～110℃，所述高温处理的时间优选为10～15min。在本发明中，所述固液分离优选为离心分离，所述离心分离的转速优选为3500～5000r/min，所述离心分离的时间优选为10～15min，所述离心分离得到上清液和固体产物。本发明优选对所述固体产物进行水洗，所述水洗的次数优选为3～5次，本发明对每次水洗时的用水量没有特殊要求，能够将所述固体产物分散得到悬浊液即可；本发明优选对水洗得到的悬浊液进行固液分离，在本发明中，当所述固液分离后得到上清液和固体产物，则继续进行水洗和固液分离，当所述固液分离后得到所述纳米纤维素的胶体溶液，无固体沉淀时则水洗结束。In the present invention, the enzymolysis reaction solution after the enzymolysis treatment is preferably post-treated to obtain the colloidal solution of the nanocellulose; in the present invention, the post-treatment preferably includes: sequentially performing biological enzyme inactivation, solid-liquid Separation and washing; in the present invention, the biological enzyme inactivation is preferably: subjecting the enzymolysis reaction solution to high temperature treatment; the temperature of the high temperature treatment is preferably 100-110°C, and the time of the high temperature treatment is preferably 10~15min. In the present invention, the solid-liquid separation is preferably centrifugation, the rotational speed of the centrifugation is preferably 3500-5000r/min, the time of the centrifugation is preferably 10-15min, and the centrifugation obtains the supernatant and solid product. In the present invention, the solid product is preferably washed with water, and the number of times of the water washing is preferably 3 to 5 times. The present invention has no special requirements on the water consumption for each washing, as long as the solid product can be dispersed to obtain a suspension The present invention preferably carries out solid-liquid separation to the suspension that washing obtains, and in the present invention, after described solid-liquid separation obtains supernatant liquid and solid product, then continue washing and solid-liquid separation, when described solid-liquid The colloidal solution of the nanocellulose is obtained after separation, and the water washing ends when there is no solid precipitation.
本发明通过固液分离和水洗，除去所述纳米纤维素中的糖分物质。The invention removes sugar substances in the nanocellulose through solid-liquid separation and water washing.
在本发明中，所述纳米纤维素的胶体溶液的固含量优选为0.13～0.46％；所述纳米纤维素的胶体溶液的残渣率优选为0％；所述纳米纤维素的胶体溶液放置1个月不产生絮状物或沉淀。In the present invention, the solid content of the colloidal solution of nanocellulose is preferably 0.13 to 0.46%; the residue rate of the colloidal solution of nanocellulose is preferably 0%; the colloidal solution of nanocellulose is placed in a Monthly does not produce flocs or sediment.
在本发明中，所述干燥的温度优选为-70～-85℃，所述干燥的时间优选为24～48h，更优选为30～40h；在本发明中，所述干燥在真空环境中进行，所述真空环境的真空度优选为0.08～0.1Pa。In the present invention, the drying temperature is preferably -70 to -85°C, and the drying time is preferably 24 to 48 hours, more preferably 30 to 40 hours; in the present invention, the drying is carried out in a vacuum environment , the vacuum degree of the vacuum environment is preferably 0.08-0.1Pa.
本发明通过干燥得到纳米纤维素粉状固体。在本发明中，所述纳米纤维素的直径优选为10～50nm，所述纳米纤维素的得率大于85％，相对结晶度优选为72～82％。The invention obtains the nano cellulose powdery solid by drying. In the present invention, the diameter of the nanocellulose is preferably 10-50 nm, the yield of the nanocellulose is greater than 85%, and the relative crystallinity is preferably 72-82%.
本发明提供的制备方法能够快速、高效的制备得到纳米纤维素产品，产量高具有绿色无污染，对环境无害。The preparation method provided by the invention can quickly and efficiently prepare nanocellulose products, has high yield, is green and pollution-free, and is harmless to the environment.
下面将结合本发明中的实施例，对本发明中的技术方案进行清楚、完整地描述。显然，所描述的实施例仅是本发明一部分实施例，而不是全部的实施例。基于本发明中的实施例，本领域普通技术人员在没有做出创造性劳动前提下所获得的所有其他实施例，都属于本发明保护的范围。The technical solutions in the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Apparently, the described embodiments are only some of the embodiments of the present invention, but not all of them. Based on the embodiments of the present invention, all other embodiments obtained by persons of ordinary skill in the art without making creative efforts belong to the protection scope of the present invention.
实施例1Example 1
将松针粉末浸渍入95％的乙醇中，在70℃浸渍8h后去除油脂后，将松针粉末水洗烘干后再浸渍入冰醋酸和亚氯酸钠的混合溶液(pH值为3～4)中75℃浸渍5次，每次1h去除木质素；水洗烘干后浸渍入17.5％氢氧化钠溶液中20℃浸渍1h得到松针纤维素，利用红外光谱检测松针纤维素并与微晶纤维素的红外光谱进行比对，松针纤维素中纤维素的含量大于99％；Immerse the pine needle powder in 95% ethanol, remove the grease after immersing at 70°C for 8 hours, wash and dry the pine needle powder and then immerse it in the mixed solution of glacial acetic acid and sodium chlorite (pH value is 3-4) Immerse 5 times at 75°C for 1 hour each time to remove lignin; after washing and drying, immerse in 17.5% sodium hydroxide solution and immerse for 1 hour at 20°C to obtain pine needle cellulose. Compared with the spectrum, the content of cellulose in pine needle cellulose is greater than 99%;
称取5g纤维素酶和95g去离子水混合，得到纤维素酶水溶液，其中纤维素酶的活性为150u/ml；Take 5g cellulase and mix with 95g deionized water to obtain the cellulase aqueous solution, wherein the activity of cellulase is 150u/ml;
称取5g果胶酶和95g去离子水混合，得到果胶酶水溶液，其中果胶酶的活性为1500u/ml；Take by weighing 5g pectinase and mix with 95g deionized water to obtain the pectinase aqueous solution, wherein the activity of pectinase is 1500u/ml;
称取2g松针纤维素、15g果胶酶水溶液和15g纤维素酶水溶液混合后，在温度50℃酶解处理60mim，搅拌速度为1000r/min；处理时间结束后立刻将酶解反应液移入100℃水浴锅中处理10min，用于生物酶灭活；然后利用离心机离心10min，转速4000r/min，去掉上清液留下沉淀产物；用蒸馏水将沉淀产物混合均匀得到悬浊液后再次用离心机离心10min，转速4000r/min；重复上述步骤4次，即可获得离心后无固体产物的乳白色的纳米纤维素的胶体溶液，固含量为0.26％；Weigh 2g of pine needle cellulose, 15g of pectinase aqueous solution and 15g of cellulase aqueous solution and mix them, and then enzymatically treat 60mim at a temperature of 50°C, with a stirring speed of 1000r/min; Treat in a water bath for 10 minutes to inactivate biological enzymes; then use a centrifuge to centrifuge for 10 minutes at a speed of 4000r/min, remove the supernatant and leave the precipitated product; mix the precipitated product with distilled water to obtain a suspension, and use the centrifuge again Centrifuge for 10 minutes at a speed of 4000r/min; repeat the above steps 4 times to obtain a milky white nanocellulose colloid solution without solid products after centrifugation, with a solid content of 0.26%;
将纳米纤维素的胶体溶液置于真空冷冻干燥机中进行冷冻干燥，真空度为0.08Pa，温度为-80℃。时间为48h；得纳米纤维素的粉末状固体；纳米纤维素得率为85％，相对结晶度为78.46％。The colloidal solution of nanocellulose was placed in a vacuum freeze dryer for freeze-drying with a vacuum degree of 0.08 Pa and a temperature of -80°C. The time is 48 hours; the powdery solid of nanocellulose is obtained; the yield of nanocellulose is 85%, and the relative crystallinity is 78.46%.
利用场发射扫描电镜对实施例1所得的纳米纤维素粉末进行观察，所得图片见图1所示，实施例1得到的纳米纤维素的直径为18～50nm。The nanocellulose powder obtained in Example 1 was observed with a field emission scanning electron microscope, and the obtained picture is shown in FIG. 1 . The diameter of the nanocellulose obtained in Example 1 was 18-50 nm.
实施例2Example 2
将杨树叶粉末浸渍入95％的乙醇中，在70℃浸渍8h去除油脂后，将杨树叶粉末水洗烘干后再浸渍入冰醋酸和亚氯酸钠的混合溶液(pH值为3～4)中75℃浸渍5次，每次1h去除木质素；水洗烘干后浸渍入17.5％氢氧化钠溶液中20℃浸渍1h得到杨树叶纤维素，利用红外光谱检测杨树叶纤维素并与微晶纤维素的红外光谱进行比对，杨树叶纤维素中纤维素的含量大于99％；Soak poplar leaf powder in 95% ethanol, soak at 70°C for 8 hours to remove grease, wash and dry poplar leaf powder, and then dip into a mixed solution of glacial acetic acid and sodium chlorite (pH value is 3-4) Immerse 5 times at 75°C for 1 hour each time to remove lignin; after washing and drying, immerse in 17.5% sodium hydroxide solution and immerse for 1 hour at 20°C to obtain poplar leaf cellulose. Use infrared spectroscopy to detect poplar leaf cellulose and combine with microcrystalline fiber Compared with the infrared spectrum of cellulose, the content of cellulose in poplar leaf cellulose is greater than 99%;
称取5g纤维素酶和95g去离子水混合，得到纤维素酶水溶液，其中纤维素酶的活性为150u/ml；Take 5g cellulase and mix with 95g deionized water to obtain the cellulase aqueous solution, wherein the activity of cellulase is 150u/ml;
称取5g果胶酶和95g去离子水混合，得到果胶酶水溶液，其中果胶酶的活性为1500u/ml；Take by weighing 5g pectinase and mix with 95g deionized water to obtain the pectinase aqueous solution, wherein the activity of pectinase is 1500u/ml;
称取1g杨树叶纤维素、15g果胶酶水溶液和15g纤维素酶水溶液混合后，在温度50℃酶解处理60mim，搅拌速度为1000r/min；处理时间结束后立刻将酶解反应液移入100℃水浴锅中处理10min，用于生物酶灭活；然后利用离心机离心10min，转速4000r/min，去掉上清液留下沉淀产物；用蒸馏水将沉淀产物混合均匀得到悬浊液后再次用离心机离心10min，转速4000r/min；重复上述步骤4次，即可获得离心后无固体产物的乳白色的纳米纤维素的胶体溶液，固含量为0.13％；Weigh 1g of poplar leaf cellulose, 15g of pectinase aqueous solution and 15g of cellulase aqueous solution and mix, enzymatically treat 60mim at a temperature of 50°C, and stir at a speed of 1000r/min; immediately transfer the enzymolysis reaction solution into 100 ℃ in a water bath for 10 minutes to inactivate biological enzymes; then use a centrifuge for 10 minutes at a speed of 4000r/min to remove the supernatant and leave the precipitated product; mix the precipitated product with distilled water to obtain a suspension and then centrifuge again Machine centrifugation for 10min, rotating speed 4000r/min; repeat the above steps 4 times, you can obtain the colloidal solution of milky white nanocellulose without solid product after centrifugation, the solid content is 0.13%;
将纳米纤维素的胶体溶液置于真空冷冻干燥机中进行冷冻干燥，真空度为0.08Pa，温度为-80℃。时间为48h；得纳米纤维素的粉末状固体；纳米纤维素得率为85％，相对结晶度为77.39％。The colloidal solution of nanocellulose was placed in a vacuum freeze dryer for freeze-drying with a vacuum degree of 0.08 Pa and a temperature of -80°C. The time is 48 hours; the powdery solid of nanocellulose is obtained; the yield of nanocellulose is 85%, and the relative crystallinity is 77.39%.
利用场发射扫描电镜对实施例2所得的纳米纤维素粉末进行观察，所得图片见图1所示，实施例2得到的纳米纤维素的直径为10～50nm。The nanocellulose powder obtained in Example 2 was observed with a field emission scanning electron microscope, and the obtained picture is shown in FIG. 1 . The diameter of the nanocellulose obtained in Example 2 was 10-50 nm.
实施例3Example 3
将竹材粉末浸渍入95％的乙醇中，在70℃浸渍8h去除油脂后，将竹材粉末水洗烘干后再浸渍入冰醋酸和亚氯酸钠的混合溶液(pH值为3～4)中75℃浸渍5次，每次1h去除木质素；水洗烘干后浸渍入17.5％氢氧化钠溶液中20℃浸渍1h得到竹材纤维素，利用红外光谱检测竹材纤维素并与微晶纤维素的红外光谱进行比对，竹材纤维素中纤维素的含量大于99％；Immerse the bamboo powder into 95% ethanol, remove the grease after soaking at 70°C for 8 hours, wash and dry the bamboo powder, and then dip into the mixed solution of glacial acetic acid and sodium chlorite (pH value is 3-4) for 75 Soak 5 times at ℃ for 1 hour each time to remove lignin; after washing and drying, immerse in 17.5% sodium hydroxide solution and immerse for 1 hour at 20 ℃ to obtain bamboo cellulose. For comparison, the content of cellulose in bamboo cellulose is greater than 99%;
称取5g纤维素酶和95g去离子水混合，得到纤维素酶水溶液，其中纤维素酶的活性为150u/ml；Take 5g cellulase and mix with 95g deionized water to obtain the cellulase aqueous solution, wherein the activity of cellulase is 150u/ml;
称取5g果胶酶和95g去离子水混合，得到果胶酶水溶液，其中果胶酶的活性为1500u/ml；Take by weighing 5g pectinase and mix with 95g deionized water to obtain the pectinase aqueous solution, wherein the activity of pectinase is 1500u/ml;
称取5g竹材纤维素、75g果胶酶水溶液和75g纤维素酶水溶液混合后，在温度50℃酶解处理60mim，搅拌速度为1000r/min；处理时间结束后立刻将酶解反应液移入100℃水浴锅中处理10min，用于生物酶灭活；然后利用离心机离心10min，转速4000r/min，去掉上清液留下沉淀产物；用蒸馏水将沉淀产物混合均匀得到悬浊液后再次用离心机离心10min，转速4000r/min；重复上述步骤4次，即可获得离心后无固体产物的乳白色的纳米纤维素的胶体溶液，固含量为0.46％；Weigh 5g of bamboo cellulose, 75g of pectinase aqueous solution and 75g of cellulase aqueous solution and mix them, and then enzymatically treat 60mim at a temperature of 50°C, with a stirring speed of 1000r/min; immediately after the processing time, transfer the enzymolysis reaction solution to 100°C Treat in a water bath for 10 minutes to inactivate biological enzymes; then use a centrifuge to centrifuge for 10 minutes at a speed of 4000r/min, remove the supernatant and leave the precipitated product; mix the precipitated product with distilled water to obtain a suspension, and use the centrifuge again Centrifuge for 10 minutes at a speed of 4000r/min; repeat the above steps 4 times to obtain a milky white nanocellulose colloid solution without solid products after centrifugation, with a solid content of 0.46%;
将纳米纤维素的胶体溶液置于真空冷冻干燥机中进行冷冻干燥，真空度为0.08Pa，温度为-80℃。时间为48h；得纳米纤维素的粉末状固体；纳米纤维素得率为85％，相对结晶度为78.46％。The colloidal solution of nanocellulose was placed in a vacuum freeze dryer for freeze-drying with a vacuum degree of 0.08 Pa and a temperature of -80°C. The time is 48 hours; the powdery solid of nanocellulose is obtained; the yield of nanocellulose is 85%, and the relative crystallinity is 78.46%.
对比例1Comparative example 1
将竹材粉末浸渍入95％的乙醇中，在70℃浸渍8h去除油脂后，将竹材粉末水洗烘干后再浸渍入冰醋酸和次氯酸钠的混合溶液(pH值为3～4)中75℃浸渍5次，每次1h去除木质素；水洗烘干后浸渍入17.5％氢氧化钠溶液中20℃浸渍1h得到竹材纤维素，利用红外光谱检测竹材纤维素并与微晶纤维素的红外光谱进行比对，竹材纤维素中纤维素的含量大于99％；Bamboo powder is soaked in 95% ethanol, after soaking at 70°C for 8h to remove grease, then the bamboo powder is washed and dried, and then immersed in a mixed solution of glacial acetic acid and sodium hypochlorite (pH value is 3-4) and soaked at 75°C for 5 hours. times, lignin was removed for 1 hour each time; after washing and drying, soak in 17.5% sodium hydroxide solution for 1 hour at 20°C to obtain bamboo cellulose, and use infrared spectroscopy to detect bamboo cellulose and compare it with the infrared spectrum of microcrystalline cellulose , the content of cellulose in bamboo cellulose is greater than 99%;
称取5g纤维素酶和95g去离子水混合，得到纤维素酶水溶液，其中纤维素酶的活性为150u/ml；Take 5g cellulase and mix with 95g deionized water to obtain the cellulase aqueous solution, wherein the activity of cellulase is 150u/ml;
称取2g竹材纤维素和30g纤维素酶水溶液混合后，在温度50℃酶解处理60mim，搅拌速度为1000r/min；处理时间结束后立刻将酶解反应液移入100℃水浴锅中处理10min，用于生物酶灭活；然后利用离心机离心10min，转速4000r/min，去掉上清液留下沉淀产物；用蒸馏水将沉淀产物混合均匀得到悬浊液后再次用离心机离心10min，转速4000r/min；重复上述步骤4次，但发现洗涤后的体系一直有固体沉淀，不能形成纳米纤维素的胶体溶液。Weigh 2g of bamboo cellulose and 30g of cellulase aqueous solution and mix them, then enzymatically treat them at 50°C for 60mim with a stirring speed of 1000r/min; immediately after the processing time is over, transfer the enzymolysis reaction solution into a water bath at 100°C for 10min. It is used to inactivate biological enzymes; then use a centrifuge to centrifuge for 10 minutes at a speed of 4000r/min, remove the supernatant and leave the precipitated product; mix the precipitated product with distilled water to obtain a suspension, and then centrifuge for 10 minutes with a centrifuge at a speed of 4000r/min min; repeat the above steps 4 times, but it is found that the washed system always has solid precipitation, and the colloidal solution of nanocellulose cannot be formed.
对比例2Comparative example 2
将竹材粉末浸渍入95％的乙醇中，在70℃浸渍8h去除油脂后，将竹材粉末水洗烘干后再浸渍入冰醋酸和亚氯酸钠的混合溶液(pH值为3～4)中75℃浸渍5次，每次1h去除木质素；水洗烘干后浸渍入17.5％氢氧化钠溶液中20℃浸渍1h得到竹材纤维素，利用红外光谱检测竹材纤维素并与微晶纤维素的红外光谱进行比对，竹材纤维素中纤维素的含量大于99％；Immerse the bamboo powder into 95% ethanol, remove the grease after soaking at 70°C for 8 hours, wash and dry the bamboo powder, and then dip into the mixed solution of glacial acetic acid and sodium chlorite (pH value is 3-4) for 75 Soak 5 times at ℃ for 1 hour each time to remove lignin; after washing and drying, immerse in 17.5% sodium hydroxide solution and immerse for 1 hour at 20 ℃ to obtain bamboo cellulose. For comparison, the content of cellulose in bamboo cellulose is greater than 99%;
称取5g果胶酶和95g去离子水混合，得到果胶酶水溶液，其中果胶酶的活性为1500u/ml；Take by weighing 5g pectinase and mix with 95g deionized water to obtain the pectinase aqueous solution, wherein the activity of pectinase is 1500u/ml;
称取2g竹材纤维素和30g果胶酶水溶液混合后，在温度50℃酶解处理60mim，搅拌速度为1000r/min；处理时间结束后立刻将酶解反应液移入100℃水浴锅中处理10min，用于生物酶灭活；然后利用离心机离心10min，转速4000r/min，去掉上清液留下沉淀产物；用蒸馏水将沉淀产物混合均匀得到悬浊液后再次用离心机离心10min，转速4000r/min；重复上述步骤4次，但发现洗涤后的体系一直有固体沉淀，不能形成纳米纤维素的胶体溶液。After weighing 2g of bamboo cellulose and 30g of pectinase aqueous solution and mixing them, enzymatically treat 60mim at a temperature of 50°C, with a stirring speed of 1000r/min; immediately transfer the enzymolysis reaction solution into a 100°C water bath for 10min, It is used to inactivate biological enzymes; then use a centrifuge to centrifuge for 10 minutes at a speed of 4000r/min, remove the supernatant and leave the precipitated product; mix the precipitated product with distilled water to obtain a suspension, and then centrifuge for 10 minutes with a centrifuge at a speed of 4000r/min min; repeat the above steps 4 times, but it is found that the washed system always has solid precipitation, and the colloidal solution of nanocellulose cannot be formed.
以上所述仅是本发明的优选实施方式，应当指出，对于本技术领域的普通技术人员来说，在不脱离本发明原理的前提下，还可以做出若干改进和润饰，这些改进和润饰也应视为本发明的保护范围。The above is only a preferred embodiment of the present invention, it should be pointed out that, for those of ordinary skill in the art, without departing from the principle of the present invention, some improvements and modifications can also be made, and these improvements and modifications can also be made. It should be regarded as the protection scope of the present invention.
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