CN100497609C - 植物性纤维分解剂及使用它的植物性废弃物的处理法 - Google Patents
植物性纤维分解剂及使用它的植物性废弃物的处理法 Download PDFInfo
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Abstract
本发明涉及植物性纤维分解剂及使用它的植物性废弃物的处理法。本发明提供一种不含有微生物、而含有微生物产生的植物性纤维分解酶，及简便处理该废弃物的方法，解决了现有技术中难于解决的前述处理费用的问题、能量问题、环境问题及处理需要的时间等问题。
Description
技术领域
本发明涉及植物性纤维分解剂及使用它的植物性废弃物的处理法。详细地说，涉及使用从可以产生好氧性细菌纤维单胞菌属、厌氧性细菌梭菌属及/或植物性纤维分解酶的微生物中获得的植物性纤维分解酶进行的稻草和稻壳等的植物性废弃物处理法。
背景技术
稻子耕种或小麦生产、玉米和甘蔗等谷物生产、和豆类生产中，作为由排出的植物纤维形成的废弃物的处理方法，有在田地焚烧的方法，在焚烧炉中通过重油或轻油等燃料焚烧的方法，或者在植物性废弃物中混合锯屑或米糠等堆积制成肥料的方法。然而，用焚烧炉处理的方法需要大量的燃料和运输费，另外在田地焚烧的方法由于产生的煤烟引起环境污染(稻草公害)的问题，所以正在讨论要禁止焚烧处置。对植物性废弃物的堆肥化所采取的方法有，通过植物性废弃物中混合稻草或锯屑、米糠等，散布熟石灰堆积在原野中使其自然腐蚀的方法；通过在切碎植物性废弃物的破碎物中混合锯屑、稻壳、鸡粪等调节水分后，加入发酵菌或处理过的堆肥等，在容器中搅拌等进行发酵堆肥化的方法。然而，随着农业的现代化和机械化，由于植物性废弃物的堆肥化处理和达到熟化的期间长所以具有不能实施的倾向。因此，该废弃物尽管认定是有效的资源但仍需要不断地产业废弃化。
作为前述以外的植物废弃物的处理方法，有通过微生物的固定化物处理植物性废弃物的方法。植物性废弃物其结构上的原因所以难分解，其主要原因是植物性废弃物中的纤维素及/或半纤维素引起的。因此，为了处理植物性纤维有效地利用，作为第一阶段，即使部分地水解这些纤维素及/或半纤维素也是必要的。作为分解这些纤维素和/或半纤维素的方法，已知有使用由多种菌混合物形成的稻杆等的分解剂的处理方法。特开平8-157285号公报中记载了将胶质、纤维素或半纤维素中的至少一种通过霉菌类或细菌类生产的植物性纤维分解酶分解，制成液状堆肥的植物性废弃物的液状堆肥化的方法。然而，该方法为确保处理厂必须设置处理设备，需要将植物性废弃物输送到该处理厂的劳动力。特开平7-2613号公报中记载了在田地直接散布的农药用细菌制剂。但用该制剂的植物性废弃物的分解从散布在田地到其效果发现之前，特别是到植物性废弃物的腐败熟化，需要很长时间。这是因为通过增殖的微生物产生的植物性纤维分解酶植物性废弃物得到分解的缘故，即，是因为用该制剂的植物性废弃物的分解必须要求土壤中该微生物的增殖。
在日本谷仓地带东北、北海道等寒冷地区中，为了有效地利用已存在的微生物制剂和石灰、氮制剂，该制剂投入田地的作业1次不充分，必须要2次。另外，在稻子耕种中不能分解的废弃物通过植入田地后发酵产生气体，阻碍苗的固定和根的生长。
本发明的目的在于，解决现有技术中难于解决的前述处理费用的问题、能量问题、环境问题及处理需要的时间等问题，不依赖于微生物的生存条件，并且不需要输送含有植物性纤维的废弃物的植物性废弃物处理法。
发明内容
本发明通过提供不含有微生物、含有微生物产生的植物性纤维分解酶的植物性纤维分解剂，进一步，通过提供作用于植物性废弃物的该植物性纤维分解剂形成的植物性废弃物处理法，解决了前述课题。
本发明的植物性纤维分解剂可以基于以下说明的本发明的实施方式得到，另外，本发明的植物性废弃物处理法也是可以基于以下本发明的实施方式进行实施。以下，对本发明进行详细地说明。
具体实施方式
所谓的本说明书中的植物性纤维分解酶是，具有在土壤中分解植物性纤维作用的微生物产生的酶类，例如，可以举出纤维素酶、キシラ—ゼ、果胶酶、木聚糖酶、支链淀粉酶、葡糖苷酶等。本说明书中的微生物只要是产生植物性纤维分解酶的就不作特别的限制，优选至少产生纤维素酶及/或木聚糖酶的微生物。例如，属于纤维单胞菌属(Cellulomonas)、杆状菌属(Bacillus)、纤维弧菌属(Cellvibrio)、假单胞菌属(Pseudomonas)、生孢噬纤维菌(Sporocytophaga)、醋弧菌属(Acetivibrio)、梭菌(Clostridium)、Bacteriodes、丁酸弧菌属(Butyrivibrio)、密螺旋体(Treponema)、瘤胃球菌属(Ruminococcus)、链霉菌属(Streptomyces)、高温放线菌属(Thermoactinomyces)、Thermonospola属、毛壳菌属(Chaetomium)、腐质霉属(Humicola)、Myceliophthola、孢子丝菌属(Spototrichium)、踝节菌属(Talaromyces)、热子囊菌属(Thermoascus)、草根霉属(Thielavia)、支顶孢属(Acremonium)、Agricus属、链格孢属(Alternaria)、曲霉属(Aspergillus)、球二孢属(Botryodiplodia)、镰孢属(Fusarium)、耙菌属(Irpex)、漆斑菌属(Myrothecium)、链孢霉属(Neurospora)、薄膜革菌属(Pellicularia)、青霉属(Penicillium)、Pestalotiopsis属、灰侧耳菌属(Pleurotus)、多孔菌属(Polyporus)、卧孔菌属(Poria)、密孔菌属(pycnoporus)、Pyricularia属、根霉属(Rhizopus)、裂褶菌属(Schizophyllum)、小核菌属(Sclerotium)、Scytalidium属、Termitomyces属、栓菌属(Trametes)、木霉属(Trichoderma)等的微生物，可以举出产生植物性纤维分解酶的微生物。这些微生物中，优选属于纤维单胞菌属或梭菌属的微生物产生的植物性纤维分解酶，进一步优选纤维单胞菌属产生的植物性纤维分解酶。作为具体的优选种，可举纤维单胞菌sp.K32A(通商产业省工业技术院生命工学工业技术研究所中的委托号FERM BP-6766，保藏日期：1999年6月25日，分类命名：Cellulomonassp.K32A)菌株及噬纤维梭菌(Clostridiumcellulovorans)ATCC 35296菌株。从在田地具有植物性纤维分解酶功能来看，植物性纤维分解酶优选中温菌产生的。这里的中温菌是指在高温区域(55℃以上)和在低温区域(0℃以下)均不能增殖的微生物。另外，因为可以通过纤维素酶或木聚糖酶活性法测定(Walfgang，H.，Analitical Biochem.，第164卷，72～77页，1987年；Wood，T.M.等，Methods of Enzymology，第160卷，87～112页，1988年)，所以本领技术人员可以适宜选择产生植物性纤维分解酶的适当的微生物。进一步，作为本说明书中的植物性纤维分解酶也包括，通过转基因技术制作含有微生物产生的植物性纤维分解酶基因的微生物或培养细胞等的转化体，培养该转化体得到的酶。
在植物性纤维分解酶中存在从植物性纤维末端分解的外源酶和从纤维内部分解的内源酶。本发明中这两种植物性纤维分解酶都优选。作为本发明中的植物性纤维分解酶优选在10℃～60℃，更优选在15℃～55℃具有酶活性的酶。酶活性可以用本领域技术人员周知的方法测定。例如纤维素酶、キシラ—ゼ、内源葡聚糖酶活性及木聚糖酶活性等的植物性纤维分解酶也可以用前述的现有方法(Walfgang，H.，1987年Analitical Biochem.，第164卷，72页～77页；Wood，T.M.等Methods of Enzymology，第160卷，87～112页，1988年)测定。
本说明书中的植物性纤维分解剂是指含有前述植物性纤维分解酶的植物性纤维分解剂，特别优选含有纤维素酶及/或木聚糖酶中的至少一种。该分解剂的形状不作特别的限定，例如可以是液状、粉末状或颗粒状等。另外，该分解剂优选含有前述的内源型和外源型这二者的酶。植物性纤维分解剂通过培养可产生前述植物性纤维分解酶的微生物得到。该微生物的培养在使用适合各自的微生物的培养基，适合的条件下实施。它们的培养基和培养条件可以由本领域技术人员适宜选择及设定。例如，作为培养基的碳源，可以使用滤纸、纤维素粉末等各种纤维质原料，作为氮源，可以使用硫酸铵、硝酸铵、醋酸铵等胺盐，硝酸盐及蛋白胨、肉提取物、玉米浸液、玉米麸质粗粉、棉籽油、脱脂大豆等有机物。也可以添加其他的微量的无机金属类、维生素类、生长促进因子，例如添加含有硫铵、生物素的酵母提取物等。这些培养基成分只要在不阻碍微生物发育的浓度内就可以，碳源通常使用0.025～0.5重量％较合适。氮源通常使用0.05～1重量％较合适。培养基通常调整在pH6.5～8.5，灭菌后使用。培养温度的范围只要是微生物能发育的温度就可以，可以通过本领域技术人员适宜设定。液体培养，例如培养好氧性微生物时，可以使用前述培养基在通常的振摇培养或通气搅拌培养下进行。培养厌氧性微生物时，液体培养也是用通常的方法，例如通过在前述培养基上添加适当浓度的还原剂保持在低的氧化还原电位，在20～40℃振摇培养24小时～1周来进行。作为还原剂，可以使用0.03～0.05％L-半胱氨酸盐酸、0.01～0.2％巯基乙酸钠、0.001％甲醛次硫酸钠或0.1％抗坏血酸。为了得到酶，培养到作为培养基的碳源的所使用物例如破碎的滤纸消失为止。
培养结束后，由培养上清或者通过离心分离或过滤培养上清得到粗酶液，可以使用这些培养上清或粗酶液作为本发明的植物性纤维分解剂。或者，可以是通过超滤法对培养上清浓缩的浓缩物，也可以是通过适当采用硫铵盐析法、溶剂沉淀法、透析法或冷冻干燥法等常法的粗酶粉末。另外，也可以将通过在粗酶液中加入アビセル等纤维素使纤维素酶活性成分等吸附后溶出吸附的成分得到的精制酶液作为植物性分解剂使用。
本说明书中的植物性废弃物是，稻子耕种或小麦生产、玉米、甘蔗等谷物、豆类或薯类等生产中排出的含有植物性纤维的废弃物。具体来说，作为植物性废弃物，例如有稻草、稻壳、麦杆、甘蔗渣(甘蔗榨完的粕、玉米芯)及薯类的蔓等，进一步有落叶和木片等。
本说明书中的植物性废弃物处理法，除了使用前述植物纤维分解剂以外，不作特别的限制，例如，可以对农作物收获后残留在田地的植物性废弃物通过直接散布植物性纤维分解剂在野外实施。从使植物性纤维分解剂高效率起作用的观点看，在该分解剂发生作用前预先破碎植物性废弃物较好。作为植物性废弃物的破碎法，可举例如通过コンバイン收获的同时切断稻秆等的方法，使用锁锯等切断树木的方法，使用对树木薄片化的破碎装置的破碎方法，或者通过设置了粉碎器具或切断用刃的处理装置裁断和粉碎的方法。另外，植物性废弃物处理优选在20℃～50℃的温度条件下实施。在利用大棚栽培等温室下使用本发明的植物性纤维分解剂时，优选室内加温到40℃～50℃。在野外的一般田地中，也可与市售的微生物制剂、石灰氮制剂等犁入剂，有机质肥料，化学肥料，堆肥和/或土壤改良剂并用。
实施例
以下，举实施例对本发明进行说明，本发明并不限于这些实施例。
实施例1
在纤维单胞菌sp.K32A的前培养中使用50mL的L-Broth(酵母提取物5g/L，蛋白胨10g/L，氯化纳5g/L)。在该前培养基中接种该K32A的单菌落，用旋转振荡机(商品名：G10 Gyrotory shaker，New Brunswick scientific制)在37℃振荡培养(250rpm，振幅10mm)一夜，进行前培养，得到前培养液。然后，使用添加了0.05％アビセル(商品名)的1/5PYT培养基(酵母提取物1g/L，胰化蛋白胨1g/L，蛋白胨1g/L)250mL作为本培养的培养基，在该本培养培养基中接种1％前培养液，37℃下振荡培养(250rpm，振幅10mm)，进行本培养(约72小时)，直到基质アビセル消失。培养结束后，离心分离(6000rpm，10分钟)，得到培养上清。过滤灭菌培养上清，加入硫酸铵(80％)直到饱和为止，沉淀上清中的蛋白质。将沉淀的蛋白质溶解在50mM吗啉丙烷磺酸(以下，简称为MOPS)、10mM CaCl2和1mM NaN3的缓冲液，透析，将其作为粗酶液。调制的粗酶液中的蛋白质浓度为2mg/mL。
在该粗酶液5mL中加入作为基质的1g稻草或稻壳，在表1和表2记载的温度下分别反应72小时或1周。反应结束后离心(3500rpm，2000×g下15分钟)，在得到的沉淀物中加入灭菌水，用离心机(商品名：KN-70，クボタ制)离心(3500rpm，15分钟)，洗涤3次，冷冻干燥后用天平测定沉淀物的重量。将从初期重量中减去反应后重量的重量占初期重量的百分率作为除去率。结果示于表1和表2。
表1
| 20℃ | 30℃ | 40℃ | 50℃ | |
| 72小时后的稻草的分解率(％) | 5.7 | 8.0 | 12.2 | 18.1 |
| 1周后的稻草的分解率(％) | 9.8 | 14.5 | 22.1 | 32.1 |
表2
| 20℃ | 30℃ | 40℃ | 50℃ | |
| 72小时后的稻壳的分解率(％) | 3.5 | 5.8 | 7.6 | 11.3 |
| 1周后的稻壳的分解率(％) | 9.7 | 13.8 | 21.8 | 18.2 |
实施例2
在含有10mM CaCl2和1mM NaN3的50mM MOPS缓冲液(pH7.0)中加入作为基质的1％稻壳50mL中，作为蛋白质质量，加入实施例1得到的K32A的粗酶液1mg，在50℃下保温24小时。保温后，离心分离，冷冻干燥沉淀成分，将干燥重量与对照组(没有加粗酶液)进行比较。其结果，加入粗酶液的重量与对照组比较减少了30～32％。通过目视观察离心分离前的溶液时，观察到稻壳一部分溶解。
实施例3
在添加0.1％磷酸处理过的アビセル厌氧调制的1/5PTY培养基250mL中接种噬纤维梭菌(ATCC 35296)的单菌落，在37℃下振荡培养(250rpm，振幅10mm)，培养约72小时。培养结束后，离心分离(6000rpm，10分钟)，得到培养上清。然后，过滤灭菌培养上清，加入硫酸铵(80％)直到饱和为止，沉淀上清中的蛋白质。将沉淀的蛋白质溶解在50mM磷酸-12mM柠檬酸缓冲液中，透析，将其作为粗酶液。调制的粗酶液中的蛋白质浓度为2mg/mL。
在该粗酶液5mL中加入基质1g稻草或稻壳，在表3和表4记载的温度下分别反应72小时或1周。反应结束后离心(3500rpm，2000×g下15分钟)，在得到的沉淀物中加入灭菌水，用离心机(商品名：KN-70，クボタ制)离心(3500rpm，15分钟)，洗涤3次，冷冻干燥后用天平测定沉淀物的重量。将从初期重量中减去反应后重量的重量占初期重量的百分率作为除去率。结果示于表3和表4。
表3
| 20℃ | 30℃ | 40℃ | 50℃ | |
| 72小时后的稻草的分解率(％) | 1.1 | 3.2 | 7.8 | 10.5 |
| 1周后的稻草的分解率(％) | 2.6 | 8.2 | 10.5 | 15.6 |
表4
| 20℃ | 30℃ | 40℃ | 50℃ | |
| 72小时后的稻壳的分解率(％) | 0.1 | 1.2 | 2.8 | 6.5 |
| 1周后的稻壳的分解率(％) | 1.2 | 2.0 | 5.6 | 8.8 |
本发明提供一种用比现有技术少的劳动力处理植物性废弃物的有效的植物性废弃物处理法。采用本发明，可以在田地等野外处理植物性废弃物，可以解决现有技术中难于解决的前述处理费用的问题、能量问题、环境问题及处理需要的时间等问题，不依赖于微生物的生存条件处理植物性纤维废弃物。
Claims (5)
1.在田地中使用的植物性纤维分解剂，所述植物性纤维分解剂中不含有微生物、含有微生物产生的植物性纤维分解酶，
前述微生物是纤维单胞菌属(Cellulomonas sp.)K32A菌株，保藏编号为FERM BP-6766。
2.植物性废弃物处理法，其特征在于，所述方法包括将含有微生物产生的植物性纤维分解酶的权利要求1所述的植物性纤维分解剂在田地中作用于植物性废弃物。
3.权利要求2所述的处理法，其中，植物性废弃物选自稻草、稻壳、麦杆、甘蔗渣、薯类的蔓、落叶或木片。
4.权利要求2所述的处理法，其包括在前述植物性纤维分解剂作用前破碎植物性废弃物的工序。
5.权利要求2所述的处理法，其在20℃～50℃的温度条件下实施。
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| CNA018218717A Division CN1486364A (zh) | 2001-01-10 | 2001-12-28 | 植物性纤维分解剂及使用它的植物性废弃物的处理法 |
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| CNA018218717A Pending CN1486364A (zh) | 2001-01-10 | 2001-12-28 | 植物性纤维分解剂及使用它的植物性废弃物的处理法 |
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| US (1) | US7160714B2 (zh) |
| JP (1) | JP4077158B2 (zh) |
| CN (2) | CN100497609C (zh) |
| AU (1) | AU2002219550B2 (zh) |
| WO (1) | WO2002055686A1 (zh) |
Cited By (1)
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|---|---|---|---|---|
| TWI420010B (zh) * | 2010-09-15 | 2013-12-21 | Yuen Foong Yu Paper Mfg Co Ltd | 製造非木材纖維漿料之方法 |
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| ES2259542B1 (es) * | 2005-01-28 | 2007-10-01 | Universidad De Sevilla | Procedimiento para la obtencion de bioestimulantes a partir de residuos agroindustriales. |
| US20150328347A1 (en) | 2005-03-24 | 2015-11-19 | Xyleco, Inc. | Fibrous materials and composites |
| US7708214B2 (en) * | 2005-08-24 | 2010-05-04 | Xyleco, Inc. | Fibrous materials and composites |
| MX373998B (es) | 2006-03-24 | 2020-07-22 | Childrens Medical Center | Uso de un modulador de células madre hematopoyeticas (hsc) para promover el crecimiento de hsc. |
| US8007558B2 (en) * | 2006-06-13 | 2011-08-30 | Maxeco Organo-Mineral (Canada) Inc. | Organo-mineral soil amendment |
| US9079786B2 (en) * | 2006-06-20 | 2015-07-14 | Johannes van Leeuwen | Purification of thin stillage from dry-grind corn milling with fungi |
| US20080302151A1 (en) * | 2007-06-11 | 2008-12-11 | Timothy Allan Stemwedel | Soluble Liquid Fertilizer for Organic Agriculture Derived From Soy Meal |
| US8481295B2 (en) * | 2007-06-20 | 2013-07-09 | Johannes van Leeuwen | Fungi cultivation on alcohol fermentation stillage for useful products and energy savings |
| CA2743833C (en) | 2011-04-15 | 2017-11-21 | 2245396 Ontario Inc. | Food waste management system |
| WO2013019857A2 (en) | 2011-08-01 | 2013-02-07 | Alnylam Pharmaceuticals, Inc. | Method for improving the success rate of hematopoietic stem cell transplants |
| CN102941210B (zh) * | 2012-11-12 | 2014-10-29 | 熊鹏 | 利用纤维素酶结合机械压榨处理植物性废渣的方法 |
| US11065656B2 (en) * | 2016-06-27 | 2021-07-20 | Shinko Tecnos Co., Ltd. | Method and apparatus for producing a product |
| CN107540850B (zh) * | 2016-07-28 | 2020-02-07 | 广西科技师范学院 | 一种植物纤维微波液化方法 |
| CN106986670A (zh) * | 2017-05-19 | 2017-07-28 | 上海绿博生物科技发展有限公司 | 一种促进还田水稻秸秆腐熟的生物制剂及其制备方法 |
| US20190202752A1 (en) * | 2018-01-04 | 2019-07-04 | EnviroPure Systems, Inc | Biological waste management systems |
| US20190202712A1 (en) * | 2018-01-04 | 2019-07-04 | EnviroPure Systems, Inc | Biological waste management systems |
| US20210355432A1 (en) * | 2018-10-18 | 2021-11-18 | Idemitsu Kosan Co., Ltd. | Plant residue-decomposing agent using liquid culture of bacillus pumilus ks-c4 strain |
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- 2001-12-28 CN CNB2006100719208A patent/CN100497609C/zh not_active Expired - Lifetime
- 2001-12-28 CN CNA018218717A patent/CN1486364A/zh active Pending
- 2001-12-28 AU AU2002219550A patent/AU2002219550B2/en not_active Ceased
- 2001-12-28 WO PCT/JP2001/011619 patent/WO2002055686A1/ja not_active Ceased
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| Publication number | Priority date | Publication date | Assignee | Title |
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| TWI420010B (zh) * | 2010-09-15 | 2013-12-21 | Yuen Foong Yu Paper Mfg Co Ltd | 製造非木材纖維漿料之方法 |
Also Published As
| Publication number | Publication date |
|---|---|
| JP4077158B2 (ja) | 2008-04-16 |
| CN1486364A (zh) | 2004-03-31 |
| JP2002204692A (ja) | 2002-07-23 |
| WO2002055686A1 (en) | 2002-07-18 |
| CN101029305A (zh) | 2007-09-05 |
| US7160714B2 (en) | 2007-01-09 |
| AU2002219550B2 (en) | 2007-08-23 |
| US20040033290A1 (en) | 2004-02-19 |
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