CN1372469A - 免疫激活剂 - Google Patents
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Abstract
本发明涉及具有激活机体免疫力作用的口服免疫激活剂。该免疫激活剂含有得自玉米外壳的半纤维素和/或其部分分解产物。上述半纤维素以从玉米外壳除去淀粉和蛋白质的剩余部分用碱提取的所得物为佳。上述半纤维素部分分解产物中较佳的是用木聚糖酶处理从玉米外壳除去淀粉和蛋白质的剩余部分用碱提取所得的半纤维素后所得的物质,其质量平均分子量以2万－20万为佳。
Description
技术领域
本发明涉及以玉米外壳为原料的免疫激活剂。
背景技术
免疫系统是排除外来病毒和微生物等的侵入和癌细胞等异物，维持机体平衡的重要的机体防御功能之一。因衰老、应激、疲劳和环境因素等因素而免疫功能降低时，容易引起各种疾病和感染等。
因此，加强作为机体防御机能的免疫力在维持机体平衡上是重要的，激活免疫系统的免疫激活剂作为食品和药品具有广泛的用途。
迄今提出了各种各样免疫活化物质及其制备方法，已知的米糠半纤维素部分分解产物是其中之一。
例如，专利公报2532899号(特开平1-153701号公报)中记载了具有免疫激活作用的半纤维素的制备方法，它是将从米糠碱提取所得的半纤维素用丝状真菌分解而得到的；特开平9-23895号公报中记载了制备植物组织水溶性多糖体后以状真菌的菌体外酶反应而生成免疫增强物质的方法。
然而，用上述制备方法所得的来自米糠的具有免疫激活作用的半纤维素和免疫力增强物质的免疫激活效果仅在体外测定了天然杀伤细胞(NK-细胞)的增加、活化效果，而实际上经口摄取时是否具有免疫激活作用尚不清楚。
另外，特别是前一制备方法，难以得到质量稳定的反应生成物，而且丝状真菌的培养耗时15天，在工业规模的制备上不能说是满意的方法。
因此，迄今未知从玉米外壳制备的半纤维素及其部分分解产物是否具有免疫激活作用。
发明的内容
因此，本发明的目的在于提供来自玉米外壳的通过口服具有激活机体免疫力作用的免疫激活剂。
本发明者为了达到上述目的进行锐意研究的结果，发现从玉米外壳制备的半纤维素及其部分分解产物具有免疫激活作用，从而完成了本发明。
按照本发明，可提供安全地、可经口服激活免疫力的免疫激活剂。
实施发明的最佳状态
本发明的免疫激活剂有效成分、得自玉米外壳的半纤维素及其部分分解产物的制备方法无特别限制，可采用公知的方法。
例如，从玉米外壳提取半纤维素的方法可采用碱或酸提取，用挤压机或高压釜加压、热水提取，用纤维素酸等酶制剂分解、抽提、或其适当组合的方法。本发明中，为了得到较高纯度的半纤维素，较理想的是将从玉米外壳除去淀粉、蛋白质，根据需要进而除去脂类、无机物等的剩余部分用碱提取，制成的半纤维素。
从玉米外壳除去淀粉、蛋白质，根据需要进而除去脂类、无机物等的方法，可采用酶处理、化学处理或物理处理，或者将它们作适当的组合。酶处理可在pH 3-9、温度30-100℃的条件下添加例如α-淀粉酶、葡糖淀粉酶等淀粉分解酶、蛋白水解酶等蛋白分解酶脂肪水解酶等脂肪分解酶、纤维素酶等纤维素分解酶，使其作用进行处理。而化学处理，可例如在玉米外壳中添加无机酸、有机酸的水溶液，在pH 2-5的条件下加热，或添加食品加工用表面活性剂，在pH 3-8的条件下进行热处理。物理处理则将例如玉米外壳用匀浆器、锤磨机等粉碎机粉碎后进行筛选。
半纤维素部分分解(部分水解)的方法较佳为采用将如上所述制备的半纤维素用纤维素分解酶(特别好的是木聚糖酶)进行处理的方法。
本发明中所用的半纤维素可按如下方法制备。
在上述预处理过的玉米外壳5-20重量份中加入水80-95重量份，添加碱化合物，调节为pH 10-13，于80-140℃搅拌0.5-10小时搅拌混合，提取半纤维素。碱化合物无特别限制，可列举氢氧化钙和氢氧化钠。碱化合物也可预先加入水中。
然后，将此溶液过滤，回收滤液，用酸中和此提取液。酸并无特别限制，可使用无机酸或有机酸。
此中和液体经脱色、脱盐后直接或浓缩后再经喷雾干燥或冷冻干燥，制成粉末，这可作为粗精制的半纤维素制品使用。
再进一步用离心分离等手段从上述中和液中分离除去沉淀的蛋白质，必要时将其上清液进行透析、离子交换树脂处理、离子交换膜处理、超滤膜处理、酒精精制、滤剂处理等单独或组合处理，可得到任意纯度的半纤维素。
此外，本发明中所用的半纤维素的部分分解产物可制备如下。
将如上所述碱提取玉米外壳所得的提取液澄清过滤后调整pH，于50-60℃温度下添加木聚糖酶，进行反应。木聚糖酶的添加量以相对于每克提取物固形成分0.0001-10单位左右为佳，反应时间以3-96小时左右为佳。
木聚糖酶效价的测定如下：以玉米外壳碱提取所得的半纤维素为基质，将pH 7.0、60℃的反应条件下1分钟内生成相当于1μmol木糖的还原糖的酶活性作为1单位。
木聚糖酶以液化型的比糖化型的为佳，既可使用真菌也可使用细菌来源的，细菌来源的木聚糖酶纯度高因而较佳。特别好的例子可列举特公昭50-13357号公报中记载的偏碱性至适当pH的碱性木聚糖酶。
然后，通过加热等使酶失活，反应液经脱色、脱盐处理，浓缩，或进一步再干燥，可得到半纤维素的部分分解产物。
还可将如前所述纯化的半纤维素配制成含3-30质量％的溶液，根据需要调节pH，与上述同样地碱性酶促反应，将酶失活后，经离心分离后将上清液浓缩、干燥。
用上述方法从玉米外壳制得的半纤维素部分分解产物市售品为“セルェ一ス”(商品名，日本食品化工株式会社制)。
本发明的来自玉米外壳的半纤维素部分分解产物质量平均分子量以2万-20万为佳，2万-10万为更佳，2万-4万为最佳。具有这样的质量平均分子量的半纤维素的部分分解产物，其5质量％水溶液的粘度为3-20cps(B型粘度计，60rpm，25℃)的低粘度，添加到饮料、食品等原料中时，饮料、食品等的口感和风味等可保持良好。半纤维素的部分分解产物的质量平均分子量超过20万时，粘度过高，难以添加到饮料、食品原料等中，粘度不到2万时，有失去作为食物纤维的生理活性效果之虞，因而不理想。
本发明的免疫激活剂含上述半纤维素和/或其部分分解产物为主要成分，可制成如含上述半纤维素和/或其部分分解产物的水溶液、浓缩液或干燥粉末等制品。除了上述半纤维素和/或其部分分解产物外，也可含有除不掉的淀粉、蛋白质及若干木素、纤维素、灰分等。
本发明的免疫激活剂即便为干燥粉末时，也易溶于水，因此可直接作为保健饮料、食品、医药品等使用。而少量添加于饮料、食品中，可赋予不损害饮料、食品风味、口感的免疫活化效果。
饮料、食品中添加本发明的免疫激活剂时，可望不损害饮料、食品口感的免疫活化效果的添加量较佳为0.1-10质量％。为了显现免疫激活作用，本发明的免疫激活剂的摄取量较好为1-10g/天，特别好的为3-5g/天。
以下列举实施例对本发明作具体的说明。
实施例
在100质量份玉米外壳中，加入1000质量份水、1质量份氢氧化钙，85℃加热3小时。将此反应液冷却至60℃后，加入硫酸，调节为pH7。
然后，每克该反应液的固形成分加入0.01单位按特公昭50-13357号公报记载的方法制备的碱性木聚糖酶(商品名“セルザィム”，日本食品化工株式会社制)，60℃反应4小时。酶促反应结束后，于90℃加热30分钟，使酶失活，回收滤过的滤液，再进行澄清过滤、脱色、脱盐精制后，用喷雾干燥器进行喷雾干燥，得到来自玉米外壳的半纤维素的部分分解产物粉末。
试验例1
4周龄的雌性小鼠(C3H/Hej)12只，进行7天预饲养后，分成2组(每组6只)，试验组连续20天用胃管经口给予上述实施例1制备的来自玉米外壳的半纤维素部分分解产物的蒸馏水溶液(每次50mg/kg)。对照组同样地给予蒸馏水。试验期间自由摄食和摄水。
每天9：00-10：00观察症状和测定体重等。关于试验期间体重变化、摄食和摄水量的结果见表1、2。
表1
| 1日 | 14日 | 28日 | ||
| 体重(g) | 对照组(n＝6) | 14.62±1.05 | 21.85±0.65 | 23.93±0.36 |
| 试验组(n＝6) | 14.28±1.12 | 21.92±0.92 | 23.68±0.98 | |
表2
| 7日 | 14日 | 21日 | 28日 | ||
| 饲料摄取量(g/只) | 对照组 | 3.10 | 3.50 | 3.70 | 4.00 |
| 试验组 | 3.20 | 3.60 | 3.70 | 3.90 | |
| 水摄取量(ml/只) | 对照组 | 3.85 | 4.22 | 4.53 | 4.67 |
| 试验组 | 3.67 | 4.50 | 4.53 | 4.83 | |
从表1、2的结果可见，通过摄取来自玉米外壳的半纤维素的部分分解产物，对体重等无影响。试验期间动物的一般情况良好，中途无死亡或频死状况的动物。
在28日试验结束后，将受试小鼠在nenbutal麻醉下打开腹腔，从腹腔静脉取血，摘出脾脏。在解剖时的剖检中，未见因给予来自玉米外壳的半纤维素部分分解产物而引起的变化。
按常规方法从摘出的脾脏制备脾细胞，用添加10(v/v)％FCS的RPMI-1640培养基调节脾细胞数目，分别以2×106/ml/孔注入24孔板。
然后，加入ConA(50μg/ml)100μl，使终浓度为5μg/ml，于37℃、5％CO2培养器内培养24小时。
培养后，将各孔的细胞悬浮液用带有0.45μm滤膜的注射滤器过滤，此滤过液作为IL-2、INF-γ和IL-4活性测定用的样品。
使用BIOSOURCE公司的Cytoscreen药盒用平板读数系统进行各种细胞因子浓度的测定。
各测定值表示为平均值±标准差，用Student’s t测验进行显著性测定，以p＜0.05、p＜0.01的危险率作有无显著性差异的判断。其结果如表3所示。
表3
| IL-2 | IFN-γ | IL-4 | ||
| 浓度(pg/ml) | 对照组 | 52.77±8.25 | 2.75±1.42 | 3.75±1.00 |
| 试验组 | 130.25±30.24** | 6.97±2.78** | 4.68±1.43 | |
平均值±标准差*p＜0.05、**p＜0.01为与对照组相比有显著性差异。
如表3所明显显示的，来自玉米外壳的半纤维素部分分解产物给药组脾细胞对ConA刺激产生细胞因子(IL-2、IFN-γ和IL-4)的能力显示明显的促进提高的作用，IL-2和IFN-γ的产生比对照组有显著意义的差异。
从这些结果确认，来自玉米外壳的半纤维素部分分解产物的摄取在日常生活中对机体不引起抗原性等反应，而且感染和应激时对免疫机能有充分的促进作用。
产业上利用的可能性
如上所述，本发明可提供由玉米外壳制得的可使免疫力活化的安全的口服免疫激活剂。
通过口服本发明的免疫激活剂，可使免疫力低下、易患各种疾病的人的免疫力增强。
本发明的免疫激活剂还可作为医药品、饮料、食品或医药品、饮料、食品中的添加物，可放心地、长期地连续服用。而且，因为半纤维素部分分解产物是低粘度的，所以在添加到饮料、食品等原料中时可良好保持饮料、食品等的口感和风味。
Claims (5)
1.免疫激活剂，其特征在于：以得自玉米外壳的半纤维素和/或其部分分解产物为有效成分。
2.如权利要求1所述的免疫激活剂，其中所述半纤维素是从玉米外壳除去淀粉和蛋白质的剩余部分用碱提取所得的物质。
3.如权利要求1所述的免疫激活剂，其中所述半纤维素的部分分解产物是将权利要求2中所述半纤维素再用木聚糖酶处理后所得的物质。
4.如权利要求3所述的免疫激活剂，其中所述半纤维素的部分分解产物的质量平均分子量为2万-20万。
5.经口给予、具有促进细胞因子产生作用的权利要求1-4之一所述的免疫激活剂。
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| JP2000141181A JP4698796B2 (ja) | 2000-05-15 | 2000-05-15 | 免疫賦活剤 |
| JP141181/00 | 2000-05-15 |
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| US (1) | US20020010151A1 (zh) |
| EP (1) | EP1287827B1 (zh) |
| JP (1) | JP4698796B2 (zh) |
| KR (1) | KR100803636B1 (zh) |
| CN (1) | CN1245989C (zh) |
| AT (1) | ATE285780T1 (zh) |
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| JP4869492B2 (ja) * | 2001-05-14 | 2012-02-08 | 日本食品化工株式会社 | 抗腫瘍組成物 |
| KR20030091101A (ko) * | 2002-05-22 | 2003-12-03 | 주식회사 삼양제넥스 | 옥피 유래의 수용성 식이섬유를 포함하는 조성물 |
| US20040231060A1 (en) * | 2003-03-07 | 2004-11-25 | Athenix Corporation | Methods to enhance the activity of lignocellulose-degrading enzymes |
| JP5107545B2 (ja) * | 2006-09-11 | 2012-12-26 | 日本食品化工株式会社 | パイエル板形成誘導組成物 |
| KR101038401B1 (ko) * | 2009-04-21 | 2011-06-03 | 스마트전자 주식회사 | 소형퓨즈 및 그 제조방법 |
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| JPH0723315B2 (ja) * | 1986-02-28 | 1995-03-15 | 日本食品化工株式会社 | 血糖値上昇抑制物質 |
| JPS63165325A (ja) * | 1986-12-27 | 1988-07-08 | Snow Brand Milk Prod Co Ltd | 整腸剤 |
| US5565492A (en) * | 1988-07-18 | 1996-10-15 | Alcon Laboratories, Inc. | Prostaglandin combinations in glaucoma therapy |
| JP2938471B2 (ja) * | 1989-07-18 | 1999-08-23 | 日本食品化工株式会社 | 水溶性食物繊維含有飲食品 |
| US5622738A (en) * | 1989-05-16 | 1997-04-22 | Nihon Shokuhin Kako Co., Ltd. | Method of preparing water-soluble dietary fiber |
| JPH0678367B2 (ja) * | 1990-05-15 | 1994-10-05 | 呉羽化学工業株式会社 | 食物繊維、その製造法及びその食物繊維を含有する生理活性剤 |
| JP2886950B2 (ja) * | 1990-07-11 | 1999-04-26 | 日本食品化工株式会社 | 水溶性食物繊維の製造法 |
| JPH0517503A (ja) * | 1991-07-15 | 1993-01-26 | Terumo Corp | 水溶性ヘミセルロースおよび水溶性ヘミセルロース含有食品 |
| AU665287B2 (en) * | 1992-12-21 | 1995-12-21 | Alcon Laboratories, Inc. | Prostaglandin combinations in glaucoma therapy |
| JP3641284B2 (ja) * | 1994-05-27 | 2005-04-20 | 日本食品化工株式会社 | 耐糖能障害改善剤 |
| US5560914A (en) * | 1995-07-12 | 1996-10-01 | Daiwa Pharmaceutical Co., Ltd. | Immunopotentiator and method of manufacturing the same |
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| Publication number | Publication date |
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| JP4698796B2 (ja) | 2011-06-08 |
| CN1245989C (zh) | 2006-03-22 |
| ATE285780T1 (de) | 2005-01-15 |
| EP1287827A4 (en) | 2003-05-02 |
| US20020010151A1 (en) | 2002-01-24 |
| DE60108116D1 (en) | 2005-02-03 |
| KR100803636B1 (ko) | 2008-02-19 |
| EP1287827A1 (en) | 2003-03-05 |
| KR20020025095A (ko) | 2002-04-03 |
| EP1287827B1 (en) | 2004-12-29 |
| WO2001087311A1 (fr) | 2001-11-22 |
| JP2001322942A (ja) | 2001-11-20 |
| DE60108116T2 (de) | 2005-12-01 |
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