CN107057086A - 金银花/薄荷复合多糖水凝胶的制备方法及用途 - Google Patents
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Abstract
金银花/薄荷复合多糖水凝胶的制备方法及用途,其方法的步骤为:(1)取金银花薄荷多糖粉末,反应釜中,然后加入去离子水,搅拌2h;(2)将金银花薄荷多糖尔量,配成饱和溶液,在避光的状态下反应8h;(3)反应结束后,向反应体系中乙二醇,搅拌;(4)反应结束后,加入氯化钠;(5)向反应体系中加入95%乙醇,搅拌后抽滤,然后用截留分子量为3500的透析袋透析72h,50℃下干燥12小时,即得氧化的金银花薄荷多糖;(6)称取氧化金银花薄荷多糖于反应釜中,加入蒸馏水;称取N,O‑羧甲基壳聚糖,溶于蒸馏水,将两种溶液等体积混合,在室温下静置,即得具有抗菌消炎作用的金银花薄荷多糖水凝胶。用途为用于抗菌消炎、伤口敷料、潜在的组织工程材料。
Description
技术领域
本发明属于生物材料制备技术,具体涉及金银花/薄荷复合多糖水凝胶的制备技术。
背景技术
水凝胶具有独特的三维网络结构,可吸水溶胀,溶胀后缓慢降解。水凝胶与细胞基质材料相似,且具有粘附性,可结合于生物体表或使一个生物体的表面与另外一个天然或合成材料的表面粘结。水凝胶良好的理化性质和生物相容性,能够有效提高载药量和生物半周期和稳定性等,使其更适合应用于药剂学领域,如作为一些缓释制剂的载体、水性软膏基质以及伤口辅料等。
制备水凝胶的材料多为无活性的天然或合成材料,难以发挥水凝胶的材料优势。而中草药中的天然高分子活性多糖根据种类不同而具有不同的功效,金银花多糖具有清热解毒抗炎及止血作用,其抗炎作用相当于皮炎平和地塞米松,薄荷多糖具有抗氧化、抗病毒等功效。将两者的优势结合起来制备复合多糖水凝胶既可以发挥两种多糖的功效又应用了水凝胶的载药优势,提高材料利用率。目前尚未见到具有抗菌消炎作用的水凝胶的报道。
发明内容
本发明的目的是提供一种金银花/薄荷复合多糖水凝胶的制备方法及用途。
本发明是金银花/薄荷复合多糖水凝胶的制备方法及用途,其方法的步骤为:
(1)取金银花薄荷多糖粉末,加入到干燥的反应釜中,然后加入与金银花薄荷多糖重量体积比w/v 为1:30~1:40的去离子水,在60℃下,150rmp/min搅拌2h;
(2)将金银花薄荷多糖尔量,以单体葡萄糖的相对分子质量计算,按照质量比n/w 1:0.40~1:0.60的NaIO4溶于适量水中,配成饱和溶液,缓慢加入到反应体系中,在避光的状态下反应8h;
(3)反应结束后,向反应体系中加入与金银花薄荷多糖摩尔体积比为n/w 1:0.10~1:0.20的乙二醇,150rmp/min搅拌5min;
(4)反应结束后,按照与金银花薄荷多糖量质量比为w/w 1:0.20~1:0.30向反应体系中加入氯化钠,150rmp/min搅拌15min;
(5)按照体积比v/v为1:3的比例向反应体系中加入95%乙醇,搅拌10min后抽滤,然后用截留分子量为3500的透析袋透析72h,50℃下干燥12小时,即得氧化的金银花薄荷多糖;
(6)称取氧化金银花薄荷多糖于反应釜中,按重量体积比w/v 为1:20加入蒸馏水;按照与氧化金银花薄荷多糖质量比为w/w1:2~1:4称取N,O-羧甲基壳聚糖,按质量体积比1:10~1:25溶于蒸馏水,将两种溶液等体积混合,在室温下静置10min,即得具有抗菌消炎作用的金银花薄荷多糖水凝胶。
金银花/薄荷复合多糖水凝胶的用途,用于抗菌消炎、伤口敷料、潜在的组织工程材料。
本发明的有益之处是:产品原料来源广泛,制备工艺简单,成本较低;水凝胶本身具有抗菌消炎活性,作为药物辅料,可以与其他抗菌消炎药构成复方,制备缓释制剂,减少主要用量,弥补缓释制剂载药量小的不足。抗菌消炎药物的使用,发挥辅料本身的优势,同时可实现药物缓释。
附图说明
图1是本发明制备的具有抗菌消炎作用的金银花薄荷复合多糖水凝胶的实物图,图2是本发明制备的缓释型金银花薄荷复合多糖水凝胶的抑制金黄色葡萄球菌图。
具体实施方式
本发明是金银花/薄荷复合多糖水凝胶的制备方法及用途,其方法的步骤为:
(1)取金银花薄荷多糖粉末,加入到干燥的反应釜中,然后加入体积为金银花薄荷多糖质量数30-40倍的去离子水,在60℃下,150rmp/min搅拌2小时;
(2)取质量为金银花薄荷多糖摩尔量(以单体葡萄糖的相对分子质量计算)的0.40-0.60倍的NaIO4溶于适量水中,配成饱和溶液,缓慢加入到反应体系中,在避光的状态下反应6-12h;
(3)反应结束后,向反应体系中加入体积数为金银花薄荷多糖摩尔量0.10-0.20倍的乙二醇,150rmp/min搅拌5min;
(4)反应结束后,向反应体系中加入4g氯化钠,150rmp/min搅拌15min;
(5)向反应体系中加入体积约3-4倍于前述反应体系的95%乙醇,搅拌10min后抽滤,然后用截留分子量为3500的透析袋透析72h,50℃下干燥12小时,即得氧化的金银花薄荷多糖;
(6)称取一定量氧化金银花薄荷多糖于反应釜中,按1:20(w/v)加入蒸馏水,称取氧化金银花薄荷多糖质量2-4倍的N,O-羧甲基壳聚糖,按质量体积比1:10-25溶于蒸馏水,将两种溶液等体积混合,在室温下静置10min,即得具有抗菌消炎作用的金银花薄荷多糖水凝胶,如图1所示。
(7)将氧化金银花薄荷多糖、N,O-羧甲基壳聚糖分别配制成20mg/ml、40mg/ml、60mg/ml、80mg/ml、100mg/ml溶液,用氧化金银花薄荷多糖和N,O-羧甲基壳聚糖制备水凝胶用于抗菌实验。
(8)菌悬液的制备 将供试的菌种进行斜面活化,取已活化的供试菌用接种环挑取2环至9mL的试管中,制成菌悬液。再用移液枪取1mL充分混匀的菌悬液于试管中,加入9mL无菌水,用显微镜直接计数法测定菌体个数,调至菌体浓度为106-107个/mL,备用。
(9)圆形纸片法 选择吸水性强的滤纸用打孔器打成直径为6 mm的圆形滤纸片,灭菌。取滤纸片在无菌的样品溶液中浸泡24h,风干备用。将灭过菌的培养皿、培养基放入超净工作台中冷却至60℃后倒平板。每皿约20mL,倒完后水平放置培养皿待培养基凝固。
(10)无菌操作,吸取0.1mL的菌液涂布做成含菌平板,平行三组,在培养基表面相应位置放上含有样品的滤纸片或用注射器将待成胶的金银花薄荷凝胶注射至平板上,每个平板3个滤纸片或三个凝胶恒温培养(37℃ ,24 h),比较抑菌效果。
表1 氧化金银花薄荷多糖、N,O-羧甲基壳聚糖、金银花薄荷多糖水凝胶的抑菌性能
注:“+”能抑制微生物生长;“++”能完全抑制微生物生长;“-”不能抑制微生物生长
金银花/薄荷复合多糖水凝胶的用途,用于抗菌消炎、伤口敷料、潜在的组织工程材料。
以下结合实施例对本发明作进一步详细描述。
实施例1:
1)取180g质量的金银花薄荷多糖粉末,加入到干燥的反应釜中,然后加入去离子水5400ml,在60℃下,150rmp/min搅拌2h;
2)取106.95gNaIO4溶于适量水中,配成饱和溶液,缓慢加入到反应体系中,在避光的状态下反应8h;
3)再向反应体系中22.32ml乙二醇,150rmp/min搅拌5min;
4)再向反应体系中加入4gNaCl,150rmp/min搅拌5min;
5)向反应体系中加入约15000ml95%乙醇,搅拌10min后抽滤,然后用截留分子量为3500的透析袋透析72h,50℃下干燥12小时,即得氧化的金银花薄荷多糖;
6)取25g氧化的金银花薄荷多糖,溶于500ml蒸馏水,制备存贮液A;
7)称取50gN,O-羧甲基壳聚糖,溶于500ml蒸馏水中,制备存贮液B;
8)将存贮液A与存贮液B充分混匀后制备成金银花薄荷多糖凝胶贴。
9)取制备好的具有抗菌消炎作用的金银花薄荷多糖水凝胶贴备用。10只雄性小鼠适应性喂养1 W后,将20μL二甲苯均匀涂抹在小鼠左耳/右耳内外两侧,30min后将制备的金银花薄荷多糖水凝胶切片贴敷于小鼠右耳,30min后断颈处死小鼠,剪下左右耳,叠放后用打孔器取双侧耳同一位置的耳片,称重后计算两耳片质量差作为耳廓肿胀度。耳廓肿胀度为:6.9(±2.2)㎎
实施例2:
1)取180g质量的金银花薄荷多糖粉末,加入到干燥的反应釜中,然后加入去离子水5400ml,在60℃下,150rmp/min搅拌2h;
2)取106.95gNaIO4溶于适量水中,配成饱和溶液,缓慢加入到反应体系中,在避光的状态下反应8h;
3)再向反应体系中22.32ml乙二醇,150rmp/min搅拌2h;
4)再向反应体系中加入4gNaCl,150rmp/min搅拌5min;
5)向反应体系中加入约15000ml95%乙醇,搅拌10min后抽滤,然后用截留分子量为3500的透析袋透析72h,50℃下干燥12小时,即得氧化的金银花薄荷多糖;
6)取25g氧化的金银花薄荷多糖,溶于500ml蒸馏水,制备存贮液A;
7)称取50gN,O-羧甲基壳聚糖,溶于500蒸馏水中,制备存贮液B;
8)将存贮液A与存贮液B充分混匀后制备成金银花薄荷多糖凝胶片。
9)取制备的具有抗菌消炎作用的金银花薄荷多糖水凝胶片备用。10只雄性小鼠适应性喂养1 W后,每日一次,每次0.5g,根据实际适当调剂给药剂量。将30只小鼠随机分为3组,每组10只,分别为:模型对照组,阳性对照组,金银花薄荷多糖凝胶组。模型对照组给予生理盐水(剂量为0.2㎎/mL),阳性对照组给予地塞米松磷酸钠注射液(浓度0.5%,剂量为0.01 mL/10g),给药组分别给予金银花薄荷多糖凝胶组(剂量为250mg/10g)。连续喂养1W后,末次给药后30min,在小鼠右耳正反两面用微量加样器各滴加10μL二甲苯致炎。同时给各鼠尾静脉注射Even's blue,每0.25mL(0.2% 溶液)。末次给药后l小时,将小鼠颈椎脱臼处死,沿耳廓基部剪下两耳,用直径9毫米的打孔器在左右耳的同一部位打下圆形耳片。称同一小鼠左右两耳片的重量,以左右两耳片重量之差表示肿胀度,并比较试验组与对照组间差异的显著性。
表2 金银花薄荷多糖水凝胶的抗炎效果
实施例3:
1)取180g质量的金银花薄荷多糖粉末,加入到干燥的反应釜中,然后加入去离子水5400ml,在60℃下,150rmp/min搅拌2h;
2)取106.95gNaIO4溶于适量水中,配成饱和溶液,缓慢加入到反应体系中,在避光的状态下反应8h;
3)再向反应体系中22.32ml乙二醇,150rmp/min搅拌2h;
4)再向反应体系中加入4gNaCl,150rmp/min搅拌5min;
5)向反应体系中加入约15000ml95%乙醇,搅拌10min后抽滤,然后用截留分子量为3500的透析袋透析72h,50℃下干燥12小时,即得氧化的金银花薄荷多糖;
6)取25g氧化的金银花薄荷多糖,溶于500ml蒸馏水,制备存贮液A;
7)称取50gN,O-羧甲基壳聚糖,溶于500蒸馏水中,制备存贮液B;
取75㎎阿司匹林粉末溶于存贮液B,迅速和存贮液A混合,为水相;与油相液体石蜡按一定比例混合后,300转/分快速搅拌120min,即得具有缓释作用的诺氟沙星/金银花薄荷多糖微球。
Claims (8)
1.金银花/薄荷复合多糖水凝胶的制备方法,其特征在于,其步骤为:
(1)取金银花薄荷多糖粉末,加入到干燥的反应釜中,然后加入与金银花薄荷多糖重量体积比w/v 为1:30~1:40的去离子水,在60℃下,150rmp/min搅拌2h;
(2)将金银花薄荷多糖尔量,以单体葡萄糖的相对分子质量计算,按照质量比n/w 1:0.40~1:0.60的NaIO4溶于适量水中,配成饱和溶液,缓慢加入到反应体系中,在避光的状态下反应8h;
(3)反应结束后,向反应体系中加入与金银花薄荷多糖摩尔体积比为n/w 1:0.10~1:0.20的乙二醇,150rmp/min搅拌5min;
(4)反应结束后,按照与金银花薄荷多糖量质量比为w/w 1:0.20~1:0.30向反应体系中加入氯化钠,150rmp/min搅拌15min;
(5)按照体积比v/v为1:3的比例向反应体系中加入95%乙醇,搅拌10min后抽滤,然后用截留分子量为3500的透析袋透析72h,50℃下干燥12小时,即得氧化的金银花薄荷多糖;
(6)称取氧化金银花薄荷多糖于反应釜中,按重量体积比w/v 为1:20加入蒸馏水;按照与氧化金银花薄荷多糖质量比为w/w1:2~1:4称取N,O-羧甲基壳聚糖,按质量体积比1:10~1:25溶于蒸馏水,将两种溶液等体积混合,在室温下静置10min,即得具有抗菌消炎作用的金银花薄荷多糖水凝胶。
2.根据权利要求1所述的金银花/薄荷复合多糖水凝胶的制备方法,其特征在于所述步骤(1)金银花薄荷多糖粉末与去离子水的质量体积比为w/v 1:30~1:40,温度为60℃下,搅拌时间为2h。
3.根据权利要求1所述的金银花/薄荷复合多糖水凝胶的制备方法,其特征在于所述步骤(2)NaIO4与金银花薄荷多糖的质量比w/w为0.40:1~0.60:1。
4.根据权利要求1所述的金银花/薄荷复合多糖水凝胶的方法,其特征在于所述步骤(3)中乙二醇与金银花薄荷多糖体积质量比v/w 为0.10:1~0.20:1。
5.根据权利要求1所述的金银花/薄荷复合多糖水凝胶的方法,其特征在于所述步骤(4)中加入氯化钠,且氯化钠与金银花薄荷多糖量质量比w/w0.20:1~0.30:1。
6.根据权利要求1所述的金银花/薄荷复合多糖水凝胶的方法,其特征在于所述步骤(5)中透析袋截留分子量为3500,透析时间为72h。
7.根据权利要求1所述的金银花/薄荷复合多糖水凝胶的方法,其特征在于所述步骤(6)中
氧化金银花薄荷多糖与N,O-羧甲基壳聚糖的质量比w/w为1:1~1:4。
8.金银花/薄荷复合多糖水凝胶的用途,其特征在于金银花/薄荷复合多糖水凝胶用于抗菌消炎、伤口敷料、潜在的组织工程材料。
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