CN113508904A - 一种具有降脂功能的复方发酵液的制备方法 - Google Patents
一种具有降脂功能的复方发酵液的制备方法 Download PDFInfo
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Abstract
本发明涉及一种具有降脂功能的复方发酵液的制备方法,将人参、荷叶、茯苓、赤小豆、陈皮和肉桂的粉末状混合物经浸提、热处理、冷却,然后再接种具有降脂、抗氧化能力的益生菌发酵剂,经发酵制得具有降脂功能的复方发酵液。所述具有降脂、抗氧化能力的益生菌发酵剂为发酵乳杆菌、植物乳杆菌、鼠李糖乳杆菌中的任意一种或者混合菌种。通过本发明,将人参、荷叶、茯苓、赤小豆、陈皮和肉桂的粉末状混合物经浸提、热处理、冷却,然后再接种具有降脂、抗氧化能力的益生菌发酵剂,经发酵制得具有降脂功能的复方发酵液。
Description
技术领域
本发明涉及一种具有降脂功能的复方发酵液的制备方法,属于功能食品技术领域。
背景技术
肥胖的形成在于长期的能量吸收大于能量消耗,因此造成能量过剩,以脂肪的形式储存在体内,形成体脂率过高或体重超标的肥胖状态,是世界上较普遍的慢性病。据美国中情局(CIA)信息显示,收录的191个国家的2016年成年人肥胖率中有127个国家的肥胖率大于15%。《中国居民营养与慢性病状况报告(2020年)》显示,我国城乡各年龄组居民超重肥胖率继续上升,慢性病患病/发病仍呈上升趋势。18岁及以上居民超重率和肥胖率分别为34.3%和16.4%;6-17岁儿童青少年超重率和肥胖率分别为11.1%和7.9%;6岁以下儿童超重率和肥胖率分别为6.8%和3.6%。肥胖会导致慢性炎症、胰岛素抵抗、高血压和血脂异常,是2型糖尿病、心血管疾病和非酒精性脂肪肝等多种慢性病的危险因素。
肥胖的治疗方法主要有药物治疗,改变不健康的饮食生活方式,增强运动以及外科手术治疗等。但是,饮食习惯的改变、增强运动较难以长时间地坚持,达不到好的减肥效果;在药物治疗方面,主要分为两类,一类是作用于中枢神经的抗肥胖剂,如西布曲明等,由于存在睡眠障碍、便秘、心跳加快、头痛、焦虑或头晕等副作用,已被禁止使用。另一类是非中枢作用的抗肥胖剂,如抑制肠道脂肪酶的奥利司他等,这类药物最常见的不良反应包括油性粪便、腹痛、排气、排便增多和大便失禁等,这些副作用限制了其临床应用。中药复方的优势刚好在于调理脏腑系统的平衡问题,发挥整体功能恢复平衡的作用。所以中药复方在治疗肥胖上具有流传久远的历史,直到现在一些经典药方(有些已沿用一千多年)在现代临床上仍发挥着较好的减肥疗效。
如专利文献CN931151651,公开了一种消肥保健口服液,由白术、天冬、朱芩、黄芪、当归、生地、云苓、制半夏、山萸肉、枸杞子、甘草、人参、柴胡、升麻、枳实、青皮、三棱、陈皮、砂仁、防风、鸡内金、山楂、莪术、泽泻、木通、瞿麦、车前子、赤小豆、肉桂、荷叶、赤白茯苓的水提取液构成,用于减肥健身,增强体质、旺盛精力,并可防治高血压、心脏病等疾病。但是,该口服液药味复杂,多达30余种,其中制半夏、柴胡、白术等属于中药范畴不能用于食品,并且药味难闻、口味不佳,会影响肥胖人群的长期服用的依从性,并且长期服用其安全性较难保证。
又如专利文献CN2006101657285,公开了一种治疗肥胖症的药酒,采用人参片、海藻、肉桂、小茴香、陈皮、山楂、茯苓、赤小豆、生薏苡仁、荷叶、嫩桑枝、配合纯优质粮白酒,具有补气养血,固脱生津、补脾益肺、益智安神、开心明目、大补元气的作用;消痰软坚、利水及理气、调中、燥湿、化痰。由于该药酒原料大多为药食两用材料,安全性较好,但是含有酒精,限制了不饮酒的肥胖人群的应用,同时也不利于服用后的日常工作、开车、外出等。
众所周知,肥胖不是一时一日形成的,同样减肥也不是一蹴而就,而是需要持之以恒的。西药基本具有明确的副作用,中成药有疗效,但存在味苦、难闻等不适合长期服用的弊端,导致减肥人群的依从性差,达不到理想的效果。因此,作为中药应具有可供人们长期服用需要一定的安全性、适口性等基本要求。
发明内容
本发明的目的就是针对上述问题,提供一种方便减肥人群长期饮用的具有降脂功能的复方发酵液的制备方法。
本发明的目的是通过以下技术方案实现的,一种具有降脂功能的复方发酵液的制备方法,其特征在于,将人参、荷叶、茯苓、赤小豆、陈皮和肉桂的粉末状混合物经浸提、热处理、冷却,然后再接种具有降脂、抗氧化能力的益生菌发酵剂,经发酵制得具有降脂功能的复方发酵液。
所述粉末状混合物中人参、荷叶、茯苓、赤小豆、陈皮和肉桂的混合质量比为5~15∶3~10∶5~20∶5~15∶1~6∶0.5~3。
所述浸提时,将粉末状混合物与去离子水混匀,常温下超声波辅助浸提30 min,然后于100℃水浴30 min,5000 rpm 离心10min取上清。
所述热处理为上清采用UHT,或者121/15-30min灭菌处理,冷却至37℃左右备用。
所述具有降脂、抗氧化能力的益生菌发酵剂为发酵乳杆菌、植物乳杆菌、鼠李糖乳杆菌中的任意一种或者混合菌种。
所述按3%接种量接种于上述经过灭菌、冷却的提取液中,在35-38℃温度下,发酵24-72 h,冷却并于4℃冷藏,制成含有活菌的参荷苓复方发酵液。
本发明方法先进科学,通过本发明,将人参、荷叶、茯苓、赤小豆、陈皮和肉桂的粉末状混合物经浸提、热处理、冷却,然后再接种具有降脂、抗氧化能力的益生菌发酵剂,经发酵制得具有降脂功能的复方发酵液。
本发明从中医对肥胖基本病机为气虚、痰湿和湿热,病位在脾、肝、肾等的认识,应用“药证对应”、“方证对应”的原则,从药食两用食物名单中优选具有补气、祛痰湿、清热功能的原料组成复方。调整恢复脏腑正常功能,祛除气虚、痰湿和湿热的症状,标本兼治达到健康减脂的目标。
针对肥胖的基本病机:气虚、痰湿和湿热,病位:脾、肝、肾等,应用“药证对应”、“方证对应”的原则,本发明从卫生部公布的药食两用食物中优选补脾益气的人参、清热利水的荷叶、利水渗湿的茯苓、利水消肿的赤小豆、理气化痰的陈皮,再兼以温补脾肾的肉桂配伍成参荷苓复方,本复方具有补气、清热、祛痰湿减脂的功效。
本发明原料配方中,人参味甘、苦,性温,归肺、脾、心、肾经,具有补脾益气的作用;含有皂苷类化合物,主要为人参皂苷,具有降脂的生理功效;人参皂苷经微生物代谢转化为稀有人参皂苷,则可发挥更强的降脂减重的作用。荷叶味苦,性平,归肝、脾、胃经,具有清暑化湿、升发清阳、凉血止血的作用;含有黄酮类、生物碱类化合物,具有抗氧化、抗衰老、降血脂、降胆固醇及治疗心脑血管疾病等功能。茯苓味甘、淡,性平,归心、肺、脾、肾经,具有利水渗湿、健脾的功效。赤小豆味甘、酸,性平,归心、小肠经,具有利水消肿、解毒排脓的功效。陈皮味苦、辛,性温,归肺、脾经,具有理气健脾,燥湿化痰的功效。肉桂味辛,性温,具有温中补肾、补中益气的作用。
本发明与专利文献CN931151651的消肥保健口服液相比较,药味极大地简少了,简化了产生工艺,避免了制半夏、柴胡、白术等属于中药范畴不能用于食品的药味难闻、口味不佳待因素,提高了减肥人群的长期服用的安全性和适应性。同时,本发明与专利文献CN2006101657285的治疗肥胖症的药酒相比较,由于避免了采用酒精的泡制成份,也利于不饮酒的减脂人群的应用,还不会对日常工作、开车、外出等造成影响。
最关键的是,针对风味、口味不佳的问题,本发明采用了接种具有降脂、抗氧化能力的益生菌发酵剂,经发酵制得具有降脂功能的复方发酵液。
本发明采用微生物发酵来解决。微生物发酵可以改善中药的不良风味,并提高疗效。益生菌有利于机体健康,可以直接被人类服用,其在肠道中定植、繁殖,直接或通过与宿主或其他微生物的相互作用发挥多种益生功能,如调节肠道功能、抗氧化、维持口腔健康、预防糖尿病、防治肥胖等。通过益生菌发酵,在增强或不降低降脂功能的基础上改善风味,达到西药或中成药的疗效水平,使得产品既有功效,又有良好的风味,适合长期服用。
进一步地,本发明所述粉末状混合物中人参、荷叶、茯苓、赤小豆、陈皮和肉桂的混合质量比为5~15∶3~10∶5~20∶5~15∶1~6∶0.5~3。上述比例的设计以人参为君药主攻补脾肾之气,以荷叶、茯苓和赤小豆为臣药用于祛湿清热,以陈皮、肉桂为佐药,用于调畅气机、温补脾肾,使得脾肾气足、正常代谢痰湿而排出体外。
所述浸提时,将粉末状混合物与去离子水混匀,常温下超声波辅助浸提30 min,然后于100℃水浴30 min,5000 rpm 离心10min取上清。采用超声波辅助浸提法可使复方粉末的成分在浸泡过程中更好地溶出。
所述热处理为上清采用UHT,或者121/15-30min灭菌处理。上述热处理可以达到灭菌效果并减少有效成分的破坏。
所述冷却方法是室温自然冷却至40℃以下。上述自然冷却节约水等能源。
所述具有降脂、抗氧化能力的益生菌发酵剂为发酵乳杆菌、植物乳杆菌、鼠李糖乳杆菌中的任意一种。上述具有降脂、抗氧化能力的益生菌可以与降脂作用的复方起到协同作用,并且经过上述益生菌的发酵,使得复方发酵液风味更佳。
所述接种发酵为常规发酵剂按3%接种量接种于复方提取液中,在35-38℃温度下,发酵24-72 h,取出放4℃冷藏,制成含有活菌的参荷苓复方发酵液。
所述的发酵剂的制备为先将乳酸菌菌种活化2代,然后按体积百分比为3%的接种量接种于经95℃/5min热处理后冷却的12%的脱脂乳粉复原乳中,于37℃培养至凝乳,制成发酵剂,4℃冷藏备用,该发酵剂的活菌数大于108 cfu/mL。
附图说明
图 1 为发酵后挥发性风味物质的总离子流图;
图 2 为发酵前挥发性风味物质的总离子流图。
具体实施方式
下面结合附图以及附图说明对本发明做进一步说明。
一、制备工艺:
实施例1:制备参荷苓复方发酵功能饮料。
(1)制备参荷苓复方提取液:
取人参、荷叶、茯苓、赤小豆、陈皮、肉桂按照10∶6∶10∶10∶3∶1重量比例混合,经粉碎得参荷苓复方粉末。
取1kg参荷苓复方粉末加20 L去离子水混匀,常温下超声波辅助浸提30 min,然后100℃水浴30 min,冷却后5000rpm离心10min,取上清,制成参荷苓复方提取液15 L。
(2)制备参荷苓复方发酵液:
将发酵乳杆菌grx08发酵剂0.45 L(3%接种量)接种于上述参荷苓复方提取液中,在37℃温度下,发酵36 h后冷却至常温再无菌灌装,并于4℃冷藏,即得15.45 L含有活性益生菌的参荷苓复方发酵功能饮料。
实施例2:制备参荷苓复方提取液的益生菌发酵乳。
(1)制备参荷苓复方提取液:
取人参、荷叶、茯苓、赤小豆、陈皮、肉桂按照10∶6∶10∶10∶3∶1重量比例混合,经粉碎得参荷苓复方粉末。
取1kg参荷苓复方粉末加10 L去离子水混匀,常温下超声波辅助浸提30 min,然后100℃水浴30 min,冷却后5000rpm离心10min,取上清,制成参荷苓复方提取液7.5 L。
(2)制备参荷苓复方益生菌发酵乳:
在150 L鲜牛乳,加入7.5 L(5%添加量)上述参荷苓复方提取液,搅拌均匀,20 Mpa压力均质,95℃热处理5-8分钟,冷却至35~40℃,接种常规制备的嗜热链球菌grx02和徳氏乳杆菌保加利亚亚种grx33发酵剂(grx02∶grx33为1∶5),接种量为3 %,37℃发酵至pH值为4.2~4.6,4℃冷藏,即得157.5 L含该复方提取液的益生菌发酵乳。
实施例3:制备参荷苓复方提取液制备益生菌发酵乳饮料。
(1)制备参荷苓复方提取液:
取人参、荷叶、茯苓、赤小豆、陈皮、肉桂按照10∶6∶10∶10∶3∶1重量比例混合,经粉碎得参荷苓复方粉末。
取2kg参荷苓复方粉末加40 kg去离子水混匀,常温下超声波辅助浸提30 min,然后100℃水浴30 min,冷却后5000 rpm离心10min,取上清,制成参荷苓复方提取液30 kg。
(2)制备参荷苓复方益生菌发酵乳饮料:
取1 kg的脱脂乳粉和1.5 kg的甜味剂,用30 kg参荷苓复方提取液进行溶解,充分混匀后,在95℃/5 min条件下热处理,待冷却至37℃,按3%接种量接入常规制备的鼠李糖乳杆菌1301发酵剂,在37℃下发酵24 h,冷却后采用无菌灌装,即得32.5 kg参荷苓复方益生菌发酵乳饮料。
实施例4:制备参荷苓复方发酵液功能补充剂。
(1)制备参荷苓复方提取液:
取人参、荷叶、茯苓、赤小豆、陈皮、肉桂按照10∶6∶10∶10∶3∶1重量比例混合,经粉碎得参荷苓复方粉末。
取1kg参荷苓复方粉末加10 L去离子水混匀,常温下超声波辅助浸提30 min,然后100℃水浴30 min,冷却后5000rpm离心10min,取上清,制成参荷苓复方提取液7.5 L。
(2)制备参荷苓复方发酵液冻干粉:
将常规制备的鼠李糖乳杆菌LV108发酵剂按3%接种量接种于上述参荷苓复方提取液中,在37℃温度下,发酵24h,即得含该复方提取液的益生菌发酵乳。
将含该复方提取液的益生菌发酵乳冷冻至-40℃~-60℃,经干燥压碎,制成含水量3%~4%的干粉0.07 kg。
将干粉加入奶粉、低聚糖、麦芽糊精、风味剂等制成粉状或片状产品,用于日常食用。
二、检测含该复方提取液的益生菌发酵液的活菌数:
将以上各例取得的含该复方提取液的益生菌发酵液采用平板计数法和pH计法进行检测,测得各含该复方提取液的益生菌发酵乳的活菌数lg(CFU/mL)值达到8.78 ±0.057,pH值降到3.98 ± 0.017。
三、功能评价:
从现代医学角度看,肥胖存在高血脂、高血糖、胰岛素抵抗以及慢性炎症等问题。因此选取脂肪酶抑制率、α-葡萄糖苷酶抑制率、总抗氧化能力以及DPPH自由基清除能力作为体外功能验证的指标。
本发明利用脂肪酶抑制率、α-葡萄糖苷酶抑制率、总抗氧化能力以及DPPH自由基清除能力进行体外功能验证。利用GC-MS对参荷苓复方益生菌发酵液进行挥发性风味物质研究。
将以上各例取得的含该复方提取液的益生菌发酵乳分别以5000rpm离心5min,取上清即为发酵上清液,用于测定以下指标。
1、脂肪酶抑制率测定:
在280 uL反应体系中依次加入180 uL pH 7.2 磷酸盐缓冲液、40 uL 发酵上清液和20 uL 10 mM的 p-NPP(对硝基苯棕榈酸酯),37℃温浴10 min,再加入40 uL 10 mg/ml胰脂肪酶溶液,充分混匀,37℃温浴15 min,立即加入200uL无水酒精终止反应。12000 rpm离心2 min,取上清测定405 nm波长下吸光值,记录为B1. 以磷酸盐缓冲液替代发酵上清液作为样品空白对照,记录为B0. 在280uL反应体系中依次加入220 uL pH 7.2 磷酸盐缓冲液、20 uL 10 mM的 p-NPP(对硝基苯棕榈酸酯),37℃温浴10 min,再加入40 uL 10 mg/ml胰脂肪酶溶液,充分混匀,37℃温浴15 min,立即加入200uL无水酒精终止反应。12000 rpm离心2 min,取上清测定405 nm波长下吸光值,记录为A1. 以磷酸盐缓冲液替代酶溶液作为空白对照,记录为A0. 脂肪酶抑制率计算方法如下:脂肪酶抑制率=(1-(B1-B0)/(A1-A0))×100%。
参荷苓复方发酵液对脂肪酶的体外抑制率为93.06 ± 1.65%,抑制能力相当于0.5 mg/ml的奥利司他溶液。发酵前的脂肪酶抑制率为60.81 ± 2.12%,发酵后提升了53.03%。
2、α-葡萄糖苷酶抑制率的测定:
在96孔板中依次加入100 uL pH 6.8 磷酸盐缓冲液、20 uL 发酵上清液和20 uL20 mM的 p-NPG,37℃温浴10 min,再加入20 uL 1U/ml α-葡萄糖苷酶溶液,充分混匀,37℃温浴反应15 min,立即用酶标仪测定405 nm波长下吸光值,记录为B1. 以磷酸盐缓冲液替代发酵上清液作为样品空白对照,记录为B0. 在96孔板中依次加入120 uL pH 6.8 磷酸盐缓冲液、20 uL 20 mM的 p-NPG,37℃温浴10 min,再加入20 uL 1U/ml α-葡萄糖苷酶溶液,充分混匀,37℃温浴反应15 min,立即用酶标仪测定405 nm波长下吸光值,记录为A1. 以磷酸盐缓冲液替代发酵上清液作为样品空白对照,记录为A0. α-葡萄糖苷酶抑制率计算方法如下:α-葡萄糖苷酶抑制率=(1-(B1-B0)/(A1-A0))×100%。
参荷苓复方发酵液对α-葡萄糖苷酶的体外抑制率为91.66±0.48%。发酵前的α-葡萄糖苷酶抑制率为93.35 ± 0.24%,发酵后降低了1.81%。
3、总抗氧化能力的测定:
FRAP工作液配制:0.3 M pH 3.6 醋酸缓冲液:0.364g无水醋酸钠+3.2mL冰乙酸定容至200mL,用1M HCl调节pH至3.6; 10 mmol/L TPTZ溶液25mL:0.078g TPTZ用40mM盐酸溶液定容至25mL; 20mmol/L FeCl3溶液50mL:2.78g用RO水定容至50mL;上述溶液以10:1:1的比例混合(现配现用)。取上述发酵上清液5uL加入180uL工作液中,37℃条件下温浴3min,于593nm处测定吸光度.
参荷苓复方发酵液的总抗氧化能力为2.34±0.14 mM FeSO4当量。发酵前的总抗氧化能力为1.77±0.04 mM FeSO4当量,发酵后提升了32.2%。
4、DPPH自由基清除能力的测定:
用无水乙醇配制 0.2mmol/L 的DPPH溶液,避光保存;在96孔板中依次加入45 uL稀释10倍的发酵上清液溶液和100 uL DPPH 溶液,混匀,室温下暗处静置 30min 后测定其516nm处吸光度A sample,同时测定 100 uL DPPH溶液与 45 uL无水乙醇混合后的吸光度A0。DPPH自由基清除能力的表示:DPPH 清除百分率=(A0-A sample)/A0*100。
参荷苓复方发酵液的DPPH自由基清除能力为82.91±0.73%。发酵前的DPPH自由基清除能力为72.31±0.71%,发酵后降低了14.7%。
四、挥发性风味物质测定:
分析采用固相微萃取-气相色谱质谱联用技术(SPME-GC-MS)测定发酵前后挥发性风味物质,取2 mL样品于15 mL顶空萃取瓶中置于4 ℃冷藏备用。
①色谱条件:载气为He,流速1.0 mL/min;不分流进样,进样口温度250 ℃。程序升温方式,起始温度为35 ℃,保持5 min后以5 ℃/min的速率上升至140 ℃,保持2 min,再以10 ℃/min的速率上升至250 ℃,保持3 min。
②质谱条件:全扫描模式,EI离子源,电子能量70 eV,离子源温度为230 ℃,质量扫描范围m/z:35-500 AMU,无溶剂延迟。
③SPME萃取条件:萃取头老化:于250 ℃进样口老化30-60 min。固相微萃取条件:45 ℃磁力搅拌器上吸附60 min。解吸附条件:250 ℃条件下解吸附3 min。
④定性与定量分析:利用随机携带Masshunter工作站NIST 2.2标准库自动检索各组分质谱数据,利用内标法计算各组分相对含量。
发酵后去除了青草药的味道,降低了肉桂的风味,增加了果香味,使得参荷苓复方发酵液具有良好的风味、口感。发酵后挥发性风味物质的总离子流图如图1所示,发酵前挥发性风味物质的总离子流图如图2所示
由图1可见发酵前(图2)挥发性风味物质峰较少,最高的为保留时间23.3 min处的肉桂醛(相对峰面积44.19%,质量浓度3402.57μg/L)具有肉桂原料的特有气味。发酵后(图1)肉桂醛的相对峰面积和质量浓度分别减少到0.32%和7.55μg/L,23.3 min处峰几乎不可见,感官上也明显降低了肉桂的特有风味;此外,发酵后产生了保留时间25.59 min处的肉桂醇的峰(相对峰面积0.55%,质量浓度12.87μg/L),肉桂醇广泛用于配制食用香精,主要有杏、桃、树莓、李等香型。正已醇(保留时间9.79 min)和正庚醇(保留时间10.01 min)具有苦涩的青草味,发酵后未检出这两种挥发性物质。
参荷苓复方提取液的制备
1、制备参荷苓复方提取液:
将人参、荷叶、茯苓、赤小豆、陈皮、肉桂粉混合,按1kg复方粉加5-30 L去离子水混匀,常温下超声波辅助浸提30 min,然后100℃水浴30 min,冷却后5000rpm离心10min,取上清液即得参荷苓复方提取液。
2、杀菌、冷却
上述提取液经采用UHT,或者121/15-30min灭菌处理后,冷却至40℃以下备用。
3、发酵剂制备
将乳酸菌菌种活化2代,按体积百分比为3%的接种量接种于经95℃/5 min热处理后冷却的12%的脱脂乳粉复原乳中,于37℃培养至凝乳,制成发酵剂,4℃冷藏备用,该发酵剂的活菌数大于108 cfu/mL。
4、发酵液的制备
将上述发酵剂按3%接种量接种于上述提取液中,在37℃下,发酵48 h,取出放4℃冷藏,制成含有活菌的参荷苓复方发酵液。
Claims (6)
1.一种具有降脂功能的复方发酵液的制备方法,其特征在于,将人参、荷叶、茯苓、赤小豆、陈皮和肉桂的粉末状混合物经浸提、热处理、冷却,然后再接种具有降脂、抗氧化能力的益生菌发酵剂,经发酵制得具有降脂功能的复方发酵液。
2.根据权利要求1所述具有降脂功能的复方发酵液的制备方法,其特征在于,所述粉末状混合物中人参、荷叶、茯苓、赤小豆、陈皮和肉桂的混合质量比为5~15∶3~10∶5~20∶5~15∶1~6∶0.5~3。
3.根据权利要求1所述具有降脂功能的复方发酵液的制备方法,其特征在于,所述浸提时,将粉末状混合物与去离子水混匀,常温下超声波辅助浸提30 min,然后于100℃水浴30min,5000 rpm 离心10min取上清。
4.根据权利要求3所述具有降脂功能的复方发酵液的制备方法,其特征在于,所述热处理为上清采用UHT,或者121/15-30min灭菌处理,冷却至37℃左右备用。
5.根据权利要求1所述具有降脂功能的复方发酵液的制备方法,其特征在于,所述具有降脂、抗氧化能力的益生菌发酵剂为发酵乳杆菌、植物乳杆菌、鼠李糖乳杆菌中的任意一种或者混合菌种。
6.根据权利要求4所述具有降脂功能的复方发酵液的制备方法,其特征在于,所述按3%接种量接种于上述经过灭菌、冷却的提取液中,在35-38℃温度下,发酵24-72 h,冷却并于4℃冷藏,制成含有活菌的参荷苓复方发酵液。
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