WO2020215265A1 - 稀有人参皂苷Rh3和Rk2混合物的制备方法及其混合物 - Google Patents

稀有人参皂苷Rh3和Rk2混合物的制备方法及其混合物 Download PDF

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WO2020215265A1
WO2020215265A1 PCT/CN2019/084173 CN2019084173W WO2020215265A1 WO 2020215265 A1 WO2020215265 A1 WO 2020215265A1 CN 2019084173 W CN2019084173 W CN 2019084173W WO 2020215265 A1 WO2020215265 A1 WO 2020215265A1
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mixture
preparing
rare ginsenosides
rare
ginsenosides
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张琦
陈小春
傅荣昭
郭涛
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邦泰生物工程(深圳)有限公司
邦泰合盛生物科技（深圳）有限公司
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Priority to CN201980042274.8A priority Critical patent/CN112313342B/zh
Priority to PCT/CN2019/084173 priority patent/WO2020215265A1/zh
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    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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  • the present invention relates to the technical field of the preparation of rare ginsenosides, in particular to a method for preparing a mixture of Rh3 and Rk2 by means of biological enzyme catalysis and the mixture thereof.
  • Rare ginsenosides refer to the secondary metabolic derivatives of prototype ginsenosides extracted from Araliaceae plants (such as ginseng, panax notoginseng, American ginseng, Korean ginseng, etc.) after transformation, because they only exist in wild ginseng and red ginseng. Ginseng, black ginseng and other processed ginseng, and the content is very low (only contains 10,000 grades), so it is called rare ginsenoside by the industry.
  • rare ginsenosides have a smaller molecular weight than prototype ginsenosides, are easier to be absorbed by the human body, have higher bioavailability, and therefore have stronger anti-cancer activity. Therefore, the research focus on ginsenosides has gradually focused on rare ginseng Saponin above. After more than half a century of research, scientists have discovered more than 60 kinds of rare ginsenosides, including Rg3, Rg5, Rk1, Rk2, Rk3, Rh1, Rh2, Rh3, Rh4, ⁇ PPT, ⁇ PPD, etc. These rare ginsenosides have The anti-cancer effects of different. Canadian scientists tested the anti-cancer activity of some rare ginsenosides and compared them.
  • Rh2 and Rg3 are the most studied and widely used rare ginsenosides in China.
  • the comparison of the above-mentioned activities shows that the anticancer activity of Rh2 and Rg3 is not dominant in rare ginsenosides. Therefore, it is necessary to develop anti-cancer activities.
  • Other rare ginsenosides with stronger cancer activity However, the stronger the anti-cancer activity of rare ginsenosides, the more difficult it is to transform from prototype ginsenosides, and it is even more difficult to realize industrial production. It is these technical barriers that hinder the domestic rare ginsenosides.
  • the development of the industry has led to the development of rare ginsenoside products with higher activity except for Rh2 and Rg3.
  • the purpose of the present invention is to develop a method for preparing a mixture of rare ginsenosides Rh3 and Rk2 that is suitable for industrialization and low cost, so as to provide a rare ginsenoside product with stronger anticancer activity. So as to solve the technical problem of the scarcity of rare ginsenoside products in the domestic market.
  • the present invention provides a method for preparing a mixture of rare ginsenosides Rh3 and Rk2, which includes the following steps:
  • step (1) After the enzyme-catalyzed reaction in step (1) is finished, adjust the pH of the reaction solution to 2-3, and stir for 1-2 hours for acidification;
  • step (3) After the acidification reaction in step (2) is over, add ammonium sulfate to the reaction solution, and after it is fully dissolved, add an extractant composed of ethanol and ethyl acetate for shaking extraction, stand still for layering and collect the upper layer solution. After the solvent is recovered, the crude product of the mixture of Rh3 and Rk2 is obtained.
  • the ⁇ -glucosidase is preferably derived from Coenella 18JY8-7 (Cohnella sp. 18JY8-7).
  • the enzyme can convert Rg3 into Rh3 and Rk2 to the greatest extent at pH 4-6 and 20-50°C.
  • the temperature of the enzyme-catalyzed reaction is 30°C.
  • the added amount of the above-mentioned ⁇ -glucosidase is preferably 0.5-3 times the weight of Rg3.
  • the added amount of ⁇ -glucosidase from the above source is the same as the weight of Rg3.
  • the co-solvent is preferably DMSO.
  • the amount of DMSO added is 0.5-3ml/g Rg3.
  • the added amount of DMSO is 1 ml/g Rg3.
  • the enzyme-catalyzed reaction in step (1) is carried out in the presence of a phosphate buffer. More preferably, the phosphate buffer is sodium phosphate buffer.
  • the purpose of the acidification reaction in step (2) is to promote the reaction in a direction that is more conducive to the generation of Rk2, so as to obtain a mixture of Rh3 and Rk2 with a larger proportion of Rk2.
  • the pH value is preferably adjusted by adding hydrochloric acid in step (2), which has the advantages of low cost of hydrochloric acid, mild reaction process, more stable generation of Rk2 and minimal impurity generation.
  • step (3) of the above preparation method provided by the present invention the ⁇ -glucosidase in the reaction solution is separated by sedimentation and separation by adding ammonium sulfate, and the amount of ammonium sulfate added is preferably 0.01-0.03 times the weight of ⁇ -glucosidase; More preferably, the amount of ammonium sulfate added is 0.02 times the weight of ⁇ -glucosidase.
  • the amount of the extractant added is preferably twice the weight of Rg3.
  • the volume ratio of ethanol and ethyl acetate is preferably 1:0.5-5. More preferably, the volume ratio of ethanol and ethyl acetate is preferably 1:3.
  • the crude product of the Rh3 and Rk2 mixture prepared by the method of the present invention can be further purified, and the purification method can be any known applicable separation and purification method.
  • the above-mentioned preparation method provided by the present invention further includes the following refining steps:
  • the elution gradient is preferably 30-50% acetonitrile aqueous solution for 10 minutes, and 75-85% acetonitrile aqueous solution for 20 minutes.
  • the elution gradient is more preferably 40% acetonitrile aqueous solution for 10 minutes and 80% acetonitrile aqueous solution for 20 minutes. Under this elution condition, the resolution of the target substance and other impurities is higher, and the peak shape of the target peak is better.
  • the present invention also provides a rare ginsenoside Rh3 and Rk2 mixture, which is prepared by the above-mentioned preparation method provided by the present invention.
  • the present invention has the following advantages:
  • the preparation method of the rare ginsenoside Rh3 and Rk2 mixture provided by the present invention is low in cost and simple in operation, suitable for industrial production, and the mixture of Rh3 and Rk2 with a higher proportion of Rk2 can be prepared by this method, and the resulting mixture product Has stronger anti-cancer activity;
  • the rare ginsenoside Rh3 and Rk2 mixture provided by the present invention has stronger anticancer activity.
  • the ⁇ -glucosidase in the following examples was obtained by the inventor using the biological fermentation method. After biological sequencing, the inventor’s self-made enzyme and ⁇ -glucosidase derived from Cohnella sp. 18JY8-7 (Cohnella sp. 18JY8-7) -The amino acid sequence of glucosidase (Genbank accession number: CP033433.1) is the same. The remaining reagents and raw materials are all commercially available products.
  • the following examples adopt the following preparation method of the rare ginsenoside Rh3 and Rk2 mixture provided by the present invention to prepare the rare ginsenoside Rh3 and Rk2 mixture:
  • step (1) After the enzyme-catalyzed reaction in step (1) is completed, add dilute hydrochloric acid to adjust the pH of the reaction solution to 2-3, stir at room temperature for 1-2 hours for acidification reaction, and measure the acidification reaction by HPLC after the acidification reaction is over The content of Rh3 and Rk2 in the solution, and calculate the proportion of Rk2 in the mixture of Rh3 and Rk2, and the conversion rate of the substrate based on the mixture of Rh3 and Rk2. The results are shown in Table 2;
  • step (3) Add ammonium sulfate to the acidification reaction solution of step (2) and stir to dissolve it.
  • Example 4-5 Compared with Example 1-3, the difference of Example 4-5 is: after the enzyme-catalyzed reaction of step (1) is completed, the acidification reaction of step (2) is not performed, and the reaction solution of the enzyme-catalyzed reaction is directly determined by HPLC. The content of each substance, and then step (3) and step (4) operations.
  • Table 2 The data of Examples 4-5 are shown in Table 2.

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Abstract

稀有人参皂苷Rh3和Rk2混合物的制备方法及其混合物，该方法以Rg3为原料，用β-葡萄糖苷酶进行酶法制备，并在酶催化反应结束后进行酸化反应，然后通过溶剂萃取进行分离获得Rh3和Rk2混合物的粗品，该混合物中Rk2的占比高于Rh3。

Description

稀有人参皂苷Rh3和Rk2混合物的制备方法及其混合物 技术领域
本发明涉及稀有人参皂苷的制备的技术领域，特别涉及一种通过生物酶催化的技术手段制备Rh3和Rk2混合物的方法及其混合物。
背景技术
稀有人参皂苷，是指从五加科植物(如人参、三七、西洋参、高丽参等)中提取出来的原型人参皂苷经过转化后的次级代谢衍生物，因其只存在于野山参、红参和黑参等炮制人参中，且含量极低(仅含万分级)，所以被业界称为稀有人参皂苷。
研究发现，稀有人参皂苷相较于原型人参皂苷，分子量更小，更容易被人体吸收，生物利用度更高，因此具有更强的抗癌活性，于是对人参皂苷的研究重点逐渐聚焦在稀有人参皂苷上面。经过半个多世纪的研究，科学家目前已经发现了60多种稀有人参皂苷，包括Rg3、Rg5、Rk1、Rk2、Rk3、Rh1、Rh2、Rh3、Rh4、αPPT、αPPD等，这些稀有人参皂苷所具有的抗癌作用各不相同。加拿大科学家通过试验测定了部分稀有人参皂苷的抗癌活性，并进行了比较，其结果如下：Rg3＜Δ(20-22)PPD＝Rg5≤αPPT＝Rk1和Rg5混合物＝Rh2≤Δ(20-21)PPD＜aPPD＝Δ(20-21)PPT＝Δ(20-22)PPT＝Rh3和Rk2混合物≤Rk2＝Δ(20-21)PPD和Δ(20-22)PPD混合物。
目前在国内，对稀有人参皂苷研究最多、应用最广的是Rh2和Rg3，但是通过上述活性比较可知，Rh2和Rg3的抗癌活性在稀有人参皂苷中并不占优势，因此，有必要开发抗癌活性更强的其它稀有人参皂苷。但是，稀有人参皂苷的抗癌活性越强，从原型人参皂苷进行转化的技术难度就越大，而要实现工业化生产，更是难上加难，正是这些技术壁垒，阻碍了国内稀有人参皂苷产业的发展，导致除去Rh2和Rg3之外尚无其它活性更高的稀有人参皂苷产品被开发出来。
发明概述
技术问题
鉴于现有技术存在的上述不足，本发明的目的在于开发一种适于工业化且成本低廉的制备稀有人参皂苷Rh3和Rk2混合物的方法，以便提供一种抗癌活性更强的稀有人参皂苷产品，从而解决现有国内市场稀有人参皂苷产品稀缺的技术问题。
问题的解决方案
技术解决方案
为实现上述目的，本发明提供了一种稀有人参皂苷Rh3和Rk2混合物的制备方法，包括以下步骤：
(1)将人参皂苷Rg3溶解于助溶剂中，加入β-葡萄糖苷酶，在pH为4-6、温度为20-50℃的条件下进行酶催化反应；
(2)待步骤(1)的酶催化反应结束后，将反应液的pH值调节为2-3，搅拌1-2小时进行酸化反应；
(3)待步骤(2)的酸化反应结束后，向反应液中加入硫酸铵，充分溶解后再加入由乙醇和乙酸乙酯组成的萃取剂进行震荡萃取，静置分层后收集上层溶液，回收溶剂后即得Rh3和Rk2混合物的粗品。
本发明提供的上述制备方法中，β-葡萄糖苷酶优选来源于柯恩氏菌18JY8-7(Cohnella sp.18JY8-7)。该酶在pH4-6、20-50℃的条件下可将Rg3最大程度地转化为Rh3和Rk2。
更优选地，酶催化反应的温度为30℃。
本发明提供的上述制备方法中，上述来源的β-葡萄糖苷酶的加入量优选为Rg3重量的0.5-3倍。
更优选地，上述来源的β-葡萄糖苷酶的加入量与Rg3的重量相同。
本发明提供的上述制备方法中，助溶剂优先选用DMSO。
优选地，DMSO的加入量为0.5-3ml/g Rg3。
更优选地，DMSO的加入量为1ml/g Rg3。
优选地，本发明提供的上述制备方法中，步骤(1)的酶催化反应在磷酸盐缓冲液存在的条件下进行。更优选地，磷酸盐缓冲液为磷酸钠缓冲液。
本发明提供的上述制备方法中，步骤(2)的酸化反应的目的在于，促使反应 向更利于生成Rk2的方向进行，从而获得Rk2占比更多的Rh3和Rk2混合物。
本发明提供的上述制备方法中，步骤(2)中优选通过加入盐酸调节pH值，其优点在于，盐酸成本低廉、反应过程温和、Rk2的生成更稳定且杂质生成量最少。
本发明提供的上述制备方法的步骤(3)中，通过加入硫酸铵将反应液中的β-葡萄糖苷酶沉降分离，硫酸铵的加入量优选为β-葡萄糖苷酶重量的0.01-0.03倍；更优选地，硫酸铵的加入量为β-葡萄糖苷酶重量的0.02倍。
本发明提供的上述制备方法中，萃取剂的加入量优选为Rg3重量的2倍。
本发明提供的上述制备方法中，乙醇和乙酸乙酯的体积比优选为1∶0.5-5。更优选地，乙醇和乙酸乙酯的体积比优选为1∶3。
为了获得纯度更高的Rh3和Rk2混合物，可对本发明方法制备得到的Rh3和Rk2混合物的粗品进行进一步纯化，纯化方法可以是任何已知的可适用的分离纯化方法。
优选地，本发明提供的上述制备方法还包括以下精制步骤：
(4)将Rh3和Rk2混合物的粗品经制备液相色谱进行梯度洗脱，流动相为乙腈和水，色谱柱为聚苯乙烯柱，收集目的峰，干燥后即得Rh3和Rk2混合物的纯品。
本发明提供的上述制备方法的精制步骤中，洗脱梯度优选为30-50％乙腈水溶液洗脱10min，75-85％乙腈水溶液洗脱20min。
本发明提供的上述制备方法的精制步骤中，洗脱梯度更优选为40％乙腈水溶液洗脱10min，80％乙腈水溶液洗脱20min。在该洗脱条件下，目标物质与其他杂质的分离度更高，且目的峰的峰形更好。
另外，本发明还提供了一种稀有人参皂苷Rh3和Rk2混合物，该混合物是通过本发明提供的上述制备方法制备得到的。
发明的有益效果
有益效果
与现有技术相比，本发明具有以下优点：
1、本发明提供的稀有人参皂苷Rh3和Rk2混合物的制备方法成本低廉且操作简 单、适于工业化生产，并且通过该方法可以制备得到Rk2占比更高的Rh3和Rk2混合物，因而得到的混合物产品的抗癌活性更强；
2、本发明提供的稀有人参皂苷Rh3和Rk2混合物具有更强的抗癌活性。
发明实施例
本发明的实施方式
下面结合具体实施例对本发明做进一步的详细说明，以下实施例是对本发明的解释，本发明并不局限于以下实施例。
以下实施例中的β-葡萄糖苷酶为发明人利用生物发酵法自制获得，经生物测序后，该发明人自制酶与来源于柯恩氏菌18JY8-7(Cohnella sp.18JY8-7)的β-葡萄糖苷酶的氨基酸序列(Genbank登录号为：CP033433.1)相同。其余试剂及原料均为市售产品。
实施例1-实施例3
从工业生产的角度，出于减少生产时间、降低生产成本的目的，以下实施例采用本发明提供的如下稀有人参皂苷Rh3和Rk2混合物的制备方法制备稀有人参皂苷Rh3和Rk2混合物：
(1)称取人参皂苷Rg3，加入DMSO和pH值为4-6的磷酸钠缓冲液中，充分搅拌至Rg3完全溶解，DMSO的加入量为1ml/g人参皂苷Rg3，磷酸钠缓冲液的加入量为DMSO的5倍体积；然后加入与Rg3重量相同的β-葡萄糖苷酶，在温度为30℃的条件下进行酶催化反应10h；
(2)待步骤(1)的酶催化反应结束后，加入稀盐酸将反应液的pH值调节为2-3，室温搅拌1-2小时进行酸化反应，待酸化反应结束后通过HPLC测定酸化反应液中Rh3及Rk2的含量，并计算Rk2在Rh3和Rk2混合物中的占比、以及以Rh3和Rk2混合物计的底物转化率，结果如表2所示；
(3)向步骤(2)的酸化反应液中加入硫酸铵，搅拌使之溶解，硫酸铵的加入量为β-葡萄糖苷酶重量的0.02倍；待硫酸铵充分溶解后，再加入Rg3重量的2倍量的萃取剂，萃取剂的组成为V 乙醇∶V 乙酸乙酯＝1∶3，充分摇匀后静置分层，取上层溶液在70℃条件下减压蒸馏至原体积的1/3，回收溶剂，趁热过滤，再用1％体积的纯水冲洗滤饼，烘干后即得Rh3和Rk2混合物的粗品；
(4)将Rh3和Rk2混合物的粗品溶于乙腈中，用0.45μm膜过滤，按照表1中的条件将滤液经制备液相色谱进行梯度洗脱，收集目的峰，干燥后即得Rh3和Rk2混合物的纯品，产品的总收率如表2所示。
表1
[Table 1]
Figure PCTCN2019084173-appb-000001
实施例4-5
与实施例1-3相比，实施例4-5的区别点在于：步骤(1)的酶催化反应结束后不进行步骤(2)的酸化反应，通过HPLC直接测定酶催化反应的反应液中的各物质含量，然后再依次进行步骤(3)和步骤(4)的操作。实施例4-5的各数据如表2所示。
表2
[Table 2]
Figure PCTCN2019084173-appb-000002
Rh3和Rk2的结构确证
1、将分离得到的Rh3和Rk2混合物的纯品，溶于甲醇，通过高分辨液相质谱检测，液相质谱图中所有峰无任何差异，m/z为663.4411的峰为[M+CH3COO] -峰，M正好为Rh 3或Rk 2的分子量。
2、将Rh3和Rk2混合物的纯品溶于氘代吡啶中，在400MHz的Bruker核磁仪中测定，测定结果如表3所示。
表3
Figure PCTCN2019084173-appb-000003

Claims (11)

  1. 稀有人参皂苷Rh3和Rk2混合物的制备方法，其特征在于，所述方法包括以下步骤：
    (1)将人参皂苷Rg3溶解于助溶剂中，加入β-葡萄糖苷酶，在pH为4-6、温度为20-50℃的条件下进行酶催化反应；
    (2)待步骤(1)的酶催化反应结束后，将反应液的pH值调节为2-3，搅拌1-2小时进行酸化反应；
    (3)待步骤(2)的酸化反应结束后，向反应液中加入硫酸铵，充分溶解后再加入由乙醇和乙酸乙酯组成的萃取剂进行震荡萃取，静置分层后收集上层溶液，回收溶剂后即得Rh3和Rk2混合物的粗品。
  2. 根据权利要求1所述的稀有人参皂苷Rh3和Rk2混合物的制备方法，其特征在于：所述β-葡萄糖苷酶来源于柯恩氏菌18JY8-7。
  3. 根据权利要求2所述的稀有人参皂苷Rh3和Rk2混合物的制备方法，其特征在于：所述β-葡萄糖苷酶的加入量为Rg3重量的0.5-3倍。
  4. 根据权利要求1所述的稀有人参皂苷Rh3和Rk2混合物的制备方法，其特征在于：所述助溶剂为DMSO。
  5. 根据权利要求1所述的稀有人参皂苷Rh3和Rk2混合物的制备方法，其特征在于：所述步骤(2)中通过加入盐酸调节pH值。
  6. 根据权利要求1所述的稀有人参皂苷Rh3和Rk2混合物的制备方法，其特征在于：所述硫酸铵的加入量为所述β-葡萄糖苷酶重量的0.01-0.03倍。
  7. 根据权利要求1所述的稀有人参皂苷Rh3和Rk2混合物的制备方法，其特征在于：所述萃取剂的加入量为Rg3重量的2倍。
  8. 根据权利要求1所述的稀有人参皂苷Rh3和Rk2混合物的制备方法，其特征在于：所述乙醇和所述乙酸乙酯的体积比为1∶0.5-5。
  9. 根据权利要求1所述的稀有人参皂苷Rh3和Rk2混合物的制备方法，其特征在于，所述方法还包括以下精制步骤：
    (4)将Rh3和Rk2混合物的粗品经制备液相色谱进行梯度洗脱，流动相为乙腈和水，色谱柱为聚苯乙烯柱，收集目的峰，干燥后即得Rh3和Rk2混合物的纯品。
  10. 根据权利要求9所述的稀有人参皂苷Rh3和Rk2混合物的制备方法，其特征在于：所述步骤(4)中的洗脱梯度为30-50％乙腈水溶液洗脱10min，75-85％乙腈水溶液洗脱20min。
  11. 稀有人参皂苷Rh3和Rk2混合物，其特征在于：所述混合物通过权利要求1-10任一项所述的稀有人参皂苷Rh3和Rk2混合物的制备方法制备得到。
PCT/CN2019/084173 2019-04-25 2019-04-25 稀有人参皂苷Rh3和Rk2混合物的制备方法及其混合物 WO2020215265A1 (zh)

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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1508147A (zh) * 2002-12-13 2004-06-30 中国科学院大连化学物理研究所 一种催化热解制备低极性人参皂苷及其苷元的方法
CN107337704A (zh) * 2017-06-15 2017-11-10 吉林大学 一种稀有人参皂苷Rk2和Rh3的分离方法
CN107338280A (zh) * 2017-06-30 2017-11-10 肖永坤 一种高活性低糖基人参次苷群及其苷元的制备方法
CN108064309A (zh) * 2017-11-30 2018-05-22 邦泰生物工程(深圳)有限公司 一种酶催化合成人参皂苷Rh2的方法

Family Cites Families (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR101776005B1 (ko) * 2015-09-01 2017-09-12 셀루스원 주식회사 효소 및 유기산 처리를 통한 진세노사이드 Rh2, Rk2 및 Rh3 제조방법
CN108430596B (zh) * 2016-12-14 2020-10-30 邦泰生物工程(深圳)有限公司 一种人参皂苷Rh2的纯化方法
KR101968004B1 (ko) * 2017-10-13 2019-04-10 재단법인 지능형 바이오 시스템 설계 및 합성 연구단 진세노사이드 Rh2 믹스의 대량제조방법

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1508147A (zh) * 2002-12-13 2004-06-30 中国科学院大连化学物理研究所 一种催化热解制备低极性人参皂苷及其苷元的方法
CN107337704A (zh) * 2017-06-15 2017-11-10 吉林大学 一种稀有人参皂苷Rk2和Rh3的分离方法
CN107338280A (zh) * 2017-06-30 2017-11-10 肖永坤 一种高活性低糖基人参次苷群及其苷元的制备方法
CN108064309A (zh) * 2017-11-30 2018-05-22 邦泰生物工程(深圳)有限公司 一种酶催化合成人参皂苷Rh2的方法

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
陈小春 等 (CHEN, XIAOCHUN ET AL.): "稀有人参皂苷Ｒh3及Ｒk2的制备及结构确认 (Isolation and structural identification of Rare Human Saponins Rh3 and Rk2)", 江西化工 (JIANGXI CHEMICAL INDUSTRY), no. 2, 15 April 2019 (2019-04-15), XP55746163, DOI: 20191212121654Y *

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