JP6473803B2 - Extraction method of chlorogenic acid from Tochu leaves - Google Patents
Extraction method of chlorogenic acid from Tochu leaves Download PDFInfo
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- CWVRJTMFETXNAD-JUHZACGLSA-N chlorogenic acid Chemical compound O[C@@H]1[C@H](O)C[C@@](O)(C(O)=O)C[C@H]1OC(=O)\C=C\C1=CC=C(O)C(O)=C1 CWVRJTMFETXNAD-JUHZACGLSA-N 0.000 title claims description 90
- CWVRJTMFETXNAD-FWCWNIRPSA-N 3-O-Caffeoylquinic acid Natural products O[C@H]1[C@@H](O)C[C@@](O)(C(O)=O)C[C@H]1OC(=O)\C=C\C1=CC=C(O)C(O)=C1 CWVRJTMFETXNAD-FWCWNIRPSA-N 0.000 title claims description 89
- PZIRUHCJZBGLDY-UHFFFAOYSA-N Caffeoylquinic acid Natural products CC(CCC(=O)C(C)C1C(=O)CC2C3CC(O)C4CC(O)CCC4(C)C3CCC12C)C(=O)O PZIRUHCJZBGLDY-UHFFFAOYSA-N 0.000 title claims description 89
- CWVRJTMFETXNAD-KLZCAUPSSA-N Neochlorogenin-saeure Natural products O[C@H]1C[C@@](O)(C[C@@H](OC(=O)C=Cc2ccc(O)c(O)c2)[C@@H]1O)C(=O)O CWVRJTMFETXNAD-KLZCAUPSSA-N 0.000 title claims description 87
- 229940074393 chlorogenic acid Drugs 0.000 title claims description 87
- 235000001368 chlorogenic acid Nutrition 0.000 title claims description 87
- FFQSDFBBSXGVKF-KHSQJDLVSA-N chlorogenic acid Natural products O[C@@H]1C[C@](O)(C[C@@H](CC(=O)C=Cc2ccc(O)c(O)c2)[C@@H]1O)C(=O)O FFQSDFBBSXGVKF-KHSQJDLVSA-N 0.000 title claims description 87
- BMRSEYFENKXDIS-KLZCAUPSSA-N cis-3-O-p-coumaroylquinic acid Natural products O[C@H]1C[C@@](O)(C[C@@H](OC(=O)C=Cc2ccc(O)cc2)[C@@H]1O)C(=O)O BMRSEYFENKXDIS-KLZCAUPSSA-N 0.000 title claims description 87
- 238000000605 extraction Methods 0.000 title claims description 60
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- 238000001914 filtration Methods 0.000 claims description 24
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- 239000012043 crude product Substances 0.000 claims description 23
- 239000000284 extract Substances 0.000 claims description 23
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical group OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 21
- 229920001661 Chitosan Polymers 0.000 claims description 19
- 239000000706 filtrate Substances 0.000 claims description 12
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- GNFTZDOKVXKIBK-UHFFFAOYSA-N 3-(2-methoxyethoxy)benzohydrazide Chemical compound COCCOC1=CC=CC(C(=O)NN)=C1 GNFTZDOKVXKIBK-UHFFFAOYSA-N 0.000 claims 1
- FGUUSXIOTUKUDN-IBGZPJMESA-N C1(=CC=CC=C1)N1C2=C(NC([C@H](C1)NC=1OC(=NN=1)C1=CC=CC=C1)=O)C=CC=C2 Chemical compound C1(=CC=CC=C1)N1C2=C(NC([C@H](C1)NC=1OC(=NN=1)C1=CC=CC=C1)=O)C=CC=C2 FGUUSXIOTUKUDN-IBGZPJMESA-N 0.000 claims 1
- 240000004670 Glycyrrhiza echinata Species 0.000 claims 1
- 235000001453 Glycyrrhiza echinata Nutrition 0.000 claims 1
- 235000006200 Glycyrrhiza glabra Nutrition 0.000 claims 1
- 235000017382 Glycyrrhiza lepidota Nutrition 0.000 claims 1
- 239000008186 active pharmaceutical agent Substances 0.000 claims 1
- 229940088679 drug related substance Drugs 0.000 claims 1
- 229940010454 licorice Drugs 0.000 claims 1
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 24
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- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 4
- 241000196324 Embryophyta Species 0.000 description 4
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- CWVRJTMFETXNAD-GMZLATJGSA-N 5-Caffeoyl quinic acid Natural products O[C@H]1C[C@](O)(C[C@H](OC(=O)C=Cc2ccc(O)c(O)c2)[C@@H]1O)C(=O)O CWVRJTMFETXNAD-GMZLATJGSA-N 0.000 description 2
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 2
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- 239000000843 powder Substances 0.000 description 2
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- QAIPRVGONGVQAS-DUXPYHPUSA-N trans-caffeic acid Chemical compound OC(=O)\C=C\C1=CC=C(O)C(O)=C1 QAIPRVGONGVQAS-DUXPYHPUSA-N 0.000 description 2
- AAWZDTNXLSGCEK-LNVDRNJUSA-N (3r,5r)-1,3,4,5-tetrahydroxycyclohexane-1-carboxylic acid Chemical compound O[C@@H]1CC(O)(C(O)=O)C[C@@H](O)C1O AAWZDTNXLSGCEK-LNVDRNJUSA-N 0.000 description 1
- ACEAELOMUCBPJP-UHFFFAOYSA-N (E)-3,4,5-trihydroxycinnamic acid Natural products OC(=O)C=CC1=CC(O)=C(O)C(O)=C1 ACEAELOMUCBPJP-UHFFFAOYSA-N 0.000 description 1
- 235000017166 Bambusa arundinacea Nutrition 0.000 description 1
- 235000017491 Bambusa tulda Nutrition 0.000 description 1
- AAWZDTNXLSGCEK-UHFFFAOYSA-N Cordycepinsaeure Natural products OC1CC(O)(C(O)=O)CC(O)C1O AAWZDTNXLSGCEK-UHFFFAOYSA-N 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 244000082204 Phyllostachys viridis Species 0.000 description 1
- 235000015334 Phyllostachys viridis Nutrition 0.000 description 1
- AAWZDTNXLSGCEK-ZHQZDSKASA-N Quinic acid Natural products O[C@H]1CC(O)(C(O)=O)C[C@H](O)C1O AAWZDTNXLSGCEK-ZHQZDSKASA-N 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 230000004103 aerobic respiration Effects 0.000 description 1
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- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 230000000259 anti-tumor effect Effects 0.000 description 1
- 230000000840 anti-viral effect Effects 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 239000011425 bamboo Substances 0.000 description 1
- 230000000975 bioactive effect Effects 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 230000036772 blood pressure Effects 0.000 description 1
- 235000004883 caffeic acid Nutrition 0.000 description 1
- 229940074360 caffeic acid Drugs 0.000 description 1
- 210000002421 cell wall Anatomy 0.000 description 1
- 210000003169 central nervous system Anatomy 0.000 description 1
- QAIPRVGONGVQAS-UHFFFAOYSA-N cis-caffeic acid Natural products OC(=O)C=CC1=CC=C(O)C(O)=C1 QAIPRVGONGVQAS-UHFFFAOYSA-N 0.000 description 1
- 238000005352 clarification Methods 0.000 description 1
- 238000004440 column chromatography Methods 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- YTJJRAWFHJBAMT-UHFFFAOYSA-N depside Natural products OC(=O)CC1=C(O)C=C(O)C=C1OC(=O)C1=CC=C(O)C(O)=C1 YTJJRAWFHJBAMT-UHFFFAOYSA-N 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000006025 fining agent Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 238000005342 ion exchange Methods 0.000 description 1
- 210000000265 leukocyte Anatomy 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000013508 migration Methods 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 238000005457 optimization Methods 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 239000003208 petroleum Substances 0.000 description 1
- 229930015704 phenylpropanoid Natural products 0.000 description 1
- -1 phenylpropanoid compounds Chemical class 0.000 description 1
- 238000002203 pretreatment Methods 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- JXOHGGNKMLTUBP-HSUXUTPPSA-N shikimic acid Chemical compound O[C@@H]1CC(C(O)=O)=C[C@@H](O)[C@H]1O JXOHGGNKMLTUBP-HSUXUTPPSA-N 0.000 description 1
- JXOHGGNKMLTUBP-JKUQZMGJSA-N shikimic acid Natural products O[C@@H]1CC(C(O)=O)=C[C@H](O)[C@@H]1O JXOHGGNKMLTUBP-JKUQZMGJSA-N 0.000 description 1
- 238000000638 solvent extraction Methods 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 229940126680 traditional chinese medicines Drugs 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C67/00—Preparation of carboxylic acid esters
- C07C67/48—Separation; Purification; Stabilisation; Use of additives
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C67/00—Preparation of carboxylic acid esters
- C07C67/48—Separation; Purification; Stabilisation; Use of additives
- C07C67/56—Separation; Purification; Stabilisation; Use of additives by solid-liquid treatment; by chemisorption
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C69/00—Esters of carboxylic acids; Esters of carbonic or haloformic acids
- C07C69/66—Esters of carboxylic acids having esterified carboxylic groups bound to acyclic carbon atoms and having any of the groups OH, O—metal, —CHO, keto, ether, acyloxy, groups, groups, or in the acid moiety
- C07C69/73—Esters of carboxylic acids having esterified carboxylic groups bound to acyclic carbon atoms and having any of the groups OH, O—metal, —CHO, keto, ether, acyloxy, groups, groups, or in the acid moiety of unsaturated acids
- C07C69/732—Esters of carboxylic acids having esterified carboxylic groups bound to acyclic carbon atoms and having any of the groups OH, O—metal, —CHO, keto, ether, acyloxy, groups, groups, or in the acid moiety of unsaturated acids of unsaturated hydroxy carboxylic acids
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
- Tea And Coffee (AREA)
- Treatment Of Liquids With Adsorbents In General (AREA)
- Extraction Or Liquid Replacement (AREA)
- Coloring Foods And Improving Nutritive Qualities (AREA)
Description
本発明は、杜仲葉からクロロゲン酸を抽出する方法に関する。 The present invention relates to a method for extracting chlorogenic acid from Tochu-nakaba.
クロロゲン酸（Ｃｈｌｏｒｏｇｅｎｉｃ ａｃｉｄ）はカフェ酸（Ｃａｆｆｅｉｃ ａｃｉｄ）とキナ酸（Ｑｕｉｎｉｃ ａｃｉｄ）とで組成されるデプシドであり、別名はカフェタンニン酸、化学名は３−Ｏ−カフェオイルキナ酸（３−Ｏ−Ｃａｆｆｅｏｙｌｑｕｉｎｉｃ ａｃｉｄ）、分子式はＣ１６Ｈ１８Ｏ９、分子量は３５４．３０であって、植物体が好気呼吸の過程においてシキミ酸経路で生成するフェニルプロパノイド類である。クロロゲン酸は重要な生物活性物質であり、抗菌、抗ウィルス、白血球増殖、肝保護・利胆、抗腫瘍、血圧降下、血中脂質降下、活性酸素除去、中枢神経系興奮等の作用がある。クロロゲン酸は多くの漢方薬の有効成分のひとつであり、一部の売薬の品質指標でもある。クロロゲン酸の応用範囲は非常に広く、主に医薬、日用化学工業品、食品等の業界で応用されている。クロロゲン酸は様々な植物に広く存在するが、含有量が高い植物は決して多くはない。近年の研究では、杜仲葉のクロロゲン酸含有量が豊富で、１％〜５％に達することがわかっている。中国では杜仲が広範囲に植林されており、毎年数百万トンの杜仲葉を生産することができる。これはクロロゲン酸の抽出に十分な原料のベースとなっている。 Chlorogenic acid is a depside composed of caffeic acid and quinic acid, also known as caffetannic acid, chemical name 3-O-caffeoylquinic acid (3-O -Caffeoylquinic acid), molecular formula C 16 H 18 O 9, the molecular weight a 354.30 a phenylpropanoid compounds that plants generated by the shikimate pathway in the process of aerobic respiration. Chlorogenic acid is an important bioactive substance, and has antibacterial, antiviral, leukocyte proliferation, hepatoprotection / biliary, antitumor, blood pressure lowering, blood lipid lowering, active oxygen removal, central nervous system excitement and the like. Chlorogenic acid is one of the active ingredients of many traditional Chinese medicines, and is also a quality indicator for some medicines. The application range of chlorogenic acid is very wide, and it is mainly applied in industries such as pharmaceuticals, daily chemical products and foods. Chlorogenic acid is widely present in various plants, but not many plants have a high content. Recent studies have shown that the chlorogenic acid content of Tochu Naka is abundant, reaching 1% to 5%. In China, Tochu is planted extensively and can produce millions of tons of Tochu every year. This is the basis for a raw material sufficient for extraction of chlorogenic acid.
現在、杜仲葉からクロロゲン酸を抽出するという報道は多く、クロロゲン酸の抽出方法は主として有機溶媒抽出法、物理的抽出法による抽出、超高圧抽出法、ホモジネート抽出法等がある。銭ホアほか、「杜仲葉クロロゲン酸の抽出分離」、中国野生植物資源、第２０巻第４期（銭ホア等、「杜仲葉緑原酸的提取分離」、中国野生植物資源、第２０巻第４期）では、以下が紹介されている。杜仲葉の乾燥粉末を、７０％エタノール（ｐＨ＝２．５）を用いて９０℃で３回抽出する。１回当たり３０分抽出し、固液比をそれぞれ１：１２、１：１０、１：８とする。抽出液を一定の体積になるまで濃縮し、前処理した後に投入する。同時に、１７種類のマクロポーラス吸着樹脂について、杜仲葉クロロゲン酸に対する吸着能、溶離能及び吸着動力学特性を比較する、というものである。また、スクリーニング試験は、ＮＫＡ−９が優れた性能の吸着剤であることを示している。王茜ほか、「杜仲葉クロロゲン酸抽出分離の工程条件と研究」、イオン交換と吸着、２００８、２４（１）：７３〜８０（王茜等、「杜仲葉中緑原酸提取分離工芸条件的研究」、離子交換与吸附、２００８、２４（１）：７３〜８０）では、杜仲葉クロロゲン酸の抽出と分離について研究を行っている。すなわち、水及び濃度の異なるエタノール、メタノール、アセトン水溶液を抽出溶媒として、クロロゲン酸得率への影響を研究しており、直交実験法を用いてクロロゲン酸抽出率に影響を与える主要因を分析し、またマクロポーラス樹脂でそれを分離している。その結果、以下のことが示された。５０℃の水により抽出されたクロロゲン酸の得率が比較的高く、１．０６％であったことから、水をクロロゲン酸抽出溶媒とする。また、水による杜仲葉クロロゲン酸抽出の最適な工程条件は、温度６０℃、固液比１：１６、ｐＨ４、抽出時間３ｈであり、スクリーニングされたＧＣ−Ｉ樹脂はクロロゲン酸の吸着分離に最適な吸着剤であって、吸着に最適なｐＨ値は３、吸着流速は３ＢＶ／ｈであった。この実験条件で得られた粗生成物の純度は３０．８８％、収率は７６．５１％であった。苗磊、「杜仲葉クロロゲン酸の抽出と純化」、北京化工大学、２０１１年６月６日（苗磊、「杜仲葉中緑原酸的提取和純化」、北京化工大学、２０１１年６月６日）によって、以下の内容が開示されている。まず水で杜仲葉粉末に対しクロロゲン酸の抽出を行い、抽出液をＮＫＡ−９マクロポーラス樹脂によって純化し、溶媒を蒸発させてクロロゲン酸生成物を得る。抽出残渣物を乾燥させた後、アルカリ液浸出により細胞壁を分解し、その後石油エーテルの熱還流で抽出した杜仲樹脂を乾燥させる。抽出液からゲル法で粗樹脂を得、アセトン洗浄で乳白色の精製樹脂を得て、杜仲樹脂生成物とする。李光鋒、「杜仲葉クロロゲン酸の抽出、純化の研究」、湖南農業大学、２００６年１０月（李光鋒、「杜仲葉中緑原酸的提取、純化研究」、湖南農業大学、２００６年１０月）の、２．クロロゲン酸の安定性についての研究の結果から、クロロゲン酸は６０℃以内で４〜６ｈは比較的安定することが示された。アルカリ性の条件において分解しやすく、ｐＨ＝３の酸性の条件において最も安定する。クロロゲン酸は強い光照射のもとでは不安定であるため、クロロゲン酸の抽出・分離は極力酸性、遮光の条件を維持して行う。クロロゲン酸の抽出溶媒及び抽出方法は、結果として水を抽出溶媒として加熱抽出するのが比較的好ましいことがわかった。その後、直交試験によって温度、時間、固液比及び粒度の４要素について考察した。加熱抽出工程を最適化した結果を以下に示す。温度６５℃、抽出時間６０ｍｉｎ、固液比１：２５、粒度４０メッシュの場合が比較的良好であった。クロロゲン酸のカラムクロマトグラフィ最適化条件の研究では、吸着剤、溶離剤等について研究した結果、ＫＬＦＣ−１５０樹脂を吸着剤とし、エタノールを溶離剤とするのが最適条件であり、この方法でクロロゲン酸を分離すれば、純度、収率ともに高くなり、純度は５６．６７％に、収率は７８．３７％に達することが示された。 At present, there are many reports that chlorogenic acid is extracted from Tochu leaves, and there are mainly chlorogenic acid extraction methods such as organic solvent extraction method, physical extraction method, ultrahigh pressure extraction method, and homogenate extraction method. Qian Hua et al., “Extraction and separation of Chung Naka leaf chlorogenic acid”, Chinese wild plant resources, Volume 20, Volume 4 (Qian Hua et al. In the 4th term, the following are introduced. The dried powder of Tochu Nakaba is extracted three times at 90 ° C. with 70% ethanol (pH = 2.5). Extraction is performed for 30 minutes per time, and the solid-liquid ratio is set to 1:12, 1:10, and 1: 8, respectively. The extract is concentrated to a constant volume and added after pretreatment. At the same time, the adsorbing ability, elution ability and adsorption kinetic characteristics of 17 kinds of macroporous adsorbent resins with respect to chunaka chlorogenic acid are compared. Screening tests also show that NKA-9 is an adsorbent with excellent performance. Wang Yin et al., “Process conditions and research for extraction and separation of Chung Nakaba chlorogenic acid”, ion exchange and adsorption, 2008, 24 (1): 73-80 (Wang Yu et al. "Research", baby-on exchange and suction, 2008, 24 (1): 73-80), researches on the extraction and separation of Tochu Nakaba chlorogenic acid. In other words, water and ethanol, methanol, and acetone aqueous solutions with different concentrations are used as extraction solvents to study the effect on chlorogenic acid yield, and the main factors affecting chlorogenic acid extraction rate are analyzed using the orthogonal experiment method. Also, it is separated with macroporous resin. As a result, the following was shown. Since the yield of chlorogenic acid extracted with water at 50 ° C. was relatively high, 1.06%, water was used as the chlorogenic acid extraction solvent. In addition, the optimum process conditions for extraction of chunaka chlorogenic acid with water are a temperature of 60 ° C., a solid-liquid ratio of 1:16, a pH of 4 and an extraction time of 3 h, and the screened GC-I resin is optimal for adsorption separation of chlorogenic acid. The optimum pH value for adsorption was 3, and the adsorption flow rate was 3 BV / h. The purity of the crude product obtained under these experimental conditions was 30.88%, and the yield was 76.51%. Miao, “Extraction and Purification of Chung Nakaba Chlorogenic Acid”, Beijing Chemical University, June 6, 2011 (Miao Xiao, “Purchasing and Purifying the Green Harmonic Acid Proposal”, Beijing Chemical University, June 6, 2011 The following content is disclosed. First, chlorogenic acid is extracted from the rice bran powder with water, the extract is purified with NKA-9 macroporous resin, and the solvent is evaporated to obtain a chlorogenic acid product. After the extraction residue is dried, the cell wall is decomposed by leaching with an alkaline solution, and then the Tochu resin extracted by hot reflux of petroleum ether is dried. A crude resin is obtained from the extract by a gel method, and a milky white purified resin is obtained by washing with acetone to obtain a Tochu resin product. Lee Mitsuobu, “Research on Extraction and Purification of Chungchuan Chlorogenic Acid”, Hunan Agricultural University, October 2006 (Li Kwanghae, “Liquor Nakanaka Green Acid Procurement, Purification Research”, Hunan Agricultural University, October 2006) 2. The results of studies on the stability of chlorogenic acid showed that chlorogenic acid is relatively stable within 60 ° C. for 4-6 h. It is easily decomposed under alkaline conditions and is most stable under acidic conditions at pH = 3. Since chlorogenic acid is unstable under strong light irradiation, extraction / separation of chlorogenic acid should be performed under the conditions of acidity and shading as much as possible. As a result, it has been found that the extraction solvent and extraction method for chlorogenic acid are relatively preferably subjected to heat extraction using water as an extraction solvent. Thereafter, four elements of temperature, time, solid-liquid ratio, and particle size were examined by an orthogonal test. The result of optimizing the heating extraction process is shown below. The case of a temperature of 65 ° C., an extraction time of 60 min, a solid-liquid ratio of 1:25, and a particle size of 40 mesh was relatively good. In the study of optimization conditions for column chromatography of chlorogenic acid, as a result of research on adsorbents, eluents, etc., the optimum condition is that KLFC-150 resin is the adsorbent and ethanol is the eluent. As a result, it was shown that both the purity and the yield were high, and the purity reached 56.67% and the yield reached 78.37%.
ところで、従来の抽出方法では、クロロゲン酸が濃縮のために長時間水中で熱にさらされ、不安定になって分解してしまうという問題がある。また、カラム投入前の濃縮工程において多数の工程を経なければならないため、生産サイクルを短縮することができない。 By the way, the conventional extraction method has a problem that chlorogenic acid is exposed to heat in water for a long time for concentration, becomes unstable and decomposes. In addition, the production cycle cannot be shortened because many steps must be performed in the concentration step before the column is charged.
本発明は、杜仲葉からクロロゲン酸を抽出する方法を提供する。 The present invention provides a method for extracting chlorogenic acid from Tochu leaves.
本発明は、以下のステップａ〜ｇを含む、杜仲葉からクロロゲン酸を抽出する方法。 The present invention is a method for extracting chlorogenic acid from Tochu leaves, comprising the following steps a to g.
ａ．杜仲葉の秤取。 a. Weighing of Nakanaka.
ｂ．水抽出：水を加えて抽出する。抽出温度は４０℃〜８０℃、抽出時間は０．５ｈ〜４．０ｈ、抽出回数は１〜３回、１回当たりの水の使用量は薬材の質量の６〜１４倍、ｐＨ値は２〜７とし、浸出又は動的抽出を行う。 b. Water extraction: Extract by adding water. Extraction temperature is 40 ° C to 80 ° C, extraction time is 0.5h to 4.0h, number of extractions is 1 to 3 times, the amount of water used per time is 6 to 14 times the mass of the drug, pH value is 2-7 and perform leaching or dynamic extraction.
ｃ．前処理：清澄剤を加える。添加量は薬材の質量の０．２％〜１．０％、沈降温度は４０℃〜８０℃、沈降時間は０．５ｈ〜２４ｈとする。 c. Pretreatment: Add clarifier. The addition amount is 0.2% to 1.0% of the mass of the chemical, the sedimentation temperature is 40 ° C. to 80 ° C., and the sedimentation time is 0.5 h to 24 h.
ｄ．ろ過：ステップｃの前処理液を、孔径５０μｍ〜２００μｍのろ過膜又はフィルターコアでろ過する。 d. Filtration: The pretreatment liquid in step c is filtered through a filtration membrane or filter core having a pore diameter of 50 μm to 200 μm.
ｅ．マクロポーラス吸着樹脂への投入：ステップｄのろ過液を、非極性又は弱極性のマクロポーラス吸着樹脂に通す。樹脂カラムの直径と高さの比は１：４〜１：１０、投入の流速は２．０〜８．０ＢＶ／Ｈ、吸着量は、薬材の質量とカラム体積の比が１：１〜１：５とする。洗浄水の使用量は０．５〜２．０ＢＶ、洗浄水の流速は２．０〜８．０ＢＶ／Ｈ、溶離溶媒の種類はメタノール又はエタノールとし、その濃度は１０〜８０％、溶離液の使用量は１．０〜６．０ＢＶ、溶離液の流速は２．０〜８．０ＢＶ／Ｈとする。 e. Charge to macroporous adsorption resin: The filtrate of step d is passed through a non-polar or weakly polar macroporous adsorption resin. The ratio of the diameter and height of the resin column is 1: 4 to 1:10, the flow rate of charging is 2.0 to 8.0 BV / H, and the adsorption amount is the ratio of the mass of the chemical to the column volume is 1: 1 to 1. 1: 5. The amount of wash water used is 0.5 to 2.0 BV, the flow rate of wash water is 2.0 to 8.0 BV / H, the type of elution solvent is methanol or ethanol, and its concentration is 10 to 80%. The amount used is 1.0 to 6.0 BV, and the flow rate of the eluent is 2.0 to 8.0 BV / H.
ｆ．濃縮：濃縮温度は４０℃〜８０℃、真空度は−０．０４ＭＰ〜−０．０９ＭＰ、相対密度は１．０２〜１．１０とする。 f. Concentration: The concentration temperature is 40 ° C. to 80 ° C., the degree of vacuum is −0.04 MP to −0.09 MP, and the relative density is 1.02 to 1.10.
ｇ．乾燥：空気入口温度１５０℃〜２００℃、空気出口温度５０℃〜９０℃である噴霧乾燥をして、クロロゲン酸の粗生成物を得る。 g. Drying: Spray drying at an air inlet temperature of 150 ° C. to 200 ° C. and an air outlet temperature of 50 ° C. to 90 ° C. yields a crude product of chlorogenic acid.
得られたクロロゲン酸の粗生成物をさらに精製、純化して、クロロゲン酸の純粋品を得る。 The obtained crude product of chlorogenic acid is further purified and purified to obtain a pure product of chlorogenic acid.
より好ましくは、以下のステップａ〜ｇを含む。 More preferably, the following steps a to g are included.
ａ．杜仲葉の秤取。 a. Weighing of Nakanaka.
ｂ．水抽出：水を加えて抽出する。抽出温度は６０℃〜８０℃、抽出時間は０．５ｈ〜１．５ｈ、抽出回数は１〜３回、１回当たりの水の使用量は薬材の質量の１０〜１４倍、ｐＨ値は６〜７とし、浸出又は動的抽出を行う。 b. Water extraction: Extract by adding water. Extraction temperature is 60 ° C to 80 ° C, extraction time is 0.5h to 1.5h, number of extractions is 1 to 3 times, the amount of water used per time is 10 to 14 times the mass of the drug, pH value is 6-7 and perform leaching or dynamic extraction.
ｃ．前処理：清澄剤を加える。添加量は薬材の質量の０．４％〜０．８％、沈降温度は５０℃〜７０℃、沈降時間は１．０ｈ〜２．０ｈとする。 c. Pretreatment: Add clarifier. The addition amount is 0.4% to 0.8% of the mass of the chemical material, the sedimentation temperature is 50 ° C. to 70 ° C., and the sedimentation time is 1.0 h to 2.0 h.
ｄ．ろ過：ステップｃの前処理液を、孔径５０μｍ〜１００μｍのろ過膜又はフィルターコアでろ過する。 d. Filtration: The pretreatment liquid in step c is filtered through a filtration membrane or filter core having a pore size of 50 to 100 μm.
ｅ．マクロポーラス吸着樹脂への投入：ステップｄのろ過液を、非極性又は弱極性のマクロポーラス吸着樹脂に通す。樹脂カラムの直径と高さの比は１：４〜１：８、投入流速は３．０〜５．０ＢＶ／Ｈ、吸着量は、薬材の質量とカラム体積の比が１：１〜１：４とする。洗浄水の使用量は１．０〜２．０ＢＶ、洗浄水の流速は３．０〜５．０ＢＶ／Ｈ、溶離溶媒の種類はメタノール又はエタノールとし、その濃度は１０〜３０％、溶離液の使用量は４．０〜６．０ＢＶ、溶離液の流速は２．０〜４．０ＢＶ／Ｈとする。 e. Charge to macroporous adsorption resin: The filtrate of step d is passed through a non-polar or weakly polar macroporous adsorption resin. The ratio of the diameter and height of the resin column is 1: 4 to 1: 8, the input flow rate is 3.0 to 5.0 BV / H, and the adsorption amount is the ratio of the mass of the chemical to the column volume is 1: 1 to 1. : 4. The amount of wash water used is 1.0 to 2.0 BV, the flow rate of wash water is 3.0 to 5.0 BV / H, the type of elution solvent is methanol or ethanol, and its concentration is 10 to 30%. The amount used is 4.0 to 6.0 BV, and the flow rate of the eluent is 2.0 to 4.0 BV / H.
ｆ．濃縮：濃縮温度は５０℃〜７０℃、真空度は−０．０７ＭＰ〜−０．０９ＭＰ、相対密度は１．０３〜１．０５とする。 f. Concentration: The concentration temperature is 50 ° C. to 70 ° C., the degree of vacuum is −0.07 MP to −0.09 MP, and the relative density is 1.03 to 1.05.
ｇ．乾燥：空気入口温度１５０℃〜１７０℃、空気出口温度６０℃〜８０℃である噴霧乾燥をして、クロロゲン酸の粗生成物を得る。 g. Drying: Spray drying at an air inlet temperature of 150 ° C to 170 ° C and an air outlet temperature of 60 ° C to 80 ° C is performed to obtain a crude product of chlorogenic acid.
ただし、ステップａで述べる杜仲葉にはクロロゲン酸が１．０〜５．０％含まれ、水分含有量は≦２０％とする。 However, the bamboo leaf described in step a contains 1.0 to 5.0% of chlorogenic acid and the water content is ≦ 20%.
ただし、ステップｃで述べる清澄剤の種類はキトサンとする。 However, the type of fining agent described in step c is chitosan.
ただし、ステップｅで述べるマクロポーラス吸着樹脂の型番は、非極性又は弱極性のＨＰＤ−１００、ＨＰＤ−４５０、ＨＰＤ−７００、ＮＫＡ−ＩＩ、ＮＫＡ−９又はＬＳ−４６とし、好ましくは、前記マクロポーラス吸着樹脂はＨＰＤ−１００、ＨＰＤ−４５０、又はＬＳ−４６とする。 However, the model number of the macroporous adsorption resin described in step e is non-polar or weakly polar HPD-100, HPD-450, HPD-700, NKA-II, NKA-9 or LS-46, preferably the macro The porous adsorption resin is HPD-100, HPD-450, or LS-46.
本発明は、水抽出、前処理、マクロポーラス吸着樹脂カラムへの投入、濃縮乾燥によってクロロゲン酸粗生成物を調製するものである。水抽出液は濃縮せずに、前処理後、直接マクロポーラス吸着樹脂に投入して純化処理を行うことで、クロロゲン酸が濃縮のために長時間水中で熱にさらされ、不安定になって分解してしまうという問題を解決するとともに、濃縮工程の段階を省いて生産サイクルを短縮する。また、異なる溶離液の濃度パラメータを制御することで、２０〜６０％の各種規格の粗生成物を生産することができる。 In the present invention, a chlorogenic acid crude product is prepared by water extraction, pretreatment, charging into a macroporous adsorption resin column, and concentration drying. Without pre-concentrating the water extract, after pre-treatment, the chlorogenic acid is exposed to heat in water for a long period of time and becomes unstable due to the purification process by directly adding it to the macroporous adsorption resin. In addition to solving the problem of decomposition, it eliminates the concentration step and shortens the production cycle. In addition, by controlling the concentration parameters of different eluents, it is possible to produce 20-60% crude products of various standards.
実施例１ 本発明のクロロゲン酸を抽出する方法
１．抽出
杜仲葉を１０ｋｇ（クロロゲン酸含有量＝１．０％）用意し、水を用いて４０℃の条件下で３回抽出した。１回当たり４．０ｈ、１回当たりの水の使用量は６０Ｌ、ｐＨは２．０とし、各回の抽出液を合わせた。
Example 1 Method for Extracting Chlorogenic Acid of the Present Invention Extraction 10 kg (Chlorogenic acid content = 1.0%) was prepared and extracted three times with water at 40 ° C. The amount of water used per time was 4.0 L, the amount of water used per time was 60 L, the pH was 2.0, and the extracts were combined each time.
２．前処理
キトサンを１００ｇ用意し、１％の酢酸水溶液で２％のキトサン水溶液を調合し、４０℃の条件下で抽出液を加え、均一に攪拌した後、２４．０ｈ置いた。
2. Pretreatment 100 g of chitosan was prepared, 2% chitosan aqueous solution was prepared with 1% acetic acid aqueous solution, the extract was added under conditions of 40 ° C., stirred uniformly, and then placed for 24.0 h.
３．ろ過
前処理で得た液を５０μｍのろ過膜でろ過し、濾液を収集した。
3. Filtration The liquid obtained in the pretreatment was filtered through a 50 μm filtration membrane, and the filtrate was collected.
４．マクロポーラス吸着樹脂カラムへの投入
（１）投入：マクロポーラス吸着樹脂ＨＰＤ−１００を用意し、径と高さの比＝１：４の樹脂カラムとなるように湿式法でカラムに充填した。カラム体積は５０Ｌとし、前処理液を２．０ＢＶ／Ｈの流速で投入した。
（２）水洗浄：０．５ＢＶの水を用いて、２．０ＢＶ／Ｈの流速で洗浄した。
（３）溶離：１．０ＢＶの８０％メタノール溶液を用いて、２．０ＢＶ／Ｈの流速で溶離を行い、溶出液を収集した。
4). Input to Macroporous Adsorption Resin Column (1) Input: Macroporous adsorption resin HPD-100 was prepared, and the column was packed by a wet method so as to obtain a resin column with a ratio of diameter to height = 1: 4. The column volume was 50 L, and the pretreatment liquid was charged at a flow rate of 2.0 BV / H.
(2) Water washing: Washed with 0.5 BV water at a flow rate of 2.0 BV / H.
(3) Elution: Elution was performed using a 1.0 BV 80% methanol solution at a flow rate of 2.0 BV / H, and the eluate was collected.
５．濃縮
溶出液を温度４０℃、真空度−０．０４ＭＰの条件下でメタノール回収し、さらに濃縮液の相対密度が１．０２（２５℃）になるまで濃縮した。
5. Concentration The eluate was recovered with methanol under conditions of a temperature of 40 ° C. and a degree of vacuum of −0.04 MP, and further concentrated until the relative density of the concentrate reached 1.02 (25 ° C.).
６．噴霧乾燥
濃縮液を空気入口温度２００℃、空気出口温度９０℃で噴霧乾燥し、クロロゲン酸粗生成物を得た。
6). Spray drying The concentrate was spray dried at an air inlet temperature of 200 ° C and an air outlet temperature of 90 ° C to obtain a crude chlorogenic acid product.
７．生成物の特徴
得られたクロロゲン酸粗生成物の純度は２４．３％、生成物得率は３．２％、クロロゲン酸移行率７７．８％であった。
7). Product characteristics The purity of the obtained chlorogenic acid crude product was 24.3%, the product yield was 3.2%, and the chlorogenic acid transfer rate was 77.8%.
実施例２ 本発明のクロロゲン酸を抽出する方法
１．抽出
杜仲葉を１０ｋｇ（クロロゲン酸含有量＝３．６％）用意し、水を用いて８０℃の条件下で１回抽出した。抽出時間は０．５ｈ、水の使用量は１４０Ｌ、ｐＨは７．０とした。
Example 2 Method for Extracting Chlorogenic Acid of the Present Invention Extraction 10 kg of chunaka leaves (chlorogenic acid content = 3.6%) were prepared and extracted once with water at 80 ° C. The extraction time was 0.5 h, the amount of water used was 140 L, and the pH was 7.0.
２．前処理
キトサンを２０ｇ用意し、１％の酢酸水溶液で２％のキトサン水溶液を調合し、８０℃の条件下で抽出液を加え、均一に攪拌した後、０．５ｈ置いた。
2. Pretreatment 20 g of chitosan was prepared, 2% chitosan aqueous solution was prepared with 1% acetic acid aqueous solution, the extract was added at 80 ° C., stirred uniformly, and then placed for 0.5 h.
３．ろ過
前処理で得た液を２００μｍのろ過膜でろ過し、濾液を収集した。
3. Filtration The liquid obtained by pretreatment was filtered through a 200 μm filtration membrane, and the filtrate was collected.
４．マクロポーラス吸着樹脂カラムへの投入
（１）投入：マクロポーラス吸着樹脂ＨＰＤ−４５０を用意し、径と高さの比＝１：１０の樹脂カラムとなるように湿式法でカラムに充填した。カラム体積は１０Ｌとし、前処理液を２．０ＢＶ／Ｈの流速で投入した。
（２）水洗浄：２．０ＢＶの水を用いて、２．０ＢＶ／Ｈの流速で洗浄した。
（３）溶離：６．０ＢＶの１０％エタノール溶液を用いて、２．０ＢＶ／Ｈの流速で溶離を行い、溶出液を収集した。
4). Input to Macroporous Adsorption Resin Column (1) Input: Macroporous adsorption resin HPD-450 was prepared, and the column was packed by a wet method so as to obtain a resin column with a ratio of diameter to height = 1: 10. The column volume was 10 L, and the pretreatment liquid was charged at a flow rate of 2.0 BV / H.
(2) Water washing: Washed with 2.0 BV water at a flow rate of 2.0 BV / H.
(3) Elution: Elution was performed using a 6.0 BV 10% ethanol solution at a flow rate of 2.0 BV / H, and the eluate was collected.
５．濃縮
溶出液を温度８０℃、真空度−０．０９ＭＰの条件下でエタノール回収し、さらに濃縮液の相対密度が１．１０（２５℃）になるまで濃縮した。
5. Concentration The eluate was recovered with ethanol under conditions of a temperature of 80 ° C. and a degree of vacuum of −0.09 MP, and further concentrated until the relative density of the concentrate reached 1.10 (25 ° C.).
６．噴霧乾燥
濃縮液を空気入口温度１５０℃、空気出口温度５０℃で噴霧乾燥し、クロロゲン酸粗生成物を得た。
6). Spray drying The concentrate was spray dried at an air inlet temperature of 150 ° C and an air outlet temperature of 50 ° C to obtain a crude product of chlorogenic acid.
７．生成物の特徴
得られたクロロゲン酸粗生成物の純度は５８．４％、生成物得率は４．８％、クロロゲン酸移行率は７７．９％であった。
7). Product characteristics The purity of the obtained chlorogenic acid crude product was 58.4%, the product yield was 4.8%, and the chlorogenic acid transfer rate was 77.9%.
実施例３ 本発明のクロロゲン酸を抽出する方法
１．抽出
杜仲葉を１０ｋｇ（クロロゲン酸含有量＝５．０％）用意し、水を用いて６０℃の条件下で２回抽出した。１回当たり１．５ｈ、１回当たりの水の使用量は８０Ｌ、ｐＨ値は３．０とし、各回の抽出液を合わせた。
Example 3 Method for Extracting Chlorogenic Acid of the Present Invention Extraction 10 kg (Chlorogenic acid content = 5.0%) of Tochu-nakaba was prepared and extracted twice with water at 60 ° C. The amount of water used per time was 1.5 L, the amount of water used was 80 L, the pH value was 3.0, and the extracts were combined each time.
２．前処理
キトサンを４０ｇ用意し、１％の酢酸水溶液で２％のキトサン水溶液を調合し、６０℃の条件下で抽出液を加え、均一に攪拌した後、１２．０ｈ置いた。
2. Pretreatment 40 g of chitosan was prepared, 2% chitosan aqueous solution was prepared with 1% acetic acid aqueous solution, the extract was added under the condition of 60 ° C., stirred uniformly, and placed for 12.0 h.
３．ろ過
前処理で得た液を２００μｍのろ過膜でろ過し、濾液を収集した。
3. Filtration The liquid obtained by pretreatment was filtered through a 200 μm filtration membrane, and the filtrate was collected.
４．マクロポーラス吸着樹脂カラムへの投入
（１）投入：マクロポーラス吸着樹脂ＨＰＤ−７００を用意し、径と高さの比＝１：８の樹脂カラムとなるように湿式法でカラムに充填した。カラム体積は２０Ｌとし、前処理液を３．０ＢＶ／Ｈの流速で投入した。
（２）水洗浄：１．０ＢＶの水を用いて、３．０ＢＶ／Ｈの流速で洗浄した。
（３）溶離：５．０ＢＶの２０％エタノール溶液を用いて、３．０ＢＶ／Ｈの流速で溶離を行い、溶出液を収集した。
4). Input to Macroporous Adsorption Resin Column (1) Input: Macroporous adsorption resin HPD-700 was prepared, and the column was packed by a wet method so as to obtain a resin column having a diameter to height ratio of 1: 8. The column volume was 20 L, and the pretreatment liquid was charged at a flow rate of 3.0 BV / H.
(2) Water washing: Washed with 1.0 BV water at a flow rate of 3.0 BV / H.
(3) Elution: Elution was performed using a 5.0 BV 20% ethanol solution at a flow rate of 3.0 BV / H, and the eluate was collected.
５．濃縮
溶出液を温度６０℃、真空度−０．０８ＭＰの条件下でエタノール回収し、さらに濃縮液の相対密度が１．０５０（２５℃）になるまで濃縮した。
5. Concentration The eluate was recovered with ethanol under conditions of a temperature of 60 ° C. and a degree of vacuum of −0.08 MP, and further concentrated until the relative density of the concentrate reached 1.050 (25 ° C.).
６．噴霧乾燥
濃縮液を空気入口温度１５０℃、空気出口温度７０℃で噴霧乾燥し、クロロゲン酸粗生成物を得た。
6). Spray drying The concentrate was spray dried at an air inlet temperature of 150 ° C and an air outlet temperature of 70 ° C to obtain a crude product of chlorogenic acid.
７．生成物の特徴
得られたクロロゲン酸粗生成物の純度は５２．４％、生成物得率は７．１％、クロロゲン酸移行率は７４．４％であった。
7). Product characteristics The purity of the obtained chlorogenic acid crude product was 52.4%, the product yield was 7.1%, and the chlorogenic acid transfer rate was 74.4%.
実施例４ 本発明のクロロゲン酸を抽出する方法
１．抽出
杜仲葉を１０ｋｇ（クロロゲン酸含有量＝３．６％）用意し、水を用いて６０℃の条件下で２回抽出した。１回当たり２．０ｈ、１回当たりの水の使用量は１００Ｌ、ｐＨ値は４．０とし、各回の抽出液を合わせた。
Example 4 Method for Extracting Chlorogenic Acid of the Present Invention Extraction 10 kg (Chlorogenic acid content = 3.6%) was prepared and extracted twice with water at 60 ° C. The amount of water used per time was 2.0 L, the amount of water used was 100 L, the pH value was 4.0, and the extracts were combined each time.
２．前処理
キトサンを６０ｇ用意し、１％の酢酸水溶液で２％のキトサン水溶液を調合し、６０℃の条件下で抽出液を加え、均一に攪拌した後、８．０ｈ置いた。
2. Pretreatment 60 g of chitosan was prepared, 2% chitosan aqueous solution was prepared with 1% acetic acid aqueous solution, the extract was added under conditions of 60 ° C., stirred uniformly, and then left for 8.0 h.
３．ろ過
前処理で得た液を５０μｍのろ過膜でろ過し、濾液を収集した。
3. Filtration The liquid obtained in the pretreatment was filtered through a 50 μm filtration membrane, and the filtrate was collected.
４．マクロポーラス吸着樹脂カラムへの投入
（１）投入：マクロポーラス吸着樹脂ＮＫＡ−ＩＩを用意し、径と高さの比＝１：８の樹脂カラムとなるように湿式法でカラムに充填した。カラム体積は４０Ｌとし、前処理液を５．０ＢＶ／Ｈの流速で投入した。
（２）水洗浄：２．０ＢＶの水を用いて、５．０ＢＶ／Ｈの流速で洗浄した。
（３）溶離：４．０ＢＶの３０％エタノール溶液を用いて、４．０ＢＶ／Ｈの流速で溶離を行い、溶出液を収集した。
4). Input to Macroporous Adsorption Resin Column (1) Input: Macroporous adsorption resin NKA-II was prepared, and the column was packed by a wet method so as to obtain a resin column with a ratio of diameter to height = 1: 8. The column volume was 40 L, and the pretreatment liquid was charged at a flow rate of 5.0 BV / H.
(2) Water washing: Washed with 2.0 BV water at a flow rate of 5.0 BV / H.
(3) Elution: Using a 30% ethanol solution of 4.0 BV, elution was performed at a flow rate of 4.0 BV / H, and the eluate was collected.
５．濃縮
溶出液を温度７０℃、真空度−０．０８ＭＰの条件下でエタノール回収し、さらに濃縮液の相対密度が１．０６（２５℃）になるまで濃縮した。
5. Concentration The eluate was recovered by ethanol under conditions of a temperature of 70 ° C. and a vacuum degree of −0.08 MP, and further concentrated until the relative density of the concentrate reached 1.06 (25 ° C.).
６．噴霧乾燥
濃縮液を空気入口温度１８０℃、空気出口温度８０℃で噴霧乾燥し、クロロゲン酸粗生成物を得た。
6). Spray drying The concentrate was spray dried at an air inlet temperature of 180 ° C and an air outlet temperature of 80 ° C to obtain a crude chlorogenic acid product.
７．生成物の特徴
得られたクロロゲン酸粗生成物の純度は４７．４％、生成物得率は５．６％、クロロゲン酸移行率は７３．７％であった。
7). Product characteristics The purity of the obtained chlorogenic acid crude product was 47.4%, the product yield was 5.6%, and the chlorogenic acid transfer rate was 73.7%.
実施例５ 本発明のクロロゲン酸を抽出する方法
１．抽出
杜仲葉を１０ｋｇ（クロロゲン酸含有量＝３．６％）用意し、水を用いて４０℃の条件下で３回抽出した。１回当たり４．０ｈ、１回当たりの水の使用量は８０Ｌ、ｐＨ値は３．０とし、各回の抽出液を合わせた。
Example 5 Method for Extracting Chlorogenic Acid of the Present Invention Extraction 10 kg (Chlorogenic acid content = 3.6%) was prepared and extracted three times with water at 40 ° C. The amount of water used per time was 4.0 L, the amount of water used per time was 80 L, the pH value was 3.0, and the extracts were combined each time.
２．前処理
キトサンを１００ｇ用意し、１％の酢酸水溶液で２％のキトサン水溶液を調合し、４０℃の条件下で抽出液を加え、均一に攪拌した後、２４．０置いた。
2. Pretreatment 100 g of chitosan was prepared, 2% chitosan aqueous solution was prepared with 1% acetic acid aqueous solution, the extract was added at 40 ° C., stirred uniformly, and then placed at 24.0.
３．ろ過
前処理で得た液を１００μｍのろ過膜でろ過し、濾液を収集した。
3. Filtration The liquid obtained in the pretreatment was filtered through a 100 μm filtration membrane, and the filtrate was collected.
４．マクロポーラス吸着樹脂カラムへの投入
（１）投入：マクロポーラス吸着樹脂ＮＫＡ−９を用意し、径と高さの比＝１：１０の樹脂カラムとなるように湿式法でカラムに充填した。カラム体積は５０Ｌとし、前処理液を８．０ＢＶ／Ｈの流速で投入した。
（２）水洗浄：２．０ＢＶの水を用いて、８．０ＢＶ／Ｈの流速で洗浄した。
（３）溶離：６．０ＢＶの４０％エタノール溶液を用いて、８．０ＢＶ／Ｈの流速で溶離を行い、溶出液を収集した。
4). Input to Macroporous Adsorption Resin Column (1) Input: Macroporous adsorption resin NKA-9 was prepared, and the column was packed by a wet method so as to obtain a resin column with a ratio of diameter to height = 1: 10. The column volume was 50 L, and the pretreatment liquid was charged at a flow rate of 8.0 BV / H.
(2) Water washing: Washed with 2.0 BV water at a flow rate of 8.0 BV / H.
(3) Elution: Elution was performed using a 6.0 BV 40% ethanol solution at a flow rate of 8.0 BV / H, and the eluate was collected.
５．濃縮
溶出液を温度５０℃、真空度−０．０６ＭＰの条件下でエタノール回収し、さらに濃縮液の相対密度が１．０８（２５℃）になるまで濃縮した。
5. Concentration The eluate was recovered by ethanol under conditions of a temperature of 50 ° C. and a vacuum degree of −0.06 MP, and further concentrated until the relative density of the concentrate reached 1.08 (25 ° C.).
６．噴霧乾燥
濃縮液を空気入口温度１６０℃、空気出口温度６０℃で噴霧乾燥し、クロロゲン酸粗生成物を得た。
6). Spray drying The concentrate was spray dried at an air inlet temperature of 160 ° C and an air outlet temperature of 60 ° C to obtain a crude chlorogenic acid product.
７．生成物の特徴
得られたクロロゲン酸粗生成物の純度は４５．８％、生成物得率は５．９％、クロロゲン酸移行率は７５．１％であった。
7). Product characteristics The purity of the obtained chlorogenic acid crude product was 45.8%, the product yield was 5.9%, and the chlorogenic acid transfer rate was 75.1%.
実施例６ 本発明のクロロゲン酸を抽出する方法
１．抽出
杜仲葉を１０ｋｇ（クロロゲン酸含有量＝３．６％）用意し、水を用いて６０℃の条件下で２回抽出した。１回当たり１．５ｈ、１回当たりの水の使用量は１２０Ｌ、ｐＨ値は６．５とし、各回の抽出液を合わせた。
Example 6 Method for Extracting Chlorogenic Acid of the Present Invention Extraction 10 kg (Chlorogenic acid content = 3.6%) was prepared and extracted twice with water at 60 ° C. The amount of water used per time was 1.5 L, the amount of water used was 120 L, the pH value was 6.5, and the extracts were combined each time.
２．前処理
キトサンを８０ｇ用意し、１％の酢酸水溶液で２％のキトサン水溶液を調合し、６０℃の条件下で抽出液を加え、均一に攪拌した後、２．０ｈ置いた。
2. Pretreatment 80 g of chitosan was prepared, 2% chitosan aqueous solution was prepared with 1% acetic acid aqueous solution, the extract was added at 60 ° C., stirred uniformly, and then left for 2.0 h.
３．ろ過
前処理で得た液を５０μｍのろ過膜でろ過し、濾液を収集した。
3. Filtration The liquid obtained in the pretreatment was filtered through a 50 μm filtration membrane, and the filtrate was collected.
４．マクロポーラス吸着樹脂カラムへの投入
（１）投入：マクロポーラス吸着樹脂ＮＫＡ−９を用意し、径と高さの比＝１：１０の樹脂カラムとなるように湿式法でカラムに充填した。カラム体積は４０Ｌとし、前処理液を６．０ＢＶ／Ｈの流速で投入した。
（２）水洗浄：１．０ＢＶの水を用いて、６．０ＢＶ／Ｈの流速で洗浄した。
（３）溶離：３．０ＢＶの５０％エタノール溶液を用いて、３．０ＢＶ／Ｈの流速で溶離を行い、溶出液を収集した。
4). Input to Macroporous Adsorption Resin Column (1) Input: Macroporous adsorption resin NKA-9 was prepared, and the column was packed by a wet method so as to obtain a resin column with a ratio of diameter to height = 1: 10. The column volume was 40 L, and the pretreatment liquid was charged at a flow rate of 6.0 BV / H.
(2) Water washing: Washed with 1.0 BV water at a flow rate of 6.0 BV / H.
(3) Elution: Elution was performed using a 3.0 BV 50% ethanol solution at a flow rate of 3.0 BV / H, and the eluate was collected.
５．濃縮
溶出液を温度６０℃、真空度−０．０８ＭＰの条件下でエタノール回収し、さらに濃縮液の相対密度が１．０４５（２５℃）になるまで濃縮した。
5. Concentration The eluate was recovered with ethanol under conditions of a temperature of 60 ° C. and a degree of vacuum of −0.08 MP, and further concentrated until the relative density of the concentrate reached 1.045 (25 ° C.).
６．噴霧乾燥
濃縮液を空気入口温度１６０℃、空気出口温度７０℃で噴霧乾燥し、クロロゲン酸粗生成物を得た。
6). Spray drying The concentrate was spray dried at an air inlet temperature of 160 ° C and an air outlet temperature of 70 ° C to obtain a crude product of chlorogenic acid.
７．生成物の特徴
得られたクロロゲン酸粗生成物の純度は４１．６％、生成物得率は６．２％、クロロゲン酸移行率は７１．６％であった。
7). Product characteristics The purity of the resulting chlorogenic acid crude product was 41.6%, the product yield was 6.2%, and the chlorogenic acid transfer rate was 71.6%.
実施例７ 本発明のクロロゲン酸を抽出する方法
１．抽出
杜仲葉を１０ｋｇ（クロロゲン酸含有量＝３．６％）用意し、水を用いて７０℃の条件下で２回抽出した。１回当たり１．０ｈ、１回当たりの水の使用量は１００Ｌ、ｐＨ値は７．０とし、各回の抽出液を合わせた。
Example 7 Method for Extracting Chlorogenic Acid of the Present Invention Extraction 10 kg (Chlorogenic acid content = 3.6%) was prepared and extracted twice with water at 70 ° C. The amount of water used per time was 1.0 L, the amount of water used per time was 100 L, the pH value was 7.0, and the extracts were combined each time.
２．前処理
キトサンを６０ｇ用意し、１％の酢酸水溶液で２％のキトサン水溶液を調合し、７０℃の条件下で抽出液を加え、均一に攪拌した後、２．０ｈ置いた。
2. Pretreatment 60 g of chitosan was prepared, 2% chitosan aqueous solution was prepared with 1% acetic acid aqueous solution, the extract was added at 70 ° C., stirred uniformly, and then left for 2.0 h.
３．ろ過
前処理で得た液を１００μｍのろ過膜でろ過し、濾液を収集した。
3. Filtration The liquid obtained in the pretreatment was filtered through a 100 μm filtration membrane, and the filtrate was collected.
４．マクロポーラス吸着樹脂カラムへの投入
（１）投入：マクロポーラス吸着樹脂ＬＳ−４６を用意し、径と高さの比＝１：８の樹脂カラムとなるように湿式法でカラムに充填した。カラム体積は２０Ｌとし、前処理液を５．０ＢＶ／Ｈの流速で投入した。
（２）水洗浄：１．０ＢＶの水を用いて、５．０ＢＶ／Ｈの流速で洗浄した。
（３）溶離：５．０ＢＶの２０％エタノール溶液を用いて、３．０ＢＶ／Ｈの流速で溶離を行い、溶出液を収集した。
4). Input to Macroporous Adsorption Resin Column (1) Input: Macroporous adsorption resin LS-46 was prepared, and the column was packed by a wet method so as to obtain a resin column with a ratio of diameter to height = 1: 8. The column volume was 20 L, and the pretreatment liquid was charged at a flow rate of 5.0 BV / H.
(2) Water wash: Washed with 1.0 BV water at a flow rate of 5.0 BV / H.
(3) Elution: Elution was performed using a 5.0 BV 20% ethanol solution at a flow rate of 3.0 BV / H, and the eluate was collected.
５．濃縮
溶出液を温度６０℃、真空度−０．０８ＭＰの条件下でエタノール回収し、さらに濃縮液の相対密度が１．０４５（２５℃）になるまで濃縮した。
5. Concentration The eluate was recovered with ethanol under conditions of a temperature of 60 ° C. and a degree of vacuum of −0.08 MP, and further concentrated until the relative density of the concentrate reached 1.045 (25 ° C.).
６．噴霧乾燥
濃縮液を空気入口温度１６０℃、空気出口温度７０℃で噴霧乾燥し、クロロゲン酸粗生成物を得た。
6). Spray drying The concentrate was spray dried at an air inlet temperature of 160 ° C and an air outlet temperature of 70 ° C to obtain a crude product of chlorogenic acid.
７．生成物の特徴
得られたクロロゲン酸粗生成物の純度は５５．８％、生成物得率は５．５％、クロロゲン酸移行率は８５．３％であった。
7). Product characteristics The purity of the obtained chlorogenic acid crude product was 55.8%, the product yield was 5.5%, and the chlorogenic acid transfer rate was 85.3%.
比較例１ クロロゲン酸を抽出する方法
１．抽出
杜仲葉を１０ｋｇ（クロロゲン酸含有量＝３．６％）用意し、水を用いて７０℃の条件下で２回抽出した。１回当たり１．０ｈ、１回当たりの水の使用量は１００Ｌ、ｐＨ値は７．０とし、各回の抽出液を合わせた。
Comparative Example 1 Method for extracting chlorogenic acid Extraction 10 kg (Chlorogenic acid content = 3.6%) was prepared and extracted twice with water at 70 ° C. The amount of water used per time was 1.0 L, the amount of water used per time was 100 L, the pH value was 7.0, and the extracts were combined each time.
２．濃縮
抽出液を温度６０℃、真空度−０．０８ＭＰの条件下で原液の半分まで濃縮した。
2. Concentration The extract was concentrated to half of the stock solution under the conditions of a temperature of 60 ° C. and a degree of vacuum of −0.08 MP.
３．前処理
キトサンを４０ｇ用意し、１％の酢酸水溶液で２％のキトサン水溶液を調合し、６０℃の条件下で抽出液を加え、均一に攪拌した後、２．０ｈ置いた。
3. Pretreatment 40 g of chitosan was prepared, a 2% chitosan aqueous solution was prepared with a 1% acetic acid aqueous solution, the extract was added at 60 ° C., stirred uniformly, and then placed for 2.0 h.
４．ろ過
前処理で得た液を１００μｍのろ過膜でろ過し、濾液を収集した。
4). Filtration The liquid obtained in the pretreatment was filtered through a 100 μm filtration membrane, and the filtrate was collected.
５．マクロポーラス吸着樹脂カラムへの投入
（１）投入：マクロポーラス吸着樹脂ＬＳ−４６を用意し、径と高さの比＝１：８の樹脂カラムとなるように湿式法でカラムに充填した。カラム体積は２０Ｌとし、前処理液を５．０ＢＶ／Ｈの流速で投入した。
（２）水洗浄：用１．０ＢＶの水を用いて、５．０ＢＶ／Ｈの流速で洗浄した。
（３）溶離：５．０ＢＶの２０％エタノール溶液を用いて、３．０ＢＶ／Ｈの流速で溶離を行い、溶出液を収集した。
5. Input to Macroporous Adsorption Resin Column (1) Input: Macroporous adsorption resin LS-46 was prepared, and the column was packed by a wet method so as to obtain a resin column with a ratio of diameter to height = 1: 8. The column volume was 20 L, and the pretreatment liquid was charged at a flow rate of 5.0 BV / H.
(2) Water washing: Washed at a flow rate of 5.0 BV / H using 1.0 BV water.
(3) Elution: Elution was performed using a 5.0 BV 20% ethanol solution at a flow rate of 3.0 BV / H, and the eluate was collected.
６．濃縮
溶出液を温度６０℃、真空度−０．０８ＭＰの条件下でエタノール回収し、さらに濃縮液の相対密度が１．０３（２５℃）になるまで濃縮した。
6). Concentration The eluate was recovered by ethanol under conditions of a temperature of 60 ° C. and a degree of vacuum of −0.08 MP, and further concentrated until the relative density of the concentrate reached 1.03 (25 ° C.).
７．噴霧乾燥
濃縮液を空気入口温度１６０℃、空気出口温度７０℃で噴霧乾燥し、クロロゲン酸粗生成物を得た。
7). Spray drying The concentrate was spray dried at an air inlet temperature of 160 ° C and an air outlet temperature of 70 ° C to obtain a crude product of chlorogenic acid.
８．生成物の特徴
得られたクロロゲン酸粗生成物の純度は４４．６％、生成物得率は４．５％、クロロゲン酸移行率は５５．８％であった。
8). Product characteristics The purity of the obtained chlorogenic acid crude product was 44.6%, the product yield was 4.5%, and the chlorogenic acid transfer rate was 55.8%.
本発明は水を抽出溶媒とし、キトサンの清澄処理を経た後、直接マクロポーラス吸着樹脂カラムに投入して溶離させ、その後濃縮し乾燥させるもので、工程は簡潔であり、作業性が高く、カラム投入前の濃縮工程において多数の工程を省くことで生産サイクルを短縮する。異なる溶離液の濃度パラメータを制御することで、２０〜６０％の各種規格の粗生成物を生産することができ、クロロゲン酸の移行率が７０〜９０％と高く、中でも実施例７の条件が最も好ましい。 In the present invention, water is used as an extraction solvent, and after passing through a clarification process of chitosan, it is directly put into a macroporous adsorption resin column for elution, and then concentrated and dried. The process is simple, the workability is high, The production cycle is shortened by omitting a number of processes in the concentration process before charging. By controlling the concentration parameters of different eluents, it is possible to produce 20-60% crude products of various standards, and the migration rate of chlorogenic acid is as high as 70-90%. Most preferred.
Claims (4)
ａ．杜仲葉の秤取、
ｂ．水抽出：水を加えて抽出し、抽出温度４０℃〜８０℃、抽出時間０．５ｈ〜４．０ｈ、抽出回数は１〜３回、１回当たりの水の使用量は薬材の質量の６〜１４倍、ｐＨ値は２〜７とし、浸出又は動的抽出を行う、
ｃ．前処理：水抽出液を濃縮せずに、清澄剤であるキトサンを加え、添加量は薬材の質量の０．２％〜１．０％、沈降温度は４０℃〜８０℃、沈降時間は０．５ｈ〜２４ｈとし、
ｄ．ろ過：ステップｃの前処理液を、孔径５０μｍ〜２００μｍのろ過膜又はフィルターコアでろ過する、
ｅ．マクロポーラス吸着樹脂への投入：ステップｄのろ過液を、非極性又は弱極性のマクロポーラス吸着樹脂に通し、樹脂カラムの直径と高さの比は１：４〜１：１０、投入の流速は２．０〜８．０ＢＶ／Ｈ、吸着量は、薬材の質量とカラム体積の比が１：１〜１：５とし、洗浄水の使用量は０．５〜２．０ＢＶ、洗浄水の流速は２．０〜８．０ＢＶ／Ｈ、溶離溶媒の種類はメタノール又はエタノールとし、その濃度は１０〜３０％、溶離液の使用量は１．０〜６．０ＢＶ、溶離液の流速は２．０〜８．０ＢＶ／Ｈとする、
ｆ．濃縮、
ｇ．乾燥してクロロゲン酸の粗生成物を得る。 A method for extracting chlorogenic acid from Tochu leaves, comprising the following steps a to g.
a. Weighing of Nakanaka,
b. Water extraction: Extraction is performed by adding water, extraction temperature is 40 ° C. to 80 ° C., extraction time is 0.5 h to 4.0 h, the number of extractions is 1 to 3 times, and the amount of water used per time is the mass of the drug substance. 6-14 times, pH value is 2-7, leaching or dynamic extraction is performed,
c. Pretreatment: Without concentrating the water extract, chitosan as a clarifier is added, the amount added is 0.2% to 1.0% of the mass of the drug, the settling temperature is 40 ° C to 80 ° C, and the settling time is 0.5h-24h,
d. Filtration: The pretreatment liquid in step c is filtered through a filtration membrane or filter core having a pore size of 50 μm to 200 μm.
e. Charge to the macroporous adsorption resin: The non-polar or weakly polar macroporous adsorption resin is passed through the filtrate in step d, the ratio of the diameter and height of the resin column is 1: 4 to 1:10, and the flow rate of the input is 2.0 to 8.0 BV / H, the amount of adsorption is such that the ratio of the mass of the chemical to the column volume is 1: 1 to 1: 5, and the amount of washing water used is 0.5 to 2.0 BV. The flow rate is 2.0 to 8.0 BV / H, the type of elution solvent is methanol or ethanol, the concentration is 10 to 30%, the amount of eluent used is 1.0 to 6.0 BV, and the flow rate of eluent is 2. .0 to 8.0 BV / H,
f. concentrated,
g. Dry to obtain a crude product of chlorogenic acid.
ステップｆで述べる濃縮条件は、濃縮温度４０℃〜８０℃、真空度−０．０４ＭＰ〜−０．０９ＭＰ、相対密度１．０２〜１．１０とし、ステップｇで述べる乾燥条件は、空気入口温度１５０℃〜２００℃、空気出口温度５０℃〜９０℃である噴霧乾燥をして、クロロゲン酸の粗生成物を得ることを特徴とする抽出方法。 The extraction method according to claim 1,
The concentration conditions described in step f are as follows: the concentration temperature is 40 ° C. to 80 ° C., the degree of vacuum is −0.04 MP to −0.09 MP, the relative density is 1.02 to 1.10, and the drying conditions described in step g are the air inlet temperature. An extraction method characterized by obtaining a crude product of chlorogenic acid by spray drying at 150 ° C to 200 ° C and an air outlet temperature of 50 ° C to 90 ° C.
以下のステップａ〜ｇを含むことを特徴とする抽出方法。
ａ．杜仲葉の秤取、
ｂ．水抽出：水を加えて抽出し、抽出温度は６０℃〜８０℃、抽出時間は０．５ｈ〜１．５ｈ、抽出回数は１〜３回、1回当たりの水の使用量は薬材の質量の１０〜１４倍、ｐＨ値は６〜７とし、浸出又は動的抽出を行う、
ｃ．前処理：清澄剤を加え、添加量は薬材の質量の０．４％〜０．８％、沈降温度は５０℃〜７０℃、沈降時間は１．０ｈ〜２．０ｈとする、
ｄ．ろ過：ステップｃの前処理液を、孔径５０μｍ〜１００μｍのろ過膜又はフィルターコアでろ過する、
ｅ．マクロポーラス吸着樹脂への投入：ステップｄのろ過液を、非極性又は弱極性のマクロポーラス吸着樹脂に通し、樹脂カラムの直径と高さの比は１：４〜１：８、投入流速は３．０〜５．０ＢＶ／Ｈ、吸着量は、薬材の質量とカラム体積の比が１：１〜１：４とし、洗浄水の使用量は１．０〜２．０ＢＶ、洗浄水の流速は３．０〜５．０ＢＶ／Ｈ、溶離溶媒の種類はメタノール又はエタノールとし、その濃度は１０〜３０％、溶離液の使用量は４．０〜６．０ＢＶ、溶離液の流速は２．０〜４．０ＢＶ／Ｈとする、
ｆ．濃縮：濃縮温度は５０℃〜７０℃、真空度は−０．０７ＭＰ〜−０．０９ＭＰ、相対密度は１．０３〜１．０５とする、
ｇ．乾燥：空気入口温度１５０℃〜１７０℃、空気出口温度６０℃〜８０℃である噴霧乾燥をして、クロロゲン酸の粗生成物を得る。 The extraction method according to claim 1,
The extraction method characterized by including the following steps ag.
a. Weighing of Nakanaka,
b. Water extraction: Extraction is performed by adding water, the extraction temperature is 60 ° C. to 80 ° C., the extraction time is 0.5 h to 1.5 h, the number of extractions is 1 to 3 times, the amount of water used per time is 10-14 times the mass, pH value is 6-7, and leaching or dynamic extraction is performed.
c. Pretreatment: A clarifier is added, the addition amount is 0.4% to 0.8% of the mass of the chemical, the sedimentation temperature is 50 ° C. to 70 ° C., and the sedimentation time is 1.0 h to 2.0 h.
d. Filtration: The pretreatment liquid in step c is filtered through a filtration membrane or filter core having a pore diameter of 50 μm to 100 μm.
e. Charging into the macroporous adsorption resin: The filtrate of step d is passed through the nonpolar or weakly polar macroporous adsorption resin, the ratio of the diameter and height of the resin column is 1: 4 to 1: 8, and the charging flow rate is 3. 0.0 to 5.0 BV / H, the amount of adsorption is such that the ratio of the mass of the chemical to the column volume is 1: 1 to 1: 4, the amount of wash water used is 1.0 to 2.0 BV, and the flow rate of the wash water Is 3.0 to 5.0 BV / H, the type of elution solvent is methanol or ethanol, the concentration is 10 to 30%, the amount of eluent used is 4.0 to 6.0 BV, and the flow rate of the eluent is 2. 0 to 4.0 BV / H,
f. Concentration: the concentration temperature is 50 ° C. to 70 ° C., the degree of vacuum is −0.07 MP to −0.09 MP, and the relative density is 1.03 to 1.05.
g. Drying: Spray drying at an air inlet temperature of 150 ° C to 170 ° C and an air outlet temperature of 60 ° C to 80 ° C is performed to obtain a crude product of chlorogenic acid.
ステップａで述べる杜仲葉にはクロロゲン酸が１．０％〜５．０％含まれることを特徴とする抽出方法。 The extraction method according to claim 1 or 2,
An extraction method characterized by containing 1.0% to 5.0% of chlorogenic acid in the licorice leaves described in step a.
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