JP4355797B2 - Method for producing caffeic acid - Google Patents
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- JP4355797B2 JP4355797B2 JP2009005071A JP2009005071A JP4355797B2 JP 4355797 B2 JP4355797 B2 JP 4355797B2 JP 2009005071 A JP2009005071 A JP 2009005071A JP 2009005071 A JP2009005071 A JP 2009005071A JP 4355797 B2 JP4355797 B2 JP 4355797B2
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Description
本発明は、カフェ酸の誘導体を主な成分とするポリフェノールからカフェ酸を製造する方法に関するものである。特に、ゴボウ葉未利用資源の有効利用に関するものである。 The present invention relates to a process for producing caffeic acid from a polyphenol containing a caffeic acid derivative as a main component. In particular, it relates to effective use of unused burdock leaf resources.
カフェ酸は機能性のカフェ酸エステル類などを合成するときの原料として利用されているが、製造が困難で多額の費用がかかり、少量しか供給されていなかった。そこで、カフェ酸の製造方法として、クロロゲン酸やその誘導体を加水分解する方法が提案されているが、カフェ酸をジエチルエーテルを用いて抽出することから、引火性の溶媒であるジエチルエーテルを多量に用いるため危険性が大きいという問題があった（例えば特許文献１）。また、甘しょ焼酎蒸留粕は処理が困難である一方で、機能性成分を含む可能性があるものの、その有効利用が十分なされていなかった。 Caffeic acid has been used as a raw material when synthesizing functional caffeic acid esters and the like, but it was difficult to produce, costing a large amount, and only a small amount was supplied. Therefore, as a method for producing caffeic acid, a method for hydrolyzing chlorogenic acid and its derivatives has been proposed. Since caffeic acid is extracted using diethyl ether, a large amount of flammable solvent diethyl ether is used. There is a problem that the danger is large because it is used (for example, Patent Document 1). Moreover, while sweet potato shochu distillery is difficult to process, it may contain functional components, but its effective use has not been sufficient.
本発明の目的は、カフェ酸を簡便かつ安全に、しかも効率良く製造する方法を提供しようとするものである。特に、ゴボウ葉未利用資源の有効に活用しようとするものである。 An object of the present invention is to provide a method for producing caffeic acid simply, safely and efficiently. In particular, it intends to make effective use of unused burdock leaf resources.
（１）ゴボウ葉から得られた抽出液に含まれるカフェ酸の誘導体を主な成分とするポリフェノールに、麹菌又はその抽出液を作用させることを特徴とするカフェ酸を製造することを特徴とする。これはゴボウ葉未利用資源を有効に活用する目的のためである。 (1) It is characterized by producing caffeic acid characterized by allowing a koji mold or its extract to act on polyphenols mainly comprising a caffeic acid derivative contained in an extract obtained from burdock leaves. . This is for the purpose of effectively utilizing unused resources of burdock leaves.
（２）本発明の第２は、カフェ酸の誘導体が、モノ又はジカフェオイルキナ酸であることを特徴とする前項（１）記載のカフェ酸の製造方法である。これらカフェ酸の誘導体としては、クロロゲン酸や、４，５ジカフェオイルキナ酸、３，５−ジカフェオイルキナ酸、３，４−ジカフェオイルキナ酸等が含まれる。 (2) A second aspect of the present invention is the method for producing caffeic acid as described in (1) above, wherein the caffeic acid derivative is mono- or dicaffeoylquinic acid. Examples of these caffeic acid derivatives include chlorogenic acid, 4,5-dicaffeoylquinic acid, 3,5-dicaffeoylquinic acid, 3,4-dicaffeoylquinic acid, and the like.
（３）本発明の第３は、使用する麹菌が白麹菌又は黒麹菌であることを特徴とする前項（１）又は（２）記載のカフェ酸の製造方法である。これら白麹菌，黒麹菌はクエン酸生成能力が高く雑菌汚染にも強いことから特にこれらを選択した。 (3) A third aspect of the present invention is the method for producing caffeic acid as described in (1) or (2) above, wherein the koji mold used is white koji mold or black koji mold. These white koji molds and black koji molds were selected because they have a high capacity to produce citric acid and are resistant to contamination.
（４）本発明の第４は、ゴボウ葉から得られた抽出液に含まれるカフェ酸の誘導体を主な成分とするポリフェノールに、麹菌又はその抽出液を作用させた後、合成吸着剤による吸着処理をして吸着画分を得、該吸着画分をアルコール水溶液を用いて溶出することを特徴とする請求項１〜３のいずれか１項記載のカフェ酸の製造方法である。この方法により高純度のカフェ酸を得ることができる。 (4) According to the fourth aspect of the present invention, after the koji mold or its extract is allowed to act on polyphenols mainly composed of caffeic acid derivatives contained in the extract obtained from burdock leaves, adsorption by a synthetic adsorbent The method for producing caffeic acid according to any one of claims 1 to 3, wherein an adsorption fraction is obtained by treatment, and the adsorption fraction is eluted using an aqueous alcohol solution. High purity caffeic acid can be obtained by this method.
（５）本発明の第５は、アルコール水溶液が、メチルアルコール、エチルアルコール、ｎ−プロピルアルコール、ｉｓｏ−プロピルアルコール、ｎ−ブチルアルコール、ｉｓｏ−ブチルアルコール、第２ブチルアルコール、第３ブチルアルコールからなる群のうちから選ばれた１又は２以上からなる水溶液であることを特徴とする前項（４）記載のカフェ酸の製造方法である。これは脂肪族低級アルコールを指定したものであり、水に対する溶解度の低いものは他のアルコールに混合して使用するものとする。 (5) In the fifth aspect of the present invention, the aqueous alcohol solution is selected from methyl alcohol, ethyl alcohol, n-propyl alcohol, iso-propyl alcohol, n-butyl alcohol, iso-butyl alcohol, secondary butyl alcohol, and tertiary butyl alcohol. The method for producing caffeic acid as described in (4) above, which is an aqueous solution comprising one or more selected from the group consisting of: This designates an aliphatic lower alcohol, and those having low solubility in water are used by mixing with other alcohols.
（６）本発明の第６は、アルコール水溶液のアルコール濃度が３０〜５０質量％であることを特徴とする前項（４）又は（５）記載のカフェ酸の製造方法である。 カフェ酸はアルコール濃度４０質量％で最も良く溶出され、アルコール濃度６０質量％以上又は３０質量％以下ではほとんど溶出しないからである。 (6) A sixth aspect of the present invention is the method for producing caffeic acid according to the above (4) or (5), wherein the alcohol concentration of the alcohol aqueous solution is 30 to 50% by mass. This is because caffeic acid is best eluted at an alcohol concentration of 40% by mass and hardly eluted at an alcohol concentration of 60% by mass or less or 30% by mass or less.
以上のように本発明では、請求項１に係る発明により、ゴボウ葉から得られた抽出液に含まれるカフェ酸の誘導体を主な成分とするポリフェノールに、麹菌又はその抽出物を作用させることで、ポリフェノールを分解し、容易にカフェ酸を生成させることができた。ゴボウの未利用資源を活用することでカフェ酸を得られることが示された。 As described above, in the present invention, by the invention according to claim 1, the koji mold or an extract thereof is allowed to act on polyphenols mainly composed of caffeic acid derivatives contained in the extract obtained from burdock leaves. The polyphenol was decomposed and caffeic acid was easily generated. It was shown that caffeic acid can be obtained by utilizing unused burdock resources.
請求項２に係る発明では、カフェ酸の誘導体がモノ又はジカフェオイルキナ酸である場合、容易にカフェ酸が生成させることがわかった。 In the invention which concerns on Claim 2, when the derivative | guide_body of caffeic acid was mono or dicaffeoylquinic acid, it turned out that a caffeic acid produces | generates easily.
請求項３に係る発明では、麹菌が焼酎製造用に広く用いられている白麹菌又は黒麹菌が有効であることを示された。 In the invention which concerns on Claim 3, it was shown that the koji mold or the black koji mold which is widely used for shochu production is effective.
請求項４に係る発明では、生成したカフェ酸を合成吸着剤で処理することにより、高純度のカフェ酸が得られることが示された。 In the invention which concerns on Claim 4, it was shown that a highly purified caffeic acid is obtained by processing the produced caffeic acid with a synthetic adsorbent.
請求項５に係る発明では、高純度のカフェ酸を得るために用いる溶媒が脂肪族低級アルコールを用いることとし、比較的安全性の高い条件で操作が行われることが示された。 In the invention according to claim 5, it was shown that the solvent used for obtaining high-purity caffeic acid was an aliphatic lower alcohol, and the operation was performed under relatively high safety conditions.
請求項６に係る発明では、使用するアルコール濃度が３０〜５０質量％の範囲であることから操作上、特に引火の危険性は低いことが示された。 In the invention which concerns on Claim 6, since the alcohol density | concentration to be used is the range of 30-50 mass%, it was shown that the danger of ignition was low on operation.
以下に本発明の実施の形態を参考例に基づいて説明する。
甘しょ焼酎蒸留粕液（酵素処理した液）と、サツマイモもろみ酢（甘しょ焼酎蒸留粕液にセルラーゼ系酵素と２０％の米麹を添加して処理し得られた液）についてポリフェノールの高速液体クロマトグラフィによる分析を行った。
分析方法は以下のとおりである。試料はセルロース膜（０．２０μｍアドバンテック）でろ過後、高速液体クロマトグラフィにより分析した。分析条件を下記に示す。
Embodiments of the present invention will be described below based on reference examples.
About sweet potato shochu distilled liquor (enzyme-treated liquid) and sweet potato moromi vinegar (liquid obtained by adding cellulase enzymes and 20% rice bran to sweet potato shochu distilled liquor) Chromatographic analysis was performed.
The analysis method is as follows. The sample was analyzed by high performance liquid chromatography after filtration through a cellulose membrane (0.20 μm Advantech). The analysis conditions are shown below.
ポンプ：ＬＣ−１０ＡＴ
オートインジェクター：ＳＩＬ−１０ＡＸＬ
検出器：ＳＰＤ−Ｍ１０ＡＶＰ フォトダイオードアッレイ
カラム：Ａ ＹＭＣ−Ｐａｃｋ ＯＤＳ−ＡＭ−３０２ ｃｏｌｕｍｎ （４．６ｍｍ ＩＤ×１５０ｍｍ，５μｍ）
温度：４０℃
溶媒：０．２％（Ｖ／Ｖ）蟻酸（Ａ），メチルアルコール（Ｂ），２％Ｂ（０−１５分），２％−４５％Ｂグラジエント（１５−５０分），４５％Ｂ（５０−６５分）
流速：１ｍｌ／分
Pump: LC-10AT
Auto injector: SIL-10AXL
Detector: SPD-M10AVP Photodiode array column: A YMC-Pack ODS-AM-302 column (4.6 mm ID × 150 mm, 5 μm)
Temperature: 40 ° C
Solvent: 0.2% (V / V) formic acid (A), methyl alcohol (B), 2% B (0-15 minutes), 2% -45% B gradient (15-50 minutes), 45% B ( 50-65 minutes)
Flow rate: 1 ml / min
高速液体クロマトグラフィによる分析結果は図１に示すとおりであって、図中、焼酎粕液とあるのは、甘しょ焼酎蒸留粕を酵素処理した液であり、もろみ酢とあるのは、甘しょ焼酎蒸留粕を酵素と２０％麹を添加して処理したものである。甘しょ焼酎蒸留粕液には少なくとも４種類のポリフェノール化合物が確認されたが、サツマイモもろみ酢ではほぼ単一のピークが確認され、このピークはカフェ酸とコクロマトグラフィにより同定された。この結果は、甘しょ焼酎蒸留粕を麹で処理することにより、他のポリフェノールがカフェ酸に変化したものと考えられる。またサツマイモもろみ酢のカフェ酸のピークの高さは、甘しょ焼酎蒸留粕液のカフェ酸のピークの高さの約２倍程度となっている。 The analysis results by high performance liquid chromatography are as shown in FIG. 1. In the figure, shochu liquor is a solution obtained by enzymatic treatment of sweet potato shochu distilled spirit, and moromi vinegar is sweet potato shochu. Distilled lees were processed by adding enzymes and 20% lees. At least four types of polyphenol compounds were confirmed in the sweet potato shochu distilled liquor, but almost a single peak was confirmed in sweet potato moromi vinegar, and this peak was identified by caffeic acid and co-chromatography. This result is considered that the other polyphenols were changed to caffeic acid by treating the sweet potato shochu distiller with boil. The height of the caffeic acid peak in sweet potato moromi vinegar is about twice the height of the caffeic acid peak in the sweet potato shochu distilled liquor.
サツマイモもろみ酢の吸着クロマトグラフィ画分のポリフェノール含量：サツマイモもろみ酢を合成吸着剤（ＭＣＩ ｇｅｌ ＣＨＰ ２０Ｐ樹脂、三菱化学製、カラムサイズ２．５×５０ｃｍ）に吸着後、吸着画分をメチルアルコールの濃度の異なる水溶液で溶出した。各画分のポリフェノール含量は、メチルアルコールの濃度が８０％画分＞４０％画分＞６０％画分＞１００％画分＞２０％画分＞非吸着画分の順であった。図２に以上の吸着クロマトグラフィの結果を示した。 Polyphenol content in the adsorption chromatographic fraction of sweet potato moromi vinegar: After adsorbing sweet potato moromi vinegar to a synthetic adsorbent (MCI gel CHP 20P resin, manufactured by Mitsubishi Chemical, column size 2.5 × 50 cm), the adsorbed fraction was concentrated to methyl alcohol. Eluted with different aqueous solutions. The polyphenol content of each fraction was such that the concentration of methyl alcohol was 80% fraction> 40% fraction> 60% fraction> 100% fraction> 20% fraction> non-adsorbed fraction. FIG. 2 shows the results of the above adsorption chromatography.
図２において、４０％画分は高速液体クロマトグラフィによる検討の結果、カフェ酸であることが確認された。６０％画分及び８０％画分は未同定の成分である。 In FIG. 2, the 40% fraction was confirmed to be caffeic acid as a result of high-performance liquid chromatography. The 60% and 80% fractions are unidentified components.
〔参考例１〕
原料の異なるもろみ酢におけるポリフェノール類の組成を高速液体クロマトグラフィで測定した結果は図３に示すとおりである。
特にサツマイモもろみ酢（図３の中で米麹・イモ蒸留粕・米麹と記載されたもの）のカフェ酸含量が多いことが特徴的である。
[Reference Example 1]
The result of having measured the composition of the polyphenols in the moromi vinegar from which raw materials differ by high performance liquid chromatography is as showing in FIG.
In particular, the sweet potato moromi vinegar (shown in FIG. 3 as rice bran, potato distilled bran, rice bran) has a high caffeic acid content.
〔参考例２〕
サツマイモの葉は高濃度のポリフェノール類を含有している。これらのポリフェノール類のほとんどはモノ及びジカフェオイルキナ酸であり、カフェ酸は極微量しか含まれていない。
そこで、麹抽出液とサツマイモ葉ポリフェノール画分を調製し、これらを反応させることとした。
麹抽出液は、麹４ｇに酢酸緩衝液（８６ｍＭ ＮａＣｌ＋０．１Ｍ酢酸緩衝液（ｐＨ５．０））２０ｍｌを加え、２０℃で３時間シェーカーで撹拌・抽出し、上澄を０．４５μｍフィルターでろ過して調製した。
[Reference Example 2]
Sweet potato leaves contain high concentrations of polyphenols. Most of these polyphenols are mono- and dicaffeoylquinic acid, and caffeic acid is contained in trace amounts.
Therefore, it was decided to prepare a koji extract and a sweet potato leaf polyphenol fraction and react them.
For the cocoon extract, add 20 ml of acetate buffer (86 mM NaCl + 0.1 M acetate buffer (pH 5.0)) to 4 g of cocoon, stir and extract with a shaker at 20 ° C. for 3 hours, and filter the supernatant with a 0.45 μm filter. Prepared.
サツマイモ葉ポリフェノール画分は、試料１００ｇを４リットルのメチルアルコールで抽出し、エバポレーターで濃縮乾固後、蒸留水５００ｍｌに溶解し、同量のヘキサンを加え、分液ロートで分層し、水層をポリフェノール画分として回収した。 For the sweet potato leaf polyphenol fraction, extract 100 g of sample with 4 liters of methyl alcohol, concentrate to dryness with an evaporator, dissolve in 500 ml of distilled water, add the same amount of hexane, separate the layers with a separatory funnel, Was recovered as a polyphenol fraction.
ゴボウ葉、ヨモギについて同様にポリフェノール画分を調製した。
麹抽出液とポリフェノール画分を６：４で混合後、５０℃で１６時間反応させ、ポリフェノールの分解状況を高速液体クロマドグラフィで確めた。
Polyphenol fractions were similarly prepared for burdock leaves and mugwort.
After the soot extract and polyphenol fraction were mixed at 6: 4, the mixture was reacted at 50 ° C. for 16 hours, and the degradation state of polyphenol was confirmed by high-speed liquid chromatography.
表１によると、サツマイモ葉のモノ及びジカフェオイルキナ酸のピークは麹抽出液処理により減少し、カフェ酸に相当するピークが著しく大きくなっている。
ゴボウ葉及びヨモギのポリフェノール類も麹抽出液処理により分解され、カフェ酸が増加していることがわかった。これらの結果は、麹菌（本実施例では白麹菌）を使えば、カフェ酸誘導体を含有する作物又はこれらを加工した際に発生する残渣等から容易にカフェ酸が製造可能なことを示している。
According to Table 1, the peaks of mono- and dicaffeoylquinic acid in sweet potato leaves are reduced by the koji extract treatment, and the peak corresponding to caffeic acid is remarkably large.
It was found that burdock leaves and mugwort polyphenols were also decomposed by the koji extract treatment and caffeic acid increased. These results show that if koji molds (white koji molds in this example) are used, caffeic acid can be easily produced from crops containing caffeic acid derivatives or residues generated when these are processed. .
Claims (6)
The method for producing caffeic acid according to claim 4 or 5, wherein the alcohol concentration of the alcohol aqueous solution is 30 to 50% by mass.
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