JP2509989B2 - Protein curculin and its use - Google Patents
Protein curculin and its useInfo
- Publication number
- JP2509989B2 JP2509989B2 JP63277717A JP27771788A JP2509989B2 JP 2509989 B2 JP2509989 B2 JP 2509989B2 JP 63277717 A JP63277717 A JP 63277717A JP 27771788 A JP27771788 A JP 27771788A JP 2509989 B2 JP2509989 B2 JP 2509989B2
- Authority
- JP
- Japan
- Prior art keywords
- curculin
- aqueous solution
- fruit
- protein
- taste
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Landscapes
- Peptides Or Proteins (AREA)
- General Preparation And Processing Of Foods (AREA)
- Seeds, Soups, And Other Foods (AREA)
- Non-Alcoholic Beverages (AREA)
- Medicinal Preparation (AREA)
Description
【発明の詳細な説明】 〔産業上の利用分野〕 本発明は、新規な蛋白質クルクリン及びその味覚修飾
剤としての用途に関するものである。DETAILED DESCRIPTION OF THE INVENTION [Industrial field of use] The present invention relates to a novel protein curculin and its use as a taste-modifying agent.
舌の受容膜に作用して、食品の味覚を変える物質(味
覚修飾物質)としては、従来、口中に含んだ後、甘味物
質を食した時、または甘味物質とともに食した時、甘味
を感じさせなくするものとしてギムネマ シルベスタ
(Gymnema sylvestre)の葉に含まれるギムネマ酸、及
びなつめ(Ziziphusjujuba)の葉に含まれるジジフィン
が知られており、また上記と同様にして酸味物質を食し
た時、甘味を感じさせるものとして、ミラクルフルーツ
(Synsepulm dulcificum)の実に含まれるミラクリンが
知られている。As a substance that acts on the receptor membrane of the tongue to change the taste of food (taste modifier), it has been conventionally made to feel sweetness when it is eaten with a sweet substance or when it is eaten with a sweet substance. Gimnema acid contained in the leaves of Gymnema sylvestre and didifin contained in the leaves of Ziziphusjujuba are known to be eliminated, and when the sour substance is eaten in the same manner as above, the sweetness is reduced. It is known that miraculin, which is contained in the fruit of miracle fruit (Synsepulm dulcificum), is a sensation.
また、クルクリゴ・ラチフォリア(Curculigo latifo
lia)は、西マレーシアやタイ南部等に自生するきんば
いざさ科の植物であり、その果実は食用に適し、食欲増
進効果があることは知られている。Also, the Curculigo latifo
lia) is a plant belonging to the family Anthrush that grows naturally in West Malaysia and southern Thailand, and its fruits are known to be edible and have an appetite enhancing effect.
上記のミラクリンは、上述の如き機能を有するもので
あるが、安定性上の問題があり、味覚修飾物質として実
用化されていない。The above-mentioned miraculin has the above-mentioned functions, but has a problem in stability and has not been put to practical use as a taste-modifying substance.
また、クリクリゴ・ラチフォリアの果実は上述以外の
性質については知られていない。Also, the fruit of Cliclygo latifolia is not known for properties other than the above.
従って、本発明の目的は、安定化度の高い新規な味覚
修飾物質を提供することにある。Therefore, an object of the present invention is to provide a novel taste modifier having a high degree of stabilization.
本発明者らは、クルクリゴ・ラチフォリアの果実を食
した後、酸味物質または水を食すると、甘味が感じられ
ることを見いだし、その誘因物質が何であるか鋭意研究
した結果、クルクリゴ・ラチフォリアの実(Fruit)に
含まれる特定の蛋白質(クルクリンと命名)が甘味を感
じさせる誘因物質であり、前記目的を達成するものであ
ることを知見した。The present inventors found that sweet taste was felt when eating sour substances or water after eating the fruit of Curculigo latifolia, and as a result of diligent research on what the inducer was, the fruit of Curculigo latifolia ( It was found that a specific protein (named curculin) contained in (Fruit) is a sweetness-inducing substance and achieves the above-mentioned object.
本発明は、上記知見に基づきなされたもので、クルク
リゴ・ラチフォリアの果実またはその乾燥物から0.01M
以上の濃度の塩の水溶液で抽出することによって得ら
れ、甘味を感じさせる味覚修飾作用を有し、等電点7.
1、分子量約12500ダルトンであるモノマーのダイマーで
ある蛋白質クルクリン、クルクリンを含有する食品、及
び蛋白質クリクリンの抽出方法を提供するものである。The present invention was made on the basis of the above findings, and was prepared from a fruit of Curculigo latifolia or a dried product thereof of 0.01M.
Obtained by extraction with an aqueous solution of salt having the above concentration, it has a taste-modifying action that makes sweet taste, and has an isoelectric point of 7.
1. To provide a protein curculin, which is a dimer of a monomer having a molecular weight of about 12500 daltons, a food containing curculin, and a method for extracting protein criclin.
以下、本発明の蛋白質クルクリンについて詳述する。 Hereinafter, the protein curculin of the present invention will be described in detail.
本発明のクルクリンは、クルクリゴ・ラチフォリア
(Curculigo latifolia)の果実またはその乾燥物から
0.01M以上の濃度の塩の水溶液で抽出することによって
得られる。The curculin of the present invention is obtained from the fruit of Curculigo latifolia or its dried product.
It is obtained by extraction with an aqueous solution of salt having a concentration of 0.01 M or more.
上記の塩としては、ナトリウム、カリウム、カルシウ
ム、マグネシウム若しくはアンモニウムの塩酸塩、ナト
リウム、カリウム、マグネシウム若しくはアンモニウム
のリン酸塩、ナトリウム、カリウム、マグネシウム若し
くはアンモニウムの炭酸塩、ナトリウム、マグネシウ
ム、カルシウム若しくはアンモニウムの硫酸塩又は亜硫
酸塩、ナトリウム若しくはカリウムの硝酸塩又は亜硝酸
塩、ナトリウム若しくはカルシウムの乳酸塩、ミョウバ
ン、焼ミョウバン、酢酸ナトリウム、ナトリウム若しく
はカリウムのピロリン酸塩、ナトリウム若しくはカルシ
ウムのプロピオン酸塩、安息香酸ナトリウム、フマル酸
−ナトリウム、ポリアクリル酸ナトリウム等が用いられ
る。Examples of the salt include sodium, potassium, calcium, magnesium or ammonium hydrochloride, sodium, potassium, magnesium or ammonium phosphate, sodium, potassium, magnesium or ammonium carbonate, sodium, magnesium, calcium or ammonium. Sulfate or sulfite, sodium or potassium nitrate or nitrite, sodium or calcium lactate, alum, baked alum, sodium acetate, sodium or potassium pyrophosphate, sodium or calcium propionate, sodium benzoate, Fumarate-sodium, sodium polyacrylate and the like are used.
また、上記のクルクリゴ・ラチフォリアの果実の乾燥
物としては、その乾燥手段に特に制限されるものではな
く、天日乾燥物、熱風乾燥物、及び凍結乾燥パルプ等の
凍結乾燥物等が用いられる。The dried product of Curculigo latifolia fruit is not particularly limited in its drying means, and sun dried product, hot air dried product, freeze-dried product such as freeze-dried pulp and the like can be used.
そして、上記塩の水溶液によるクルクリンの抽出方法
の代表的な一例を挙げると次の通りである。Then, a typical example of the method for extracting curculin with the above aqueous solution of salt is as follows.
クルクリゴ・ラチフォリアの果実またはその乾燥物に
上記の塩の水溶液を加えてホモジナイズした後、濾過、
遠心分離等を行って抽出できるが、クルクリンはクルク
リゴ・ラチフォリアの果実の果肉中の水不溶部分に含ま
れるため、上記果実またはその乾燥物に水を加えてホモ
ジナイズし、充分水洗いして水可溶部を除去した後、そ
の残査から上記の塩の水溶液で抽出する方がクルクリン
を高純度で得られるので好ましい。After adding the above salt aqueous solution to the fruit of Curculigo latifolia or its dried product and homogenizing, filtration,
It can be extracted by centrifugation, etc., but since curculin is contained in the water-insoluble part of the fruit pulp of Curculigo latifolia, it is homogenized by adding water to the fruit or a dried product thereof, and washed thoroughly with water to dissolve in water. After removing a part, it is preferable to extract the residue with an aqueous solution of the above salt, because curculin can be obtained in high purity.
抽出に用いられる塩の水溶液が0.01M未満の濃度のも
のではクルクリンを充分に抽出することができないの
で、0.01M以上の濃度の塩の水溶液が必要である。一
方、あまり高濃度では、抽出後の脱塩に手間がかかるの
で、抽出効率と抽出以後の精製の手間のかね合いから0.
1〜1.0Mの濃度の塩の水溶液が好ましい。If the aqueous salt solution used for extraction has a concentration of less than 0.01 M, curculin cannot be sufficiently extracted, so an aqueous salt solution of 0.01 M or more is required. On the other hand, if the concentration is too high, desalting after extraction takes time, so it is not possible to balance the efficiency of extraction and the purification after extraction.
An aqueous solution of salt with a concentration of 1 to 1.0 M is preferred.
上記塩の水溶液による抽出液を脱塩、乾燥することに
より、充分実用に供するクルクリン含有物質が得られる
が、抽出液をさらにCM−セファローズによるイオン交
換、ゲルカラム使用中のHPLCにかけて精製することによ
り、純度を上げることができ、その後、脱塩、乾燥して
純クルクリンが得られる。もちろん、上記の精製法の
他、抽出液を塩析、溶剤沈殿その他の公知の蛋白精製法
により分画することによっても、クルクリン純度を上げ
ることができる。By desalting and drying the extract with an aqueous solution of the above salt, a curculin-containing substance that can be sufficiently put to practical use can be obtained. , The purity can be increased, and then desalted and dried to obtain pure curculin. Of course, in addition to the above-mentioned purification method, the curculin purity can be increased by fractionating the extract solution by a known protein purification method such as salting out, solvent precipitation or the like.
このようにして得られる本発明のクルクリンの具体例
としては、分子量約12500ダルトン(dalton)、等電点
7.1の蛋白質が挙げられ、この蛋白質は分子量約26000ダ
ルトン(dalton)のダイマー(dimer)として存在す
る。また、上記蛋白質のアミノ酸組成は下記の第1表の
通りであり、上記蛋白質は比較的多量のアスパラギン
酸、ロイシン及びグリシンを含むものである。Specific examples of the curculin of the present invention thus obtained include a molecular weight of about 12500 daltons and an isoelectric point.
The protein of 7.1 is mentioned, and this protein exists as a dimer with a molecular weight of about 26,000 daltons. The amino acid composition of the protein is shown in Table 1 below, and the protein contains a relatively large amount of aspartic acid, leucine and glycine.
上記の本発明の蛋白質クルクリンは、甘味修飾剤とし
て好適に用いられる。 The above-mentioned protein curculin of the present invention is suitably used as a sweetness modifier.
上記クルクリンからなる本発明の味覚修飾剤は、その
まま摂取してもよいが、食品、飲料または薬剤等に適宜
配合して用いることができる。食品等に対する配合量
は、後述するクルクリンの甘味発現活性を参考にして、
目的及び用途に応じ定めることができる。The taste-modifying agent of the present invention comprising curculin may be ingested as it is, but may be used by appropriately blending it with a food, a beverage, a drug or the like. The amount to be mixed with food, etc., with reference to the sweetness expressing activity of curculin described later,
It can be set according to the purpose and use.
また、クルクリンからなる味覚修飾剤を含有する食品
等は、その食品等の性状に応じて、粉末状、溶液状、シ
ート状、スプレー状または乳化物状等に加工することが
できる。Further, a food or the like containing a taste modifier comprising curculin can be processed into a powder, a solution, a sheet, a spray, an emulsion or the like depending on the properties of the food or the like.
本発明のクルクリンは、口中に含んだ後、酸味物質、
無味物質または水等を食した時、または酸味物質、無味
物質または水等とともに食した時、舌の受容膜に作用し
て甘味を感じさせる効果を有しており、本発明のクルク
リンあるいは該クルクリンを含む製剤または食品等を事
前に食しておけば、30分後までは、酸味物質、無味物質
または水を食した時に甘味を感じさせることができる。Curculin of the present invention, after containing in the mouth, a sour substance,
The curculin of the present invention or the curculin of the present invention has the effect of acting on the receptive membrane of the tongue to make it taste sweet when eaten with a tasteless substance or water, or when eaten with a sour substance, tasteless substance, water, etc. By pre-eating a preparation or food containing the above, sweetness can be felt when eating a sour substance, a non-taste substance or water until 30 minutes later.
各種水溶液に対する純クルクリン(クルクリン4×10
-5M濃度)の甘味発現活性は下記第2表に示す通りであ
り、酸味物質に対し、特に活性が高い。Pure curculin for various aqueous solutions (curcurin 4 × 10
The sweetness expressing activity at a concentration of −5 M) is as shown in Table 2 below, and the activity is particularly high for sour substances.
また、本発明のクルクリンは、ミラクルフルーツの実
に含まれるミラクリンと異なり、水溶液中でも安定であ
るので、クルクリン及びクルクリンを含む製剤または食
品等を粉末状、錠剤状、シート状の他、溶液状、乳化物
状、スプレー状とすることが容易である。 Further, curculin of the present invention, unlike miraculin contained in miracle fruit, is stable in an aqueous solution, so that curculin and a formulation containing curcurin or a food or the like is powdered, tableted, sheet-shaped, solution-shaped or emulsified. It is easy to make a product or a spray.
また、本発明のクルクリンは、それ自体弱い甘味を有
する。Moreover, the curculin of the present invention has a weak sweetness in itself.
次に実施例を示し、本発明を具体的に説明する。 Next, the present invention will be specifically described with reference to examples.
実施例1 クルクリゴ・ラチフォリア(Curculigo latifolia)
の果実の凍結乾燥パルプ30gに600mlの水を加え、ミキサ
ーで2分間ホモジナイズした後、10000r.p.m.で30分間
遠心分離した。上澄(着色している)を除去後、残査に
600mlの水を加え、ホモジナイズ、遠心分離、上澄除去
を、上澄が着色しなくなるまで4回くりかえし、残査を
得た。Example 1 Curculigo latifolia
600 ml of water was added to 30 g of freeze-dried pulp of the fruit of Example 1, homogenized with a mixer for 2 minutes, and then centrifuged at 10000 rpm for 30 minutes. After removing the supernatant (colored), leave the residue
600 ml of water was added, homogenization, centrifugation, and removal of the supernatant were repeated 4 times until the supernatant did not become colored, and a residue was obtained.
次に、この残査に0.5MNaCl水溶液250mlを加え、ミキ
サーで2分間ホモジナイズした後、吸引濾過した。濾液
分取後、残査にさらに0.5MNaCl水溶液250mlを加え、ホ
モジナイズ及び吸引濾過を行い、濾液を分取した。Next, 250 ml of 0.5 M NaCl aqueous solution was added to this residue, and the mixture was homogenized with a mixer for 2 minutes and then suction filtered. After collecting the filtrate, 250 ml of 0.5 M NaCl aqueous solution was further added to the residue, homogenization and suction filtration were performed, and the filtrate was collected.
次に、濾液を合わせた後、30000r.p.m.で1時間遠心
分離し、上澄(クルクリン粗抽出液)を得た。Next, the filtrates were combined and then centrifuged at 30,000 rpm for 1 hour to obtain a supernatant (cruculin crude extract).
この粗抽出液から脱塩、凍結乾燥して、クルクリン含
有物質(粗クルクリン)を得た。The crude extract was desalted and freeze-dried to obtain a curculin-containing substance (crude curculin).
実施例2 実施例1で得られたクルクリン粗抽出液500mlが限外
濾過により、30mlまで濃縮した後、この濃縮液に0.01M
リン酸バッファー(pH6.8)70mlを加え、100mlとしたも
のを試料液として、CM−セファローズカラムクロマトグ
ラフィーを行った〔CM−セファローズCL−6B。0.01Mリ
ン酸バッファー(pH6.8)で平衡化したもの。〕。ベッ
ドボリューム130ml及びベッド高さ17cmのカラムに試料
液を流下し、0.01Mリン酸バッファー(pH6.8)で洗浄し
た後、0−1.0MNaCl/0.01Mリン酸バッファー(pH6.8)
のグラディエントで溶出し、活性画分を集めた。Example 2 500 ml of the crude curculin extract obtained in Example 1 was concentrated to 30 ml by ultrafiltration, and then 0.01 M was added to the concentrate.
70 ml of a phosphate buffer (pH 6.8) was added to make 100 ml, and CM-Sepharose column chromatography was performed using the sample solution as a sample solution [CM-Sepharose CL-6B. Equilibrated with 0.01M phosphate buffer (pH 6.8). ]. The sample solution was flowed down to a column with a bed volume of 130 ml and a bed height of 17 cm, washed with 0.01 M phosphate buffer (pH 6.8), and then 0-1.0 M NaCl / 0.01 M phosphate buffer (pH 6.8).
Was eluted with a gradient of and the active fractions were collected.
この活性画分から、脱塩、凍結乾燥して、クルクリン
含有物質(粗精製クルクリン)を得た。The active fraction was desalted and lyophilized to obtain a curculin-containing substance (crude curculin).
実施例3 実施例2で得られた活性画分を限外濾過で濃縮した
後、ゲルカラム(東洋ソーダ製TSKゲルG3000SW)を用い
たHPLCにかけ、0.01Mリン酸バッファー(pH6.8)で溶出
した。Example 3 The active fraction obtained in Example 2 was concentrated by ultrafiltration and then subjected to HPLC using a gel column (TSK gel G3000SW manufactured by Toyo Soda) and eluted with 0.01M phosphate buffer (pH 6.8). .
クルクリンは、強い活性を持つシャープな1ピークと
して溶出された。Curculin was eluted as one sharp peak with strong activity.
この活性画分を脱塩、凍結乾燥して、精製クルクリン
を得た。This active fraction was desalted and lyophilized to obtain purified curculin.
なお、この精製クルクリンを8.0M尿素の存在下でSDS
−PAGEにかけたところ、分子量12500ダルトン(dalto
n)の所に単一バンドを示し、クルクリン純品であるこ
とが確認された。The purified curculin was treated with SDS in the presence of 8.0M urea.
When subjected to −PAGE, the molecular weight was 12500 daltons (dalto
A single band was shown at n), and it was confirmed to be pure curculin.
クルクリゴ・ラチフォリアの果実の凍結乾燥パルプ30
gあたりの、各実施例で得られたクルクリンの蛋白量、
活性収率及び比活性は下記第3表に示す通りであった。Curculigo latifolia fruit lyophilized pulp 30
Protein amount of curculin obtained in each example per g,
The activity yield and specific activity were as shown in Table 3 below.
実施例4 実施例1で得られた粗クルクリン10mgと食塩1mgを水2
0mlに溶解し、0.05%水溶液とした。この水溶液は室温
(25℃前後)に1ケ月放置した後も安定であった。この
水溶液1mlを口中に1分間含み吐き出した後、0.02Mクエ
ン酸、水、紅茶(ブラック)をそれぞれ飲食した。その
時の甘味の感じ方は下記第4表に示す通りであった。 Example 4 10 mg of crude curculin obtained in Example 1 and 1 mg of sodium chloride were added to water 2
It was dissolved in 0 ml to give a 0.05% aqueous solution. This aqueous solution was stable even after being left at room temperature (around 25 ° C) for 1 month. After 1 ml of this aqueous solution was put into the mouth for 1 minute and spit out, 0.02 M citric acid, water, and black tea (black) were respectively consumed. The way of feeling the sweetness at that time was as shown in Table 4 below.
紅茶(ブラック)の場合は普通の砂糖添加の紅茶と同
様の味を感じた。 In the case of black tea (black), I felt the same taste as ordinary sugar-added black tea.
実施例5 下記第5表に示す配合にて、ブラックのコーヒーゼリ
ー(A)及び砂糖入りのコーヒーゼリー(B)を作成し
た。Example 5 Black coffee jelly (A) and sugar-containing coffee jelly (B) were prepared according to the formulation shown in Table 5 below.
一方、市販の生クリーム(C)に実施例1で得られた
粗クルクリン0.005%を加え、クルクリン含有生クリー
ム(D)を作成した。 On the other hand, 0.005% of the crude curculin obtained in Example 1 was added to a commercially available fresh cream (C) to prepare a curculin-containing fresh cream (D).
コーヒーゼリー(A)を溶解後、カップに注ぎ、冷却
してゼリー状に固めたものの上に、生クリーム(D)に
砂糖10%を加えてホイップしたものを約10g絞って、ク
リームコーヒーゼリーを得た。また、コーヒーゼリー
(B)の上に、生クリーム(C)に砂糖10%を加えてホ
イップしたものをのせたクリームコーヒーゼリーを、上
記と同様にして得た。After dissolving the coffee jelly (A), pour it into a cup, cool and solidify it into a jelly, squeeze about 10 g of whipped fresh cream (D) with 10% sugar and squeeze the cream coffee jelly. Obtained. Also, cream coffee jelly obtained by placing whipped sugar (10%) on fresh cream (C) on coffee jelly (B) was obtained in the same manner as above.
ホイップクリーム(D)を食した後コーヒーゼリー
(A)を食したところ、ホイップクリーム(C)を食し
た後コーヒーゼリー(B)を食した場合と同様の風味を
感じた。When the coffee jelly (A) was eaten after eating the whipped cream (D), the same flavor as when the coffee jelly (B) was eaten after eating the whipped cream (C) was felt.
実施例6 実施例5で用いたホイップクリーム(D)の代りに、
実施例1で得られた粗クルクリンを0.2%加えて作成し
た可食性フィルム0.2gを乾燥防止をかねて、実施例5の
コーヒーゼリー(A)の表面に薄膜状においた。この可
食性フィルムを食した後コーヒーゼリー(A)を食した
ところ、実施例5のコーヒーゼリー(B)を食した場合
と同様の風味を感じた。Example 6 Instead of the whipped cream (D) used in Example 5,
0.2 g of the edible film prepared by adding 0.2% of the crude curculin obtained in Example 1 was placed on the surface of the coffee jelly (A) of Example 5 in a thin film form for the purpose of preventing drying. When the coffee jelly (A) was eaten after eating this edible film, the same flavor as when the coffee jelly (B) of Example 5 was eaten was felt.
実施例7 クルクリゴ・ラチフォリア(Curculigo latifolia)
の果実の乾燥物30gに600mlの水を加え、ミキサーで2分
間ホモジナイズした後、10000r.p.m.で30分間遠心分離
した。上澄(着色している)を除去後、残査に600mlの
水を加え、ホモジナイズ、遠心分離、上澄除去を、上澄
が着色しなくなるまで4回くりかえし、残査を得た。Example 7 Curculigo latifolia
After adding 600 ml of water to 30 g of the dried fruit, the mixture was homogenized with a mixer for 2 minutes and then centrifuged at 10,000 rpm for 30 minutes. After removing the supernatant (colored), 600 ml of water was added to the residue, homogenization, centrifugation, and removal of the supernatant were repeated 4 times until the supernatant was no longer colored, and a residue was obtained.
次に、この残査に0.5MNaCl水溶液250mlを加え、ミキ
サーで2分間ホモジナイズした後、吸引濾過した。濾液
分取後、残査にさらに0.5MNaCl水溶液250mlを加え、ホ
モジナイズ及び吸引濾過を行い、濾液を分取した。Next, 250 ml of 0.5 M NaCl aqueous solution was added to this residue, and the mixture was homogenized with a mixer for 2 minutes and then suction filtered. After collecting the filtrate, 250 ml of 0.5 M NaCl aqueous solution was further added to the residue, homogenization and suction filtration were performed, and the filtrate was collected.
次に、濾液を合わせた後、30000r.p.m.で1時間遠心
分離し、上澄(クルクリン粗抽出液)を得た。Next, the filtrates were combined and then centrifuged at 30,000 rpm for 1 hour to obtain a supernatant (cruculin crude extract).
この粗抽出液から脱塩、乾燥して、クルクリン含有物
質(粗クルクリン)を得た。The crude extract was desalted and dried to obtain a curculin-containing substance (crude curculin).
実施例8 実施例7で得られたクルクリン粗抽出液500mlを限外
濾過により、30mlまで濃縮した後、この濃縮液に0.01M
リン酸バッファー(pH6.8)70mlを加え、100mlとしたも
のを試料液として、CM−セファローズカラムクロマトグ
ラフィーを行った〔CM−セファローズCL−6B。0.01Mリ
ン酸バッファー(pH6.8)で平衡化したもの。〕。ベッ
ドボリューム130ml及びベッド高さ17cmのカラムに試料
液を流下し、0.01Mリン酸バッファー(pH6.8)で洗浄し
た後、0−1.0MNaCl/0.01Mリン酸バッファー(pH6.8)
のグラディエントで溶出し、活性画分を集めた。Example 8 500 ml of the crude curculin extract obtained in Example 7 was concentrated to 30 ml by ultrafiltration, and then 0.01 M was added to the concentrate.
CM-Sepharose column chromatography was performed using 70 ml of phosphate buffer (pH 6.8) added to 100 ml as a sample solution [CM-Sepharose CL-6B. Equilibrated with 0.01M phosphate buffer (pH 6.8). ]. The sample solution was allowed to flow down to a column with a bed volume of 130 ml and a bed height of 17 cm, washed with 0.01 M phosphate buffer (pH 6.8), and then 0-1.0 M NaCl / 0.01 M phosphate buffer (pH 6.8).
Was eluted with a gradient of and the active fractions were collected.
この活性画分から、脱塩、乾燥して、クルクリン含有
物質(粗精製クルクリン)を得た。The active fraction was desalted and dried to obtain a curculin-containing substance (crude purified curculin).
実施例9 実施例8で得られた活性画分を限外濾過で濃縮した
後、ゲルカラム(東洋ソーダ製TSKゲルG3000SW)を用い
たHPLCにかけ、0.01Mリン酸バッファー(pH6.8)で溶出
した。Example 9 The active fraction obtained in Example 8 was concentrated by ultrafiltration, subjected to HPLC using a gel column (TSK gel G3000SW manufactured by Toyo Soda), and eluted with 0.01M phosphate buffer (pH 6.8). .
クルクリンは、強い活性を持つシャープな1ピークと
して溶出された。Curculin was eluted as one sharp peak with strong activity.
この活性画分を脱塩、乾燥して、精製クルクリンを得
た。This active fraction was desalted and dried to obtain purified curculin.
なお、この精製クルクリンを8.0M尿素の存在下でSDS
−PAGEにかけたところ、分子量12500ダルトン(dalto
n)の所に単一バンドを示し、クルクリン純品であるこ
とが確認された。The purified curculin was treated with SDS in the presence of 8.0M urea.
When subjected to −PAGE, the molecular weight was 12500 daltons (dalto
A single band was shown at n), and it was confirmed to be pure curculin.
クルクリゴ・ラチフォリアの果実の乾燥物30gあたり
の、各実施例で得られたクルクリンの蛋白量、活性収率
及び比活性は下記第6表に示す通りであった。The amount of protein, activity yield and specific activity of curculin obtained in each example per 30 g of dried product of Curculigo latifolia fruit were as shown in Table 6 below.
実施例10 実施例7で得られた粗クルクリン100mgと食塩1mgを水
20mlに溶解し、0.5%水溶液とした。この水溶液は室温
(25℃前後)に1ケ月放置した後も安定であった。この
水溶液1mlを口中に1分間含み吐き出した後、0.02Mクエ
ン酸、水、紅茶(ブラック)をそれぞれ飲食した。その
時の甘味の感じ方は下記第7表に示す通りであった。 Example 10 100 mg of crude curculin obtained in Example 7 and 1 mg of sodium chloride were added to water.
It was dissolved in 20 ml to give a 0.5% aqueous solution. This aqueous solution was stable even after being left at room temperature (around 25 ° C) for 1 month. After 1 ml of this aqueous solution was put into the mouth for 1 minute and spit out, 0.02 M citric acid, water, and black tea (black) were respectively consumed. The way of feeling the sweetness at that time was as shown in Table 7 below.
紅茶(ブラック)の場合は普通の砂糖添加の紅茶と同
様の味を感じた。 In the case of black tea (black), I felt the same taste as ordinary sugar-added black tea.
実施例11 下記第8表に示す配合にて、ブラックのコーヒーゼリ
ー(A)及び砂糖入りのコーヒーゼリー(B)を作成し
た。Example 11 Black coffee jelly (A) and sugar-containing coffee jelly (B) were prepared according to the formulation shown in Table 8 below.
一方、市販の生クリーム(C)に実施例7で得られた
粗クルクリン0.05%を加え、クルクリン含有生クリーム
(D)を作成した。 On the other hand, 0.05% of the crude curculin obtained in Example 7 was added to a commercially available fresh cream (C) to prepare a curculin-containing fresh cream (D).
コーヒーゼリー(A)を溶解後、カップに注ぎ、冷却
してゼリー状に固めたものの上に、生クリーム(D)に
砂糖10%を加えてホイップしたものを約10g絞って、ク
リームコーヒーゼリーを得た。また、コーヒーゼリー
(B)の上に、生クリーム(C)に砂糖10%を加えてホ
イップしたものをのせたクリームコーヒーゼリーを、上
記と同様にして得た。After dissolving the coffee jelly (A), pour it into a cup, cool and solidify it into a jelly, squeeze about 10 g of whipped fresh cream (D) with 10% sugar and squeeze the cream coffee jelly. Obtained. Also, cream coffee jelly obtained by placing whipped sugar (10%) on fresh cream (C) on coffee jelly (B) was obtained in the same manner as above.
ホイップクリーム(D)を食した後コーヒーゼリー
(A)を食したところ、ホイップクリーム(C)を食し
た後コーヒーゼリー(B)を食した場合と同様の風味を
感じた。When the coffee jelly (A) was eaten after eating the whipped cream (D), the same flavor as when the coffee jelly (B) was eaten after eating the whipped cream (C) was felt.
実施例12 実施例11で用いたホイップクリーム(D)の代りに、
実施例7で得られた粗クルクリンを0.5%加えて作成し
た可食性フィルム0.2gを乾燥防止をかねて、実施例11の
コーヒーゼリー(A)の表面に薄膜状においた。この可
食性フィルムを食した後コーヒーゼリー(A)を食した
ところ、実施例11のコーヒーゼリー(B)を食した場合
と同様の風味を感じた。Example 12 Instead of the whipped cream (D) used in Example 11,
0.2 g of the edible film prepared by adding 0.5% of the crude curculin obtained in Example 7 was put on the surface of the coffee jelly (A) of Example 11 in a thin film form for the purpose of preventing drying. When the coffee jelly (A) was eaten after eating this edible film, the same flavor as when the coffee jelly (B) of Example 11 was eaten was felt.
実施例13〜18 実施例7で用いた0.5MNaCl水溶液に代えて、0.5MKC1
水溶液(実施例13)、0.5MCaCl2水溶液(実施例14)、
0.5MNaHCO3水溶液(実施例15)、0.5MMgCO3水溶液(実
施例16)、0.2MKH2PO4水溶液(実施例17)、0.2MNaH2PO
4水溶液(実施例18)をそれぞれ用いた以外は、実施例
7と同様にして、クルクリン含有物質(粗クルクリン)
をそれぞれ得た。Examples 13 to 18 Instead of the 0.5 M NaCl aqueous solution used in Example 7, 0.5 MKC1
Aqueous solution (Example 13), 0.5M CaCl 2 aqueous solution (Example 14),
0.5M NaHCO 3 aqueous solution (Example 15), 0.5MMgCO 3 aqueous solution (Example 16), 0.2MKH 2 PO 4 aqueous solution (Example 17), 0.2M NaH 2 PO
4 Curculin-containing substance (crude curculin) in the same manner as in Example 7 except that 4 aqueous solutions (Example 18) were used respectively.
Respectively obtained.
本発明の新規物質クルクリンは、安定化度が高く、味
覚修飾物質として好適に用いられ、例えば、本発明のク
ルクリンまたはその含有物を事前に食しておけば、酸味
物質または水等を飲食した時に甘味を感じさせることが
でき、保存性等の理由から酸性にせざるを得ない食品、
または健康や栄養上の理由から砂糖を含有しない食品等
を食する際に好ましい甘味を感じさせることができるの
で、広範な用途に利用可能である。The novel substance curculin of the present invention has a high degree of stabilization and is suitably used as a taste-modifying substance.For example, if the curculin of the present invention or a content thereof is eaten in advance, when a sour substance or water is consumed, Foods that have a sweet taste and must be acidified for reasons such as shelf life,
Alternatively, it can be used for a wide range of purposes because it can have a preferable sweet taste when eating foods containing no sugar for health or nutritional reasons.
Claims (4)
の乾燥物から0.01M以上の濃度の塩の水溶液で抽出する
ことによって得られ、甘味を感じさせる味覚修飾作用を
有し、等電点7.1、分子量約12500ダルトンであるモノマ
ーのダイマーである蛋白質クルクリン。1. Obtained by extracting from a fruit of Curculigo latifolia or a dried product thereof with an aqueous solution of a salt having a concentration of 0.01 M or more, and having a taste-modifying action to make sweet taste, an isoelectric point of 7.1 and a molecular weight of about The protein curculin, which is a dimer of the monomer that is 12500 daltons.
有する食品。2. A food containing the protein curculin according to claim 1.
レー状または乳化物状に加工されている請求項(2)記
載の食品。3. The food according to claim 2, which is processed into powder, solution, sheet, tablet, spray or emulsion form.
の乾燥物から0.01M以上の濃度の塩の水溶液で、請求項
(1)記載の蛋白質クルクリンを抽出することを特徴と
する蛋白質クルクリンの抽出方法。4. A method for extracting the protein curculin according to claim 1, which comprises extracting the protein curculin according to claim 1 from a fruit of Curculigo latifolia or a dried product thereof with an aqueous solution of a salt having a concentration of 0.01 M or more.
Priority Applications (43)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP63277717A JP2509989B2 (en) | 1988-06-21 | 1988-11-02 | Protein curculin and its use |
NZ229419A NZ229419A (en) | 1988-06-21 | 1989-06-06 | Curuculigo latifolia fruits, protein extract from them and their use as taste modifiers |
NO892332A NO175261C (en) | 1988-06-21 | 1989-06-07 | Curuculin protein and its use |
MYPI89000766A MY104031A (en) | 1988-06-21 | 1989-06-08 | Protein curculin and application of the same |
CA000602872A CA1336855C (en) | 1988-06-21 | 1989-06-15 | Protein curculin and application of the same |
CA000602870A CA1337025C (en) | 1988-06-21 | 1989-06-15 | Method for processing taste-modifier |
CA000602871A CA1337026C (en) | 1988-06-21 | 1989-06-15 | Reinforcer for taste-modifier |
CA000602869A CA1337024C (en) | 1988-06-21 | 1989-06-15 | Method for stabilizing taste-modifier |
FI892986A FI98782C (en) | 1988-06-21 | 1989-06-19 | Curculin protein, its method of preparation and its use |
DE8989111180T DE68903145T2 (en) | 1988-06-21 | 1989-06-20 | METHOD FOR TREATING A TASTE CHANGER. |
ES89111180T ES2052821T3 (en) | 1988-06-21 | 1989-06-20 | PREPARATION METHOD OF A FLAVOR MODIFICATION AGENT. |
EP89111180A EP0351567B1 (en) | 1988-06-21 | 1989-06-20 | Method for processing taste-modifier |
AT89111181T ATE124212T1 (en) | 1988-06-21 | 1989-06-20 | METHOD FOR STABILIZING A FLAVOR MODIFYING AGENT. |
DE68927083T DE68927083T2 (en) | 1988-06-21 | 1989-06-20 | The protein curuculin and use of the same |
EP89111178A EP0350667B1 (en) | 1988-06-21 | 1989-06-20 | Composition containing reinforcer for taste-modifier |
ES89111181T ES2075838T3 (en) | 1988-06-21 | 1989-06-20 | PROCEDURE TO STABILIZE A FLAVOR MODIFIER. |
EP89111181A EP0347832B1 (en) | 1988-06-21 | 1989-06-20 | Method for stabilizing taste-modifier |
AU36623/89A AU633827B2 (en) | 1988-06-21 | 1989-06-20 | Protein curculin and application of the same |
DK303789A DK303789A (en) | 1988-06-21 | 1989-06-20 | PROTEIN CURUCULIN AND ITS USE AS A TASTE MODIFACTOR |
ES198989111178T ES2040413T3 (en) | 1988-06-21 | 1989-06-20 | COMPOSITION CONTAINING A FLAVOR MODIFYING BOOSTER. |
AT89111178T ATE87430T1 (en) | 1988-06-21 | 1989-06-20 | MIXTURE CONTAINING ENHANCEMENT AGENT FOR A TASTE MODIFIER. |
ES89111179T ES2092989T3 (en) | 1988-06-21 | 1989-06-20 | CURCULIN PROTEIN AND USE OF THE SAME. |
DE68923226T DE68923226T2 (en) | 1988-06-21 | 1989-06-20 | Process for stabilizing a flavor changing agent. |
AT89111179T ATE142228T1 (en) | 1988-06-21 | 1989-06-20 | THE PROTEIN CURUCULIN AND USE OF THE SAME |
AT89111180T ATE81260T1 (en) | 1988-06-21 | 1989-06-20 | METHOD OF TREATMENT OF A TASTE MODIFIER. |
SU894614373A RU2033057C1 (en) | 1988-06-21 | 1989-06-20 | Method for preparation of taste modifier, composition for manufacture of chewing gum and mouthwash |
DE8989111178T DE68905690T2 (en) | 1988-06-21 | 1989-06-20 | MIXTURE CONTAINING REINFORCEMENT AGENT FOR A TASTE CHANGER. |
DE68928820T DE68928820T2 (en) | 1988-06-21 | 1989-06-20 | Process for stabilizing a flavor changing agent |
EP89111179A EP0351566B1 (en) | 1988-06-21 | 1989-06-20 | Protein curuculin and application of the same |
EP93111742A EP0577147B1 (en) | 1988-06-21 | 1989-06-20 | Method for stabilizing taste-modifier |
AT93111742T ATE171342T1 (en) | 1988-06-21 | 1989-06-20 | METHOD FOR STABILIZING A FLAVOR MODIFYING AGENT |
KR1019890008590A KR930006205B1 (en) | 1988-06-21 | 1989-06-21 | Protein curuculin and application of the same |
SG1996003520A SG50520A1 (en) | 1988-06-21 | 1989-06-21 | Protein curuculin and application of the same |
US07/598,799 US5178900A (en) | 1988-06-21 | 1990-10-16 | Method for stabilizing taste-modifier |
US07/649,373 US5176937A (en) | 1988-06-21 | 1991-01-31 | Reinforcer for taste-modifier |
US07/651,060 US5178899A (en) | 1988-06-21 | 1991-02-05 | Method for processing taste-modifier |
US07/655,184 US5242693A (en) | 1988-06-21 | 1991-02-13 | Protein curuculin and application of the same |
US07/963,916 US5256439A (en) | 1988-06-21 | 1992-10-20 | Method for stabilizing taste-modifier |
GR920402384T GR3006059T3 (en) | 1988-06-21 | 1992-10-22 | |
GR950402358T GR3017247T3 (en) | 1988-06-21 | 1995-08-30 | Method for stabilizing taste-modifier. |
CN96122700A CN1158721A (en) | 1988-06-21 | 1996-10-28 | Protein colukrine and its use |
CN96122699A CN1158704A (en) | 1988-06-21 | 1996-10-28 | Protein colukrine and its use |
GR960403017T GR3021646T3 (en) | 1988-06-21 | 1996-11-14 | Protein curuculin and application of the same |
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP15314388 | 1988-06-21 | ||
JP63-153143 | 1988-06-21 | ||
JP63277717A JP2509989B2 (en) | 1988-06-21 | 1988-11-02 | Protein curculin and its use |
Publications (2)
Publication Number | Publication Date |
---|---|
JPH02104263A JPH02104263A (en) | 1990-04-17 |
JP2509989B2 true JP2509989B2 (en) | 1996-06-26 |
Family
ID=26481853
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
JP63277717A Expired - Fee Related JP2509989B2 (en) | 1988-06-21 | 1988-11-02 | Protein curculin and its use |
Country Status (1)
Country | Link |
---|---|
JP (1) | JP2509989B2 (en) |
Families Citing this family (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPH0427356A (en) * | 1990-05-22 | 1992-01-30 | Yoshie Kurihara | Method for stabilizing taste modifying composition and taste modifying substance |
RU2113440C1 (en) * | 1991-03-04 | 1998-06-20 | Курихара Есие | Dna fragment encoding curculin b showing taste modifying agent activity, curculin b showing taste modifying agent activity and a method of its preparing |
JPH0597896A (en) * | 1991-10-01 | 1993-04-20 | Yoshie Kurihara | Taste modifying substance |
Family Cites Families (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPS63277721A (en) * | 1987-05-09 | 1988-11-15 | Nkk Corp | Manufacture of rail combining high strength with high toughness |
JPS63277718A (en) * | 1987-05-11 | 1988-11-15 | Kawasaki Steel Corp | Manufacture of grain-oriented silicon steel sheet excellent in magnetic characteristic |
JPS63285474A (en) * | 1987-05-18 | 1988-11-22 | Keiyo Seiki Kk | Measuring apparatus of radiated interference wave |
-
1988
- 1988-11-02 JP JP63277717A patent/JP2509989B2/en not_active Expired - Fee Related
Also Published As
Publication number | Publication date |
---|---|
JPH02104263A (en) | 1990-04-17 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
US11351214B2 (en) | Methods for making yerba mate extract composition | |
RU2062037C1 (en) | Chewing rubber | |
US5242693A (en) | Protein curuculin and application of the same | |
JP2509989B2 (en) | Protein curculin and its use | |
JP2509993B2 (en) | Taste modifier for substances with bitterness, astringency or harshness | |
JP3246738B2 (en) | Saponin and amino acid-containing composition | |
KR930006205B1 (en) | Protein curuculin and application of the same | |
RU2090097C1 (en) | Emulsified composition for taste modification of food products | |
US5178899A (en) | Method for processing taste-modifier | |
JP2509990B2 (en) | Taste modifier for sour substances | |
CA1337025C (en) | Method for processing taste-modifier | |
JP2509991B2 (en) | Beverages containing novel taste modifiers | |
JPH11180999A (en) | Purified silk fibroin solution, its production and beverage and food containing the same | |
JPH0284161A (en) | Method for processing taste modifying substance | |
JPS60160858A (en) | Low-salt seasoning solution of oyster flavor | |
JPH0614857B2 (en) | Taste modifier spray | |
US4567054A (en) | Sweetener composition | |
JPH0813253B2 (en) | Processing method of taste modifier | |
JPH0813244B2 (en) | Chewing gum composition | |
JPH03147742A (en) | Coffee drink | |
JPS62175155A (en) | Vinegared solid egg | |
JPH0813254B2 (en) | Effect enhancer of taste modifier | |
JPS62208253A (en) | Production of novel konjak | |
JPH11146759A (en) | Coffee-containing drink compounded with 10-hydroxy-2-decenoic acid | |
JPH0279951A (en) | Treatment of fruit |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
S533 | Written request for registration of change of name |
Free format text: JAPANESE INTERMEDIATE CODE: R313533 |
|
R350 | Written notification of registration of transfer |
Free format text: JAPANESE INTERMEDIATE CODE: R350 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
LAPS | Cancellation because of no payment of annual fees |