JP2534043B2 - Method for separating active ingredients from cruciferous plants - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial field of application] The present invention relates to a method for separating and recovering from the cruciferous plants such as horseradish and radish, the pungency component and the active component of active peroxidase contained in the plant. In particular, the present invention relates to a method for separating an active ingredient from a crucifer plant capable of extracting and collecting these active ingredients simultaneously and separately.

Cruciferous plants such as horseradish and radish mainly contain an endogenous substrate thioglycoside (sinigrin), an endogenous thioglycosidase (enzyme myrosinase), peroxidase and the like.

Sinigrin is usually used as a spice ingredient by causing an enzymatic reaction in the presence of water and the enzyme myrosinase to produce an allyl isothiocyanate which is a spicy ingredient. Further, peroxidase is used, for example, as a clinical test reagent for the determination of glucose, cholesterol, phospholipids or uric acid, and is also widely used as a reagent for food analysis, and these are all cruciferous plants such as horseradish. Has been extracted and separated from.

[Conventional technology]

Conventionally, the above-mentioned active ingredients, the pungent ingredient and peroxidase, have conventionally been separately extracted and separated. That is, for example, the spicy ingredient is grated with horseradish, which undergoes an enzymatic reaction and is then extracted and separated by steam distillation at a temperature of about 110 ° C. Peroxidase is grated with horseradish separately and then in saline. The solution was extracted and separated by immersing in agitating.

[Problems to be solved by the invention]

However, in the above-mentioned extraction and separation of the pungency component, peroxidase is inactivated due to the high temperature, and in the extraction and separation of peroxidase, the pungency component is volatilized or deteriorated without being recovered. The simultaneous extraction and recovery of oxidase has been strongly desired.

An object of the present invention is to provide a method for separating an active ingredient from a cruciferous plant, which is capable of simultaneously extracting and recovering a pungent ingredient and peroxidase, and which has improved the above-mentioned drawbacks of the prior art.

[Means for solving problems]

In order to achieve the above-mentioned object, according to the present invention, a cruciferous plant as a raw material is subjected to an enzymatic reaction in an extraction tank to generate a pungency component, and supercritical carbon dioxide or liquefied carbonic acid is added to this tank. Introducing and extracting the pungent component into the supercritical carbon dioxide or liquefied carbonic acid, then introducing supercritical carbon dioxide or liquefied carbonic acid containing the pungent component into a separation tank to separate and recover the pungent component, The active peroxidase is separated and recovered from the residue in the extraction tank, and the pungency component and the active component of the active peroxidase contained in the cruciferous plant are separately collected at the same time.

Hereinafter, the present invention will be described in detail with reference to the accompanying drawings and using horseradish as a crucifer.

First, raw horseradish is put into the cooling tank 1 and frozen at a low temperature below the freezing point, and then freeze-pulverized by the pulverizer 2 while maintaining this frozen state. The coolant in the cooling tank 1 is liquid nitrogen or the like. The crusher 2 is an ordinary crusher or a grater.

Next, the frozen substance pulverized by the pulverizer 2 is filled in the extraction tank 3 and heated to cause an enzymatic reaction to generate a pungency component (allyl isothiocyanate). When dry horseradish is used as the raw material, water is introduced into the extraction tank 3 by an entrainer device and the water content is adjusted to 10% or more, preferably 30 to 50% (both by weight) to carry out an enzymatic reaction. Cause Further, in the case of crushing at room temperature or grazing at room temperature, the enzyme reaction is caused as it is.

Furthermore, a solvent of supercritical carbon dioxide or liquefied carbonic acid is introduced into the extraction tank 3 at the same time as the above-mentioned enzymatic reaction, and the pungent component produced by the enzyme reaction is extracted into the solvent of supercritical carbon dioxide or liquefied carbonic acid. . At the time of the introduction, in the case of supercritical carbon dioxide, a temperature of 30 ° C or higher, a pressure of 75
It is introduced under atmospheric pressure or higher, and in the case of liquefied carbonic acid, for example, the temperature is 20 ° C. and the pressure is 200 atmospheric pressure.

The solvent containing the pungency component thus obtained is introduced into the separation tank 4, decompressed and phase-separated into a gas phase and a liquid phase, and released from the separation tank 4 as a gas, which is a useful substance. Only the separated and recovered.

Further, in the present invention, peroxidase is extracted and separated from the residue remaining in the extraction tank 3 by a usual method.

[Work]

Since supercritical carbon dioxide or liquefied carbonic acid is used as the extraction solvent in the present invention, the peroxidase in the residue in the extraction tank 3 remains without being killed, and by recovering this from the residue, the pungency component and the peroxidase are separated. Simultaneous extraction and collection becomes possible.

〔Example〕

Example 1 100 kg of fresh horseradish (water content about 70%) was grated at room temperature and filled in an extraction tank. The temperature inside this extraction tank is 40 ℃
Kept at 40 ° C and 200 atm of supercritical carbon dioxide was introduced,
Extraction was performed for about 2 hours with stirring. Next, this supercritical carbon dioxide was introduced into a separation tank through a pressure reducing valve, and was depressurized to 50 atm to release supercritical carbon dioxide as carbon dioxide gas to obtain 0.3 kg of allyl isothiocyanate as a pungency component.

Then, the paste-like horseradish residue remaining in the extraction tank was transferred to another extraction tank, and peroxidase was extracted and separated. This extraction separation was performed by the following method.

To the extraction residue, add 140 water, stir well, let stand overnight, add a mixture of ethanol and chloroform (2: 1) 54, and add 0.
Stir for 20 minutes at ℃ and centrifuge (3,000 rpm for 15 minutes) to obtain a supernatant. Then, it is adjusted to 5.5 with 2N sodium hydroxide and concentrated under reduced pressure at 30 ° C or lower to 40. Ammonium sulfate was added to this (250 g
/) And remove the resulting precipitate by centrifugation. Ammonium sulfate was further added to the supernatant (250 g /), and the resulting precipitate was collected by centrifugation. The precipitate is suspended in 0.2M phosphate buffer (PH7) 1 and dialyzed overnight. The precipitate is removed by centrifugation to obtain a supernatant. It is cooled and 1.5 volumes of ethanol are added and the resulting precipitate is collected. This is dissolved in water and purified by alcoholic precipitation. That is, at the first time, the sections that precipitate by the addition of ethanol (1.2 to 2.0 volumes) are collected, and the sections that dissolve in water and precipitate again by the addition of ethanol (1.3 to 1.9 volumes) are collected. Dissolve this in water 1 and add ammonium sulphate to take a section where the degree of saturation precipitates at 0.56 to 0.70. Dissolve in a small amount of water and add ammonium sulphate saturated solution dropwise to give a slight turbidity. When cooled and added with crystal seeds and left at room temperature, crystals were precipitated in several hours to obtain peroxidase.

Example 2 100 kg of horseradish was grated in the same manner as in Example 1 and filled in an extraction tank. The temperature in this extraction tank was kept at 20 ° C., liquefied carbonic acid at 20 ° C. and 200 atm was introduced, and extraction was carried out for 2 hours while stirring. Next, this liquefied carbonic acid was introduced into a separation tank through a pressure reducing valve, and the pressure was reduced to 50 atm to release the liquefied carbonic acid as carbon dioxide gas to obtain 0.32 kg of allyl isothiocyanate as a pungency component.

Then, the paste-like horseradish residue remaining in the extraction tank was transferred to another extraction tank, and peroxidase was extracted and separated by the same method as in Example 1.

〔The invention's effect〕

As described above, since supercritical carbon dioxide or liquefied carbonic acid is used as the extraction solvent in the present invention, it is possible to separate and recover both active ingredients of the pungency component and peroxidase at the same time.

[Brief description of drawings]

The drawings are block diagrams showing the steps of the present invention. 1 ... Cooling tank, 2 ... Crusher, 3 ... Extraction tank, 4 ... Separation tank.

Claims (1)

(57) [Claims] 1. A cruciferous plant as a raw material is subjected to an enzymatic reaction in an extraction tank to produce a pungent ingredient, and supercritical carbon dioxide or liquefied carbonic acid is introduced into this tank to remove the pungent ingredient from the above-mentioned pungent ingredient. Extraction into critical carbon dioxide or liquefied carbonic acid, then introducing supercritical carbon dioxide or liquefied carbonic acid containing the pungent component into a separation tank to separate and recover the pungent component, and further activate from the residue in the extraction tank A method for separating an active ingredient from a crucifer plant, which comprises separating and collecting peroxidase to simultaneously separate and collect the pungency component and the active component of active peroxidase contained in the crucifer plant.