WO2015186806A1

WO2015186806A1 - Placenta extract and method for producing placenta extract

Info

Publication number: WO2015186806A1
Application number: PCT/JP2015/066253
Authority: WO
Inventors: 暁史保母; 靖晃伊東; 中村　真也; 広瀬　直宏
Original assignee: イビデン株式会社
Priority date: 2014-06-04
Filing date: 2015-06-04
Publication date: 2015-12-10
Also published as: JPWO2015186806A1

Abstract

Provided is a placenta extract that contains an active ingredient at a high concentration and is scarcely colored and, therefore, suitably usable for cosmetics and foods in which appearance and flavor are highly appreciated. The placenta extract having an absorbance of 0.10 or less at a wavelength of 390 nm and containing 0.10 mass% or more of nitrogen is suitably usable for cosmetics and foods in which appearance and flavor are highly appreciated.

Description

Placenta extract and method for producing placenta extract

The present invention relates to a placenta extract using placenta as a raw material and a method for producing a placenta extract. In particular, it relates to a placenta extract exhibiting specific absorbance and nitrogen content and a method for producing a placenta extract.

Placenta is the placenta of mammals. In recent years, it has been used as health foods, cosmetic materials, pharmaceuticals, etc. due to its superior functionality represented by its antioxidant properties, ability to promote collagen production, collagenase inhibitory activity, and antihypertensive effect. Has been. In these uses, in order to exhibit the excellent functionality of the placenta, it is necessary to lower the molecular weight and solubilize the protein in the placenta to a peptide having excellent absorbability and functionality.

Conventionally, a method using an enzyme treatment is known as a method for obtaining a placenta extract in which placenta is reduced in molecular weight and solubilized. For example, Patent Document 1 includes adding a yeast extract to a supernatant obtained by hydrolyzing a residue removed when extracting a high molecular protein such as a water-soluble protein from human placenta tissue with a proteolytic enzyme. A method for producing a melanin production inhibitor is disclosed. Patent Document 2 discloses a cosmetic composition containing a water-soluble component obtained by hydrolyzing pig and / or horse placenta by enzymatic treatment, and a phenol derivative.

JP-A-53-142515 Japanese Patent Laid-Open No. 2002-212046

However, in the conventional extraction method by enzyme treatment, the addition of the enzyme is natural, but it is necessary to add other auxiliary agents. Specifically, in addition to the enzyme, an enzyme reaction auxiliary agent such as a pH adjuster or an enzyme stabilizer is added to the placenta material. There is a problem that the concentration of the active ingredient is diluted by the inclusion of these additives and a problem that a component derived from non-placenta is mixed.

Furthermore, in order to blend placenta extract into cosmetics, foods, etc., particularly liquid products, it is required to have a high concentration of active ingredients such as low molecular weight peptides, and at the same time, it is also required to have low color and odor. Therefore, it is common to subject the placenta extract treated with an enzyme to a decolorization step using an ion exchange resin or activated carbon. However, when the decolorization step is performed on the enzyme-treated placenta extract, there is a problem that the active ingredients are significantly reduced along with the color and odor.

As a means for increasing the concentration of the active ingredient, a method of concentrating the extract obtained through the extraction process and decoloring process by enzyme treatment is conceivable, but the color and odor increase along with the active ingredient concentration. Not suitable for use in cosmetics and foods where appearance and flavor are important factors. In addition, as a means for preventing the reduction of the active ingredient, it is conceivable that the decolorization process is a simple method, but in this case, the decolorization is incomplete, and a placenta extract colored yellow to brown is obtained. Therefore, it is not suitable for use in cosmetics and foods where appearance and flavor are important factors.

The present invention has been made in view of these problems, and an object thereof is to provide a placenta extract having a high concentration of active ingredients and little coloring.

The present invention relates to a placenta extract having an absorbance at a wavelength of 390 nm of 0.10 or less and a nitrogen content of 0.10% by mass or more.

A placenta extract having an absorbance of 0.01 to 0.10 is preferred.

A placenta extract having a nitrogen content of 0.10 to 1.0% by mass is preferred.

It is preferable that the placenta extract has a total peptide content of 70 to 99.5% by mass in the total solid content of the placenta extract.

The placenta extract is preferably a placenta extract in which the content of low molecular weight peptides having a molecular weight of 3000 or less in the total solid content of the placenta extract is 40 to 99.5% by mass.

It is preferable that the placenta extract has a free amino acid content of 0.01 to 30% by mass in the total solid content of the placenta extract.

A placenta extract in which the content of components having a molecular weight of 3000 or less in the total solid content of the placenta extract is 100% is preferable.

It is preferably a placenta extract that is extracted through a subcritical treatment of the placenta raw material.

Furthermore, a placenta extract extracted through activated carbon treatment is preferred.

It is preferable that the temperature of the subcritical treatment is 160 to 200 ° C. and the pressure is equal to or higher than the saturated vapor pressure.

The subcritical processing time is preferably 5 to 30 minutes.

In the present invention, the absorbance at a wavelength of 390 nm is 0.10 or less,
A placenta extract production method for obtaining a placenta extract having a nitrogen content of 0.10% by mass or more,
An extraction process for subcritical processing of the placenta to obtain a subcritical processed product,
The present invention relates to a method for producing a placenta extract, comprising a solid-liquid separation step of separating the subcritical processed product into an extract and a raw material residue.

It is preferable that the placenta extract production method includes a decolorization step after the subcritical processing extraction step.

In the subcritical process, a placenta extract production method in which the temperature of the subcritical process is 160 to 200 ° C. and the pressure is equal to or higher than the saturated vapor pressure is preferable.

A method for producing a placenta extract in which the subcritical processing time in the subcritical processing is 5 to 30 minutes is preferable.

A method for producing a placenta extract in which the treatment temperature in the decolorization step is 0 to 80 ° C. is preferred.

A method for producing a placenta extract in which the treatment time in the decolorization step is 3 to 120 minutes is preferable.

A method for producing a placenta extract in which the number of treatments in the decolorization step is 2 to 10 times is preferable.

In the placenta extract of the present invention, the placenta extract having an absorbance at a wavelength of 390 nm of 0.10 or less and a nitrogen content of 0.10% by mass or more has a high active ingredient concentration and is colored. It is possible to provide a placenta extract that is suitable for use in cosmetics and foods, which has a small amount of placenta extract, and its appearance and flavor are important factors. Further, in the method for producing a placenta extract of the present invention, the absorbance at a wavelength of 390 nm is 0.10 or less, the nitrogen content is 0.10% by mass or more, the concentration of the active ingredient is high, and the placenta with little coloring. It is possible to provide a production method for obtaining a placenta extract suitable for use in cosmetics and foods, which is an extract, the appearance and flavor of which are important factors.

It is a graph which shows an example of the calibration curve in elution time-molecular weight (common logarithm) in the molecular weight distribution measurement of a placenta extract. It is a graph which shows an example of the result of having measured the placenta extract by the high performance liquid chromatography. It is a graph which shows the light absorbency (390 nm) and nitrogen content of the extract of an Example and a comparative example.

The placenta extract of the present invention is an extract using placenta as a raw material, and has an absorbance at a wavelength of 390 nm of 0.10 or less and a nitrogen content of 0.1% by mass or more. By using a placenta extract having an absorbance at a wavelength of 390 nm and a nitrogen content in this range, it is a placenta extract having a high active ingredient concentration and low coloration, and a cosmetic or cosmetic product whose appearance and flavor are important factors. Placenta extract suitable for food use.

The absorbance at a wavelength of 390 nm in the placenta extract of the present invention is an index for coloring the extract from yellow to brown. This absorbance can be measured with a commercially available visible ultraviolet spectrophotometer (such as GeneQuant 1300 manufactured by GE Healthcare Japan).

The absorbance is measured by a single beam method. A cell containing a reference solution (water) is prepared. The cell of the reference solution is set in a visible ultraviolet spectrophotometer and measured at a wavelength of 390 nm. Confirm that the measurement result is 0.00. Thereafter, a cell in which the sample solution is put in the same cell as that used for the reference solution is prepared, set in a visible ultraviolet spectrophotometer, measured at a wavelength of 390 nm, and the measurement result becomes the absorbance of the sample solution.

In the placenta extract of the present invention, the absorbance at a wavelength of 390 nm is 0.10 or less, preferably 0.08 or less, and more preferably 0.05 or less. If the absorbance exceeds 0.10, the placenta extract tends to be yellowish or odorous. In addition, the closer the absorbance value is to 0, the more colorless and transparent the placenta extract is. However, the placenta extract that has undergone subcritical treatment is less than 0.01 even if the decolorization step is performed. Since it has been confirmed that this does not occur, the lower limit of the absorbance of the placenta extract of the present invention at a wavelength of 390 nm is 0.01. The absorbance of the placenta extract of the present invention at a wavelength of 390 nm is more preferably 0.028 to 0.084, and it is a practically effective concentration of active ingredient and becomes a placenta extract with little coloration. Easy to apply. In order to make the absorbance within the above numerical range, it can be adjusted by subjecting the placenta to subcritical processing and then performing decolorization processing.

The nitrogen content of the placenta extract of the present invention is the nitrogen content of free amino acids and peptides contained in the extract, and is an index of the active ingredient content in the placenta extract. This nitrogen content is measured by the Kjeldahl method (a method in which sulfuric acid is added to the test solution, organic nitrogen in the solution is converted to ammonium sulfate, the liquid is made alkaline and heated, and the generated ammonia is quantified by titration). be able to. Specifically, it can be performed according to the method described in JIS K010244.

The measurement of nitrogen content by the Kjeldahl method is performed according to the following procedure.
1. An appropriate amount (3-10 mL) of the sample solution is placed in 500 mL of a beaker, sulfuric acid (1 + 35) is added to make it weakly acidic, and the mixture is heated to 30 mL to prepare a test solution.
2. After allowing the test solution to cool, a small amount of water is added and washed into a 200 mL Kjeldahl flask.
3. Add 10 mL of sulfuric acid, 5 g of potassium sulfate and 2 g of copper (II) sulfate pentahydrate, and heat to generate white smoke of sulfuric acid, followed by heating for about 30 minutes to decompose organic matter.
4). After standing to cool, add a small amount of water, heat to dissolve, wash to a distillation flask with water to make about 300 mL.
5. A distillation flask is connected, and 200 mL of a graduated cylinder (plugged type) is used as a receiver, and 50 mL of sulfuric acid (25 mmol / L) is added. After adding 40 mL of sodium hydroxide solution (500 g / L) from the top of the distillation flask, the distillation flask is heated and distilled at a distillation rate of 5 to 7 mL / min. When about 140 mL is distilled, distillation is performed. Remove the stopper, cooler and backflow stopper, wash the contents of the cooler and the backflow inside and outside with a small amount of water, put the wash into a 200 mL graduated cylinder, add water to the 200 mL mark, Prepare a sample for sum measurement.
6). As a blank test, take 30 mL of water, perform steps 3 to 5, and prepare a blank test sample for neutralization measurement.
7). Using the whole amount of the solution obtained from the sample for neutralization measurement, titrate with a 50 mmol / L sodium hydroxide solution until the color of the solution becomes gray purple (pH 4.8). Similarly, a blank test sample for neutralization measurement is similarly titrated with a sodium hydroxide solution.
8). Add sulfuric acid (25 mmol / L) to a 500 mL Erlenmeyer flask correctly, add 5 to 7 drops of methyl red-promocresol green mixed solution until the color of the solution becomes gray purple (pH 4.8) with 50 mmol / L sodium hydroxide solution. Titration is performed to obtain a titer (mL) of a 50 mmol / L sodium hydroxide solution corresponding to sulfuric acid (25 mmol / L).

The ammonium ion concentration is determined by the following formula.
Formula X A = (ba) × f × 1000 / F × 0.902
A: Ammonium ion (NH ⁴⁺ mg / L)
b: 50 mmol / L sodium hydroxide solution (mL) corresponding to 50 mL of sulfuric acid (25 mmol / L)
a: 50 mmol / L sodium hydroxide solution (mL) required for titration
f: Factor F of 50 mmol / L sodium hydroxide solution: Sample (mL)
In addition, 0.902 is an ammonium ion equivalent amount of 1 mL of 50 mmol / L sodium hydroxide solution.

The concentration of nitrogen content is calculated by the following formula Y.
Formula YN = (b1-a1) × f × (1000 × 0.700) /V×0.7766
N: Nitrogen content (mg / L)
b1: 50 mmol / L sodium hydroxide solution (mL) required for titration of blank test
a1: 50 mmol / L sodium hydroxide solution (mL) required for titration
f: Factor V of 50 mmol / L sodium hydroxide solution: Amount of sample solution (mL)
A: Ammonium ion concentration determined by Formula X (NH ⁴⁺ mg / L)
In addition, 0.700 is a nitrogen equivalent (mg) of 1 mL of 50 mmol / L sodium hydroxide solution, and 0.7766 is a coefficient when converting ammonium ion into nitrogen equivalent. Further, the nitrogen content in the placenta extract of the present invention is expressed in mass%.

The nitrogen content of the placenta extract of the present invention is 0.10% by mass or more, preferably 0.15% by mass or more, and more preferably 0.20% by mass or more. When the nitrogen content is less than 0.10% by mass, the concentration of the active ingredient is not sufficient, and the excellent function of the placenta extract tends to be insufficient. Moreover, although the upper limit of nitrogen content is not specifically limited, It can be 1.0 mass% or less. Further, the nitrogen content of the placenta extract of the present invention is more preferably 0.15 to 0.21% by mass, and it becomes a placenta extract having a practical active ingredient concentration and little coloration. Easy to apply. In addition, in order to make nitrogen content into said numerical range, it can adjust by performing a decoloring process after performing a subcritical process of a placenta.

The extraction components in the placenta extract of the present invention are roughly classified into four components: free amino acids, low molecular peptides having a molecular weight of 3000 or less, peptides having a molecular weight of more than 3000, and other components.

In addition, the content of free amino acids, peptides and other components in the present specification is indicated by mass% of each component with respect to the total solid content of the placenta extract. In the case where the placenta extract contains a liquid component such as moisture, the placenta extract is dried or freeze-dried to remove the liquid component as the total solid content of the placenta extract.

Free amino acids include arginine, lysine, histidine, phenylalanine, tyrosine, leucine, isoleucine, methionine, valine, alanine, glycine, proline, glutamic acid (including glutamine), serine, threonine, aspartic acid (including asparagine), cystine, and tryptophan. The amino acid which 18 types of amino acids exist independently is pointed out.

Free amino acids inhibit the absorption of functional peptides into the body and may reduce the absorbability and functionality of functional peptides. In addition, when the content of free amino acids is high, the peptide content is relatively low. And the functionality of the placenta extract may be reduced. Therefore, the content of free amino acids in the total solid content of the placenta extract is preferably as small as possible, but the placenta extract that has undergone the subcritical treatment should not be smaller than 0.01 even if the decolorization step is performed. Therefore, the lower limit of the content of free amino acid is 0.01% by mass. For the reason that the functionality of the placenta extract can be sufficiently exhibited, the content of free amino acids in the total solid content of the placenta extract is preferably 30% by mass or less, more preferably 10% by mass or less, and more preferably 5% by mass or less. Is most preferred.

A low molecular weight peptide having a molecular weight of 3000 or less refers to all peptides having a molecular weight of 3000 or less, in which at least one of the 18 kinds of amino acids is bound. These low molecular weight peptides with a molecular weight of 3000 or less have good absorbability to the human body, and the absorbed components are excellent in placenta extract typified by antioxidant properties, collagen production promoting ability, collagenase inhibitory activity, and blood pressure elevation inhibiting action. It is a component that easily exhibits high functionality.

The peptide having a molecular weight of greater than 3000 refers to all peptides having a molecular weight of greater than 3000 among peptides in which at least one of the 18 amino acids is bound.

Other components refer to all components other than peptides and amino acids, including inorganic components such as minerals, lipids, and carbohydrates. These other components are difficult to remove all by separation and extraction, and are contained in an amount of 0.5% by mass or more in the total solid content of the placenta extract.

Since the absorbability to the human body is good, and the absorbed component is a component that easily exhibits excellent functionality such as antioxidant properties and collagen production promoting ability, the molecular weight in the total solid content of the placenta extract is 3000 or less. The content of the components (free amino acid, low molecular peptide having a molecular weight of 3000 or less and other components having a molecular weight of 3000 or less) is preferably 100%, and preferably no component having a molecular weight of more than 3000 is contained. By performing the subcritical treatment of the placenta and hydrolyzing it, the content of components having a molecular weight of 3000 or less in the total solid content of the placenta extract is close to 100%. In addition, in the placenta extract that has undergone the subcritical treatment and the decolorization step, the content of components having a molecular weight of 3000 or less in the total solid content of the placenta extract is further close to 100%.

The content of all peptides in the total solid content of the placenta extract of the present invention (the total of low molecular weight peptides with a molecular weight of 3000 or less and peptides with a molecular weight of more than 3000) is that the components absorbed by the human body are antioxidant and promote collagen production 70% by mass or more is preferable because it is easy to exhibit functionality such as performance. In the placenta extract that has undergone the subcritical treatment, the total peptide content in the total solid content of the placenta extract can be 70% by mass or more. Further, the total peptide content in the total solid content of the placenta extract is more preferably 75% by mass or more. When the content of the peptide is less than 70% by mass, the functionality of the placenta extract may be lowered. Further, the upper limit of the peptide content is preferably as much as possible, but it is estimated that the upper limit is 99.5% by mass excluding the other components from the total solid content.

Among peptides, the inclusion of 40-99.5% by mass of a low molecular weight peptide having a molecular weight of 3000 or less in the total solid content of the placenta extract is said to enhance the functionality of the placenta extract. Preferred for reasons. In other words, the total solid content of the placenta extract preferably contains 40 to 99.5% by mass of a low molecular weight peptide having a molecular weight of 3000 or less. The solid component can contain 40 to 99.5% by mass of a low molecular weight peptide having a molecular weight of 3000 or less.

The molecular weight distribution in the placenta extract of the present invention is measured by size exclusion chromatography using the separation mode of high performance liquid chromatography. The sample solution injected into the high performance liquid chromatography is separated in the instrument, and the molecular weight is measured. It is carried out by a method of measuring the molecular weight utilizing the fact that the components are eluted in order from the largest. For this purpose, a plurality of standard reagents with different molecular weights are specified, each standard reagent is measured by high performance liquid chromatography, peak elution time (X) and common logarithm of molecular weight (Y) By creating a calibration curve based on the relationship, the relationship between elution time and molecular weight can be determined. The elution time can be converted into molecular weight from this calibration curve.

】 The calibration curve will be explained based on examples. Seven standard reagents are used to create a calibration curve. The elution time of the peak in the high performance liquid chromatography for each molecular weight of the standard reagent is 6.85 minutes for the standard reagent 1: molecular weight 12,500 (reagent name: Cytochrome C), and the standard reagent 2: molecular weight For 6,512 (reagent name: Aprotinin), the elution time was 8.58 minutes. For standard reagent 3: molecular weight 1,423 (reagent name: Bacitracin), the elution time was 9.60 minutes. For the molecular weight 1046 (reagent name: Angiotensin II), the elution time is 9.79 minutes, and for the standard reagent 5: molecular weight 451 (reagent name: Gly-Gly-Tyr-Arg), the elution time is 10.7 minutes. The elution time was 11.7 minutes for reagent 6: molecular weight 189 (reagent name: Gly-Gly-Gly), and the elution time was 12.2 minutes for standard reagent 7: molecular weight 75 (reagent name: glycine). Based on this measurement result and the molecular weight of each standard reagent, the elution time was plotted on the X axis and the common logarithm of the molecular weight was plotted on the Y axis, and a calibration curve was created from the results. The prepared calibration curve is shown in FIG. With the calibration curve in FIG. 1, the molecular weight corresponding to the peak elution time in high performance liquid chromatography can be confirmed. The molecular weight distribution can be calculated based on the confirmed molecular weight and the peak area of each peak. In this calibration curve, it was confirmed that the molecular weight was 3000 at an elution time of 9.1 minutes. Therefore, as a result of measuring the placenta extract by constrained chromatography, if the elution time of the detected peak is within 9.1 minutes, the molecular weight exceeds 3000 and the elution time at the peak peak exceeds 9.1 minutes. In this case, the molecular weight is 3000 or less.

Here, 200 g of distilled water was put into a pig placenta, and the result of measurement by high performance liquid chromatography of a placenta extract that was subcritically processed at a processing temperature of 195 ° C., a processing pressure of 1.6 MPa, and a processing time of 30 minutes is shown. As shown in FIG. In FIG. 2, since the detected peak exceeds the elution time of 9.1 minutes, it can be confirmed that the molecular weight of the extract is 3000 or less.

The method for producing a placenta extract of the present invention includes an extraction step for subcritically treating a raw placenta to obtain a subcritical treatment, and a solid-liquid separation step for separating the extract from the raw material residue. Furthermore, you may include the decoloring process by activated carbon treatment. According to the production method, it is possible to produce a placenta extract having a high active ingredient concentration and little coloration.

In the method for producing the placenta extract of the present invention, the placenta as a raw material is not particularly limited as long as it is a mammalian placenta. From the viewpoint of easy availability, placenta such as pigs, cows, horses, sheep, wild boars, etc. preferable. Since placenta as a raw material may contain blood and other parts, it is preferable to perform a purification step such as washing before the extraction step.

The extraction step in the placenta extract production method of the present invention is a step of obtaining a subcritical processed product by subjecting the placenta raw material to subcritical processing. The subcritical treatment is a process in which a subcritical fluid as an extracting agent brought into a subcritical state under a predetermined temperature and pressure is brought into contact with a raw material to be extracted (in the present invention, a placenta) to thereby extract predetermined components from the extracted raw material. Is extracted. For example, when water is raised to a pressure of 22.12 MPa and a temperature of 374.15 ° C., it shows a state where it is neither liquid nor gas. This point is called the critical point of water, and hot water at a temperature and pressure lower than the critical point is called subcritical water. This subcritical water has an excellent component extraction action and hydrolysis action due to a decrease in dielectric constant and an improvement in ionic product.

In the method for producing the placenta extract of the present invention, when water is used as the extractant used for the subcritical treatment, it can be used in a liquid state or a gas state as long as it is a high-temperature water treatment. That is, water vapor may be supplied to the subcritical processing tank, water may be supplied, or both of them may be supplied. The temperature of water or water vapor is desirably 100 ° C. or higher, and the desired reaction field is more liable to proceed in the liquid state than in the gas. The use of water in the state of More specifically, placenta and water as an extractant are placed in a pressure vessel such as metal or ceramics to bring it into a closed state, and the water is in a subcritical state (temperature: 100 ° C or higher, pressure: saturated vapor pressure or higher). ), The extract obtained by performing the contact between them for a certain time or more can be used as a subcritical processed product.

Examples of the extractant used for the subcritical treatment in the method for producing the placenta extract of the present invention include, in addition to water, ethylene, ethane, propane, carbon dioxide, methanol, ethanol, and mixtures thereof. Among these, it is most preferable to use water from the viewpoint of safety. Next, processing conditions when the extractant is water will be described.

The subcritical processing temperature of the placenta in the method for producing the placenta extract of the present invention is preferably between 160 and 200 ° C. By setting this temperature range, it is easy to produce a low molecular peptide having a molecular weight of 3000 or less, which is a functional component. The molecular weight distribution can be measured by measuring the molecular weight distribution by high performance liquid chromatography. When the temperature of the subcritical treatment is less than 160 ° C., it tends to be difficult to produce a low molecular peptide having a molecular weight of 3000 or less. Moreover, when the temperature of subcritical processing exceeds 200 degreeC, the produced | generated low molecular peptide will raise | generate a subcritical reaction further, the production amount of a free amino acid will increase, and the production amount of the low molecular peptide whose molecular weight is 3000 or less There is a tendency to decrease.

Furthermore, the subcritical processing temperature in the method for producing the placenta extract of the present invention is more preferably 180 to 195 ° C. By setting it within this range, it becomes easy to produce a low molecular peptide having a molecular weight of 3000 or less in a shorter time.

The subcritical processing pressure of the placenta in the placenta extract production method of the present invention is equal to or higher than the saturated vapor pressure of the subcritical processing temperature (for example, 0.61 MPa or higher at 160 ° C., 1. It is preferable to carry out at 55 MPa or more. By using this pressure, it tends to be easy to produce a low molecular weight peptide having a molecular weight of 3000 or less. Although the upper limit of the subcritical processing pressure is not particularly defined, it is preferably suppressed to around 20 to 30 MPa due to the specifications of the high pressure apparatus.

The placenta subcritical treatment time in the placenta extract production method of the present invention is preferably 5 to 30 minutes. By making it into the range of this processing time, there exists a tendency which becomes easy to produce | generate a low molecular peptide. When the subcritical processing time is less than 5 minutes, it tends to be difficult to produce a low molecular peptide. When the subcritical processing time exceeds 30 minutes, the produced low molecular weight peptide is further excessively decomposed, and the amount of free amino acid produced tends to increase and the amount of low molecular weight peptide produced tends to decrease.

Furthermore, it is more preferable that the subcritical processing time of the placenta in the method for producing the placenta extract of the present invention is 10 to 30 minutes. If it is this range, it will become easy to produce | generate a low molecular peptide more efficiently.

In the production method of the placenta extract of the present invention, the hydrolysis conditions by the subcritical treatment of the placenta when water is used as the subcritical treatment extractant, the treatment temperature is 160 to 200 ° C., and the treatment pressure is saturated steam at the treatment temperature. The treatment time is preferably 5 to 30 minutes or more. By carrying out under these conditions, a low molecular peptide can be produced with high efficiency. Furthermore, the treatment temperature is 180 to 195 ° C., the treatment pressure is equal to or higher than the saturated vapor pressure of the treatment temperature, and the treatment time is 10 to 30 minutes, whereby a low molecular peptide having a molecular weight of 3000 or less can be obtained with the maximum efficiency. This is preferable.

The solid-liquid separation step in the method for producing the placenta extract of the present invention is a step of separating the subcritical processed product into the extract and the raw material residue (solid material). Specific solid-liquid separation processes include filtration using filter paper, centrifugation, decantation, screw press, roller press, rotary drum screen, belt screen, vibrating screen, multi-plate vibrating filter, vacuum dehydration, pressure dehydration, Belt press, centrifugal concentration dehydration, multiple disk dehydration and the like can be mentioned. Among these, filtration is preferable because the operation is simple and the separation efficiency is excellent.

The extract obtained by the solid-liquid separation step in the method for producing the placenta extract of the present invention can be transferred to the next decolorization step as it is, and can be decolorized, but is stored after the drying step of drying the extract. It may be solid with good properties. As this drying step, a general drying method can be used. Naturally, it is allowed to stand naturally, heat transfer drying such as box-type drying or spray drying which is a heating system, internal heat drying such as microwave drying, non-drying, etc. A heating system such as freeze drying, vacuum drying, suction drying, pressure drying, ultrasonic drying, and the like are possible. Of course, it is acceptable to dry using a general and simple oven and thermostat. Since the obtained solid matter is soluble in water, when it is desired to decolorize it, it can be dissolved again in water and transferred to the decolorization step.

The decolorization step in the method for producing the placenta extract of the present invention is the browning of the extract obtained in the solid-liquid separation step (including the extract obtained by dissolving the solid obtained in the drying step in water, the same applies hereinafter). Is a process of removing the color and making the color lighter.

In general, when a protein, which is the main tissue of the placenta, is reduced in molecular weight by enzymatic treatment or high-temperature treatment, the amino group of the protein reacts with the sugar contained in the raw material (so-called Maillard reaction) to form a substance called melanoidin And the protein browns to produce a blackish color. This reaction is not limited to placenta, and this reaction is common for components containing protein, for example, charring of fish and meat, coloring of coffee and miso.

In the method for producing the placenta extract of the present invention, examples of the decolorization step for removing the browning of the extract include a method using an adsorbent such as activated carbon or ion exchange resin. Especially, the activated carbon treatment which adsorb | sucks the produced | generated melanoidin to the fine hole of activated carbon and decolorizes from the reason that it can decolorize simply and efficiently is preferable.

In the method for producing a placenta extract of the present invention, the decolorization step by the activated carbon treatment is performed by adding a predetermined amount of activated carbon to the extract obtained in the solid-liquid separation step and stirring the mixture for a predetermined time. And solid-liquid separation.

The treatment temperature of the activated carbon treatment in the method for producing the placenta extract of the present invention is preferably 0 to 80 ° C, more preferably 10 to 60 ° C. When the processing temperature is less than 0 ° C., the extract may be frozen, and even when it is not frozen, the amount of adsorption tends to decrease. On the other hand, when the treatment temperature exceeds 80 ° C., the solid content concentration changes due to evaporation of the liquid, and there is a tendency that efficient decolorization reproduction cannot be obtained.

The treatment time of the activated carbon treatment in the method for producing the placenta extract of the present invention is preferably 3 minutes or more, and more preferably 5 minutes or more. When the treatment time is less than 3 minutes, the adsorption does not proceed as expected and decolorization tends to be insufficient. Moreover, the processing time of the decoloring process is preferably within 120 minutes, and more preferably within 60 minutes. When the processing time of the decolorization process exceeds 120 minutes, the adsorption reaction reaches equilibrium, and there is a tendency that the efficiency of decolorization does not change and there is no change in the decolorization effect. When the processing time of the decoloring process is less than 3 minutes, the browning of the extract cannot be removed.

In the method for producing the placenta extract of the present invention, the solid-liquid separation method in the decolorization step by the activated carbon treatment is not particularly limited, and various methods mentioned as the solid-liquid separation step can be adopted, but the operation is simple, and the separation efficiency. Filtration and decantation are preferred because of their excellent performance.

In the method for producing the placenta extract of the present invention, the activated carbon used for the activated carbon treatment is not particularly limited, and a commercially available activated carbon is appropriately selected and used depending on the properties of melanoidin to be produced and availability. Can do. As activated carbon for melanoidin adsorption, commercially available melanoidin or activated carbon for caramel adsorption can be used, and the production method is to control the pore size by general chemical (zinc) or steam activation treatment etc. Can be used. For example, the Shirataka series and the carborafin series manufactured by Nippon Enviro Chemicals Co., Ltd. can be mentioned.

The amount of the activated carbon used in the method for producing the placenta extract of the present invention can be adjusted as appropriate according to the amount of melanoidin produced (that is, depending on the color intensity), but 100 mass of the liquid to be decolored. The amount is preferably 0.1 to 30 parts by mass with respect to parts. When the amount is less than 0.1 parts by mass, the decolorization performance tends to be insufficient, and the effect of lowering the chromaticity tends to be insufficient. When the amount exceeds 30 parts by mass, the decolorization performance is improved, but yield loss occurs due to the penetration of the liquid component simultaneously with decolorization, and there is a tendency that a sufficient amount cannot be secured in the end. For the reason that the decolorization operation can be performed more efficiently, it is more preferable to add 1 to 10 parts by mass of the activated carbon.

In addition, in the method for producing the placenta extract of the present invention, the activated carbon treatment can be repeated a plurality of times with a small amount of activated carbon used instead of adjusting the amount of activated carbon. Furthermore, by performing the activated carbon treatment a plurality of times, the decolorization behavior / process is revealed in more detail, and the minimum necessary amount of activated carbon can be used. The number of repetitions can be appropriately selected according to the amount of melanoidin (that is, depending on the color intensity), but is preferably about 2 to 10 times and more preferably about 2 to 5 times in view of the complexity of the process. By performing the treatment a plurality of times, the brown color of the extract can be further removed.

In the placenta extract production method of the present invention, the extractant is water, the subcritical processing temperature is 190 to 195 ° C., the subcritical processing pressure is equal to or higher than the saturated vapor pressure of the subcritical processing temperature, and the subcritical processing time is 20 to It is desirable that the placenta be subcritically treated for 30 minutes to obtain a placenta subcritically treated product, and then decolorized by 2-5 times of activated carbon treatment. The placenta extract obtained by the production method under the above conditions can have an absorbance at a wavelength of 390 nm in the range of 0.028 to 0.084, and the nitrogen content of the placenta extract can be 0.15 to 0.21. It can be in the range of mass%.

In the conventional extraction method using an enzyme treatment, it is natural to add an enzyme, but it is necessary to use other auxiliary agents. Specifically, in addition to enzymes, enzyme reaction aids such as pH adjusters and enzyme stabilizers are added to the placenta raw material, and the inclusion of these additives dilutes the concentration of the active ingredient, resulting in an extract. The active ingredient concentration per unit becomes low.

On the other hand, placenta extract obtained by subcritical processing uses subcritical water extraction as a means of placenta extraction. Basically, only water is used to reduce the molecular weight of proteins and peptides. It is possible not to use any other additives. Therefore, the placenta extract obtained by using this extraction method basically has a high peptide concentration with a low molecular weight, and an extract having a higher active ingredient concentration than the prior art can be obtained. Therefore, if the placenta extract obtained by the conventional technique and the subcritical treatment is subjected to the same decolorization operation by the activated carbon treatment, the adsorbed amount of main components other than melanoidine that is necessarily removed by the activated carbon treatment, that is, the peptide amount is not changed. As a result, the amount of peptide in the solution after decolorization obtained is larger in the present invention. That is, the concentration of the active ingredient can be kept high even after decolorization.

The protein constituting the placenta is composed mainly of various amino acids, that is, about 20 kinds of amino acids constituting the living body, and naturally there are various types of peptide bonds, whereas In the enzyme treatment of the technology, only a low molecular weight is obtained by cleaving a specific peptide bond due to the substrate specificity of the enzyme. That is, since only a specific peptide bond can be cleaved by the method using enzyme treatment, the degree of molecular weight reduction is limited, and a molecular weight below a certain level cannot be reached.

In the method for producing the placenta extract of the present invention, in the extraction by subcritical water treatment, the peptide bond is cleaved by the hydrolytic power of water under high temperature and high pressure. By cleaving, the protein can be reduced in molecular weight. Therefore, it is possible to obtain a molecular weight distribution with more low molecular weight peptides than conventional enzyme treatment depending on temperature and time conditions.

Furthermore, the size of the low-molecular peptide obtained by the enzymatic treatment method matches the size of the pore size of the activated carbon for melanoidin adsorption, and the low-molecular peptide is also adsorbed along with the adsorption of melanoidin. It is presumed that the amount of peptide is reduced. On the other hand, the size of low molecular peptides produced by extraction by subcritical water treatment exists as lower molecular peptides than the conventional method as described above. Adsorption of molecular peptides is unlikely to occur, and as a result, it is possible to obtain a decolored extract despite the large amount of low molecular peptides that are active ingredients.

In addition, the placenta extract of the present invention thus obtained may be diluted or concentrated in a timely manner according to the purpose of use. Dilution is simple addition of water, addition of organic solvents such as ethanol, other liquids and food additives containing active ingredients, fragrances, phenoxyethanol and butanediol, glycerin, antiseptics for solids, or organic solvents in which they are dissolved And natural preservatives can be added. Although the dilution increases the transparency, the amount of nitrogen decreases, but the dilution can be diluted while keeping the amount of nitrogen higher than that of the conventional extract. Concentration can be performed by a general concentration method such as heat concentration or reduced pressure concentration. After drying by the above-described drying method, an appropriate amount of a solvent such as water can be added and dissolved. Although the concentrate increases the nitrogen content, the transparency decreases, but the concentration can be performed while keeping the transparency higher than that of the conventional extract.

The present invention will be described based on examples, but the present invention is not limited to the examples.

Example 1
<Extraction process>
200 g of pig placenta (Tokyo Shibaura Organ Co., Ltd.) and 200 g of distilled water were placed in a pressure-resistant container having a volume of 2 L, and subcritical processing was performed at a processing temperature of 195 ° C., a processing pressure of 1.6 MPa, and a processing time of 30 minutes. The pig placenta was washed with water and blood and other parts were removed.
<Solid-liquid separation process>
After finishing the subcritical treatment, the treated product in the pressure vessel was collected and suction filtered with a cellulose filter paper (pore size: 1 μm, 5C manufactured by Advantec), and the filtrate was collected. The collected filtrate was brown.
<Decolorization process (activated carbon treatment) 1st time>
50 g of the obtained filtrate was weighed into a beaker and stirred using a stirrer in a constant temperature water bath at 30 ° C. Separately, 3 parts by mass of activated carbon (using commercially available activated melanoidin activated carbon) for 100 parts by mass of the filtrate was added to the beaker in which the filtrate was being stirred. Further, after stirring for 30 minutes, the beaker was taken out from the thermostatic bath and suction filtered with the same filter paper as in the solid-liquid separation step, and the filtrate was recovered. The collected filtrate was a little dark yellow.
<Decolorization process (activated carbon treatment) second time>
The same decolorization process was performed on the filtrate obtained in the first decolorization step, and the filtrate was collected again. The color of the resulting extract was visually colorless.

Example 2
The same decolorization process was performed on the extract obtained in Example 1, and the filtrate was collected again. The color of the resulting extract was visually colorless.

Comparative Examples 1 to 4
Commercially available placenta lotions 1 to 4 (all derived from porcine placenta, extraction process: enzyme treatment, decolorization process: activated carbon treatment) were designated as Comparative Examples 1 to 4. The apparent colors of each of Comparative Examples 1 and 2 were yellow, and Comparative Examples 3 and 4 were colorless.

Comparative Example 5
<Extraction process>
To 200 g of pig placenta (Tokyo Shibaura Organ Co., Ltd.), 50 g of distilled water, 4 mL of protein hydrolase Alcalase (manufactured by Novozymes) and 2 mL of 25 mass% sodium hydroxide were added and reacted at 60 ° C. for 3 hours. Thereafter, the enzyme was inactivated by treatment at 90 ° C. for 1 hour. In order to substantially match the solid content concentration with Examples 1 and 2, distilled water was added after the deactivation treatment to make the total amount 400 g.
<Solid-liquid separation process>
The reaction liquid after enzyme deactivation was filtered through cellulose filter paper (pore diameter: 1 μm, 5C manufactured by Advantec), and the filtrate was recovered. The collected filtrate was dark brown <Decolorization step (activated carbon treatment) 1st time>
50 g of the obtained filtrate was weighed into a beaker and stirred using a stirrer in a constant temperature water bath at 30 ° C. Separately, 3 parts by mass of activated carbon (commercial activated carbon for melanoidin adsorption) was added to 100 parts by mass of the filtrate, and the filtrate was added to the beaker in which the filtrate was being stirred. Further, after stirring for 30 minutes, the beaker was taken out from the thermostatic bath and suction filtered with the same filter paper as in the solid-liquid separation step, and the filtrate was recovered. The collected filtrate was a little dark yellow.
<Decolorization process (activated carbon treatment) second time>
The same decolorization process was performed on the filtrate obtained in the first decolorization step, and the filtrate was collected again. The color of the obtained extract was light yellow.
<Decolorization process (activated carbon treatment) third time>
The same decolorization process was performed on the filtrate obtained in the first decolorization step, and the filtrate was collected again. The color of the resulting extract was apparently colorless.

Absorbance measurement Absorbance at 390 nm of the extract solutions and skin lotions of Examples 1 and 2 and Comparative Examples 1 to 5 was measured using a visible ultraviolet spectrophotometer (GeneQuant 1300 manufactured by GE Healthcare Japan Co., Ltd.). The results are shown in Table 1 and FIG.

Nitrogen content measurement The nitrogen content in the extracts or lotions of Examples 1 and 2 and Comparative Examples 1 to 5 was determined using the Kjeldahl method (sulfuric acid was added to the test liquid, and the organic nitrogen in the liquid was converted to ammonium sulfate to obtain a liquid. Was made alkaline and heated, and the generated ammonia was determined by titration. Specifically, it was performed in accordance with the method described in JIS K0102244. The results are shown in Table 1 and FIG.

Molecular Weight Distribution Measurement The extracts or lotions of Examples 1-2 and Comparative Examples 1-5 were filtered through a 0.45 μm membrane filter and measured by high performance liquid chromatography (HP1100 series manufactured by Agilent Technologies).

For analysis conditions, a column (product number: TSK guard column SWXL (6.0 mm ID × 40 mm) and TSKgel G2000SWXL (7.8 mm ID × 300 mm) manufactured by Tosoh Corporation) was used, and the eluent was 0.1 w. / V% TFA in MeCN / H ₂ O = 45/55, column temperature 35 ° C., flow rate 1.0 mL / min, detection UV 220 nm, introduction amount 20 μL, analysis time 30 min. Cytochrome C (standard reagent for molecular weight distribution measurement) Wako Pure Chemical Industries, Ltd.) Molecular weight 12,500, Aprotinin (CALBOCHEM) molecular weight 6512, Bacitracin (Dr. Ehrentorfer) molecular weight 1,423, Angiotensin II (Peptide Institute) Molecular weight 1,026, Gly-Gly-Tyr-Arg (peptide) Laboratory) Create a calibration curve with the same measurement based on molecular weight 451 and Gly-Gly-Gly (Peptide Institute) molecular weight 189, determine the molecular weight position relative to the elution time, find the molecular weight distribution, and the peak area in the molecular weight fractionation range The molecular weight ratio was determined and the results are shown in Table 1.

In addition, the molecular weight obtained this time is not more than the extra exclusion molecular weight (about 100,000) of the column used for the upper limit in all samples, and the maximum molecular weight calculated from the time when elution starts is about 20,000. there were. Therefore, it was found that all the proteins contained in the raw materials in the above Examples and Comparative Examples were converted into peptide forms having a molecular weight of about 100,000 or less, and further about 20,000 or less and contained in the sample. .

From the results in Table 1, it can be seen that the placenta extract of the present invention is a placenta extract having a high concentration of active ingredients and little coloring. Furthermore, it can be seen that the placenta extract produced by the production method including the subcritical process described above is a placenta extract having a high concentration of active ingredients and little coloring. Moreover, in the comparative example 5, the smell which feels a raw odor was confirmed. However, in Examples 1 and 2, no odor felt as a raw odor was confirmed.

The molecular weight distribution (area) value for Comparative Examples 1 to 4 is very large. This is because Comparative Examples 1 to 4, which are commercially available placenta lotions, contain other ingredients (preservatives, This is presumed to be due to containing a large amount of (including the addition of an ultraviolet absorber, etc.).

Claims

Absorbance at a wavelength of 390 nm is 0.10 or less,
A placenta extract having a nitrogen content of 0.10% by mass or more.
The placenta extract according to claim 1, wherein the absorbance is 0.01 to 0.10.
The placenta extract according to claim 1 or 2, wherein the nitrogen content is 0.10 to 1.0 mass%.
The placenta extract according to any one of claims 1 to 3, wherein the content of all peptides in the total solid content of the placenta extract is 70 to 99.5% by mass.
The placenta extract according to any one of claims 1 to 4, wherein the content of the low molecular weight peptide having a molecular weight of 3000 or less in the total solid content of the placenta extract is 40 to 99.5% by mass.
6. The placenta extract according to claim 1, wherein the content of free amino acids in the total solid content of the placenta extract is 0.01 to 30% by mass.
The placenta extract according to any one of claims 1 to 6, wherein the content of a component having a molecular weight of 3000 or less in the total solid content of the placenta extract is 100%.
The placenta extract according to any one of claims 1 to 7, wherein the placenta extract is a placenta extract extracted through a subcritical treatment of a placenta raw material.
The placenta extract according to any one of claims 1 to 8, wherein the placenta extract is a placenta extract extracted through an activated carbon treatment.
The temperature of subcritical processing is 160-200 ° C,
The placenta extract according to claim 8, wherein the pressure is equal to or higher than a saturated vapor pressure.
The placenta extract according to claim 8 or 10, wherein the subcritical processing time is 5 to 30 minutes.
Absorbance at a wavelength of 390 nm is 0.10 or less,
A placenta extract production method for obtaining a placenta extract having a nitrogen content of 0.10% by mass or more,
An extraction process for subcritical processing of the placenta to obtain a subcritical processed product,
A method for producing a placenta extract, comprising a solid-liquid separation step of separating the subcritical processed product into an extract and a raw material residue.
The method for producing a placenta extract according to claim 12, further comprising a decolorization step after the extraction step by the subcritical treatment.
The method for producing a placenta extract according to claim 12 or 13, wherein a temperature of the subcritical treatment in the subcritical treatment is 160 to 200 ° C, and a pressure is equal to or higher than a saturated vapor pressure.
The method for producing a placenta extract according to any one of claims 12 to 14, wherein the subcritical treatment time in the subcritical treatment is 5 to 30 minutes.
The method for producing a placenta extract according to claim 13, wherein the treatment temperature in the decolorization step is 0 to 80 ° C.
The method for producing a placenta extract according to claim 13 or 16, wherein the treatment time in the decolorization step is 3 to 120 minutes.
The method for producing a placenta extract according to claim 13, 16 or 17, wherein the number of treatments in the decoloring step is 2 to 10 times.