JP2006158354A - Method for extracting eggshell membrane hydrolysis protein containing collagen from eggshell membrane using ultrasonic wave - Google Patents
Method for extracting eggshell membrane hydrolysis protein containing collagen from eggshell membrane using ultrasonic wave Download PDFInfo
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- JP2006158354A JP2006158354A JP2004358460A JP2004358460A JP2006158354A JP 2006158354 A JP2006158354 A JP 2006158354A JP 2004358460 A JP2004358460 A JP 2004358460A JP 2004358460 A JP2004358460 A JP 2004358460A JP 2006158354 A JP2006158354 A JP 2006158354A
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- eggshell membrane
- eggshell
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- hydrolysis
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- 229940012466 egg shell membrane Drugs 0.000 title claims abstract description 47
- 238000000034 method Methods 0.000 title claims abstract description 15
- 102000008186 Collagen Human genes 0.000 title claims abstract description 14
- 108010035532 Collagen Proteins 0.000 title claims abstract description 14
- 229920001436 collagen Polymers 0.000 title claims abstract description 14
- 102000004169 proteins and genes Human genes 0.000 title claims abstract description 11
- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 11
- 230000007062 hydrolysis Effects 0.000 title abstract 7
- 238000006460 hydrolysis reaction Methods 0.000 title abstract 7
- 230000001678 irradiating effect Effects 0.000 claims abstract description 4
- 238000002137 ultrasound extraction Methods 0.000 claims 1
- 210000003278 egg shell Anatomy 0.000 abstract description 21
- 102000002322 Egg Proteins Human genes 0.000 abstract description 18
- 108010000912 Egg Proteins Proteins 0.000 abstract description 18
- 239000002253 acid Substances 0.000 abstract description 5
- 239000003513 alkali Substances 0.000 abstract description 3
- 238000000354 decomposition reaction Methods 0.000 abstract description 3
- 238000007796 conventional method Methods 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 8
- 239000007857 degradation product Substances 0.000 description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 6
- 239000000047 product Substances 0.000 description 5
- 235000013601 eggs Nutrition 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 230000003301 hydrolyzing effect Effects 0.000 description 3
- 239000012528 membrane Substances 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 230000011382 collagen catabolic process Effects 0.000 description 2
- 238000001962 electrophoresis Methods 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- 238000002604 ultrasonography Methods 0.000 description 2
- 102000029816 Collagenase Human genes 0.000 description 1
- 108060005980 Collagenase Proteins 0.000 description 1
- 102100033962 GTP-binding protein RAD Human genes 0.000 description 1
- 108050007570 GTP-binding protein Rad Proteins 0.000 description 1
- 108010009736 Protein Hydrolysates Proteins 0.000 description 1
- 239000007983 Tris buffer Substances 0.000 description 1
- 206010052428 Wound Diseases 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 239000003929 acidic solution Substances 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 210000000991 chicken egg Anatomy 0.000 description 1
- 229960002424 collagenase Drugs 0.000 description 1
- 238000004040 coloring Methods 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 238000010612 desalination reaction Methods 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000000909 electrodialysis Methods 0.000 description 1
- 230000003628 erosive effect Effects 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000000796 flavoring agent Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 238000010979 pH adjustment Methods 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 239000003531 protein hydrolysate Substances 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
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- Cosmetics (AREA)
Abstract
Description
この発明は、超音波を用いた卵殻膜からのコラーゲンを含む卵殻膜加水分解タンパク質の抽出方法に関する。 The present invention relates to a method for extracting eggshell membrane hydrolyzed protein containing collagen from eggshell membranes using ultrasound.
卵殻膜は、古くから創傷の治療などに利用されてきた。この卵殻膜をいろんなものに添加して使用し易くするため、卵殻膜を強酸や強アルカリの溶液に入れ、加熱し還流状態で卵殻膜を加水分解して卵殻膜分解物を得る方法がある（例えば、特許文献１）。 Egg shell membranes have long been used for wound treatment and the like. In order to add this eggshell membrane to various things for easy use, there is a method in which the eggshell membrane is placed in a strong acid or strong alkali solution and heated to hydrolyze the eggshell membrane under reflux to obtain an eggshell membrane degradation product ( For example, Patent Document 1).
さらに、異臭、着色がない卵殻膜分解物を作るため、アルカリ性の含水有機溶媒中で卵殻膜を加水分解して卵殻膜分解物を得る方法（特開平１−２７５５１２）がある（例えば、特許文献２）。 Furthermore, there is a method (JP-A-1-275512) for obtaining an eggshell membrane decomposition product by hydrolyzing the eggshell membrane in an alkaline water-containing organic solvent in order to produce an eggshell membrane decomposition product free from off-flavors and coloring. 2).
また、酢酸な等の弱酸性の溶液に浸漬することにより卵殻膜からコラーゲンを含むタンパク質が溶液中に抽出されることが知られている。
これまでの方法は卵殻膜を加水分解する前に、あらかじめ卵殻と卵殻膜を分離しておく必要があった。卵殻膜と卵殻の分離には、破砕した卵殻を水中に入れて撹拌や超音波照射によって剥離する方法や、卵殻を酸性水溶液に浸漬して卵殻のみを溶かす方法があるが、機械的な分離では分離の効率が悪い。超音波照射による分離では超音波発生のために大量のエネルギーを必要とする。さらに、卵殻の溶解は酸に浸漬する必要があり、長時間を要する。いずれの方法でも経済的に効率が悪いと考えられる。 Previous methods required the eggshell and eggshell membrane to be separated in advance before the eggshell membrane was hydrolyzed. There are two methods for separating eggshell membranes and eggshells: a method in which crushed eggshell is placed in water and separated by stirring or ultrasonic irradiation, and a method in which eggshell is immersed in an acidic aqueous solution to dissolve only eggshell. Separation efficiency is poor. Separation by ultrasonic irradiation requires a large amount of energy to generate ultrasonic waves. Furthermore, the eggshell needs to be dissolved in an acid, which takes a long time. Either method is considered to be economically inefficient.
卵殻膜を酸やアルカリで加水分解するとことによって得られる卵殻膜加水分解物は、卵殻膜のタンパク質が分解されすぎてしまって、コラーゲン、コラーゲン分解物のみを分離精製することは困難である。 In the eggshell membrane hydrolyzate obtained by hydrolyzing the eggshell membrane with acid or alkali, the protein of the eggshell membrane is excessively degraded, and it is difficult to separate and purify only collagen and collagen degradation products.
化学物質により卵殻膜を加水分解すると未溶解の卵殻膜も化学物質の浸食を受けて大きく変性してしまう。 When the eggshell membrane is hydrolyzed by a chemical substance, the undissolved eggshell membrane is also greatly denatured by the erosion of the chemical substance.
発明者は、上記の目的を達しようと鋭意研究した結果、本発明を完成した。卵殻膜に超音波を照射することによってタンパク質加水分解物を抽出することを特徴とするものである。 As a result of intensive research aimed at achieving the above object, the inventors completed the present invention. The protein hydrolyzate is extracted by irradiating the eggshell membrane with ultrasonic waves.
以下、本発明を詳細に説明する。本発明でいう卵殻膜の加水分解物とは、鳥卵の卵殻の内側にある卵殻膜を加水分解したものをいう。卵殻膜の代表的な例として鶏卵の卵殻膜が挙げられる。 Hereinafter, the present invention will be described in detail. The hydrolyzate of eggshell membrane as used in the present invention refers to a product obtained by hydrolyzing the eggshell membrane inside the eggshell of a bird egg. A typical example of the eggshell membrane is a eggshell membrane of a chicken egg.
本発明の卵殻膜分解物は、化粧品、医薬、食品などの原料として使用することができる。 The eggshell membrane degradation product of the present invention can be used as a raw material for cosmetics, medicines, foods and the like.
以下、本発明の卵殻膜分解物の代表的製造方法について述べる。まず、殻付鶏卵を割卵して卵液を除去して卵殻を得、この卵殻を水洗した後、破砕してから水中に入れ超音波を照射することにより、卵殻膜加水分解物を抽出し、卵殻膜加水分解物含有溶液をえる。 Hereinafter, representative methods for producing the eggshell membrane degradation product of the present invention will be described. First, egg shell membrane hydrolyzate is extracted by breaking eggs in shells and removing egg liquid to obtain eggshells, washing the eggshells with water, crushing them, placing them in water, and irradiating them with ultrasonic waves. Obtain a solution containing eggshell membrane hydrolyzate.
このようにして得られた卵殻膜の加水分解物は、分解物が溶液中に存在したままとしてもよいが、通常溶解していない成分と分離するためのフィルター濾過遠心分離、酸剤を添加してのｐＨ調整（中和処理）、ミクロフィルターによる除菌処理、電気透析などの脱塩処理などを行って精製する。さらに、必要に応じて乾燥して製品とする。 The hydrolyzate of eggshell membrane thus obtained may remain in the solution, but it is usually filtered by filtration to separate it from undissolved components, and an acid agent is added. Purification is performed by pH adjustment (neutralization treatment), sterilization treatment using a microfilter, and desalination treatment such as electrodialysis. Further, the product is dried as necessary.
本発明によれば、超音波によって抽出される卵殻膜加水分解物中にはコラーゲン、コラーゲン分解物が含まれており、またこれをゲル化することによりコラーゲンを容易に分離精製することが可能である。 According to the present invention, the eggshell membrane hydrolyzate extracted by ultrasound contains collagen and collagen degradation product, and it is possible to easily separate and purify collagen by gelling it. is there.
以下、実施例により本発明の方法をさらに具体的に説明する。卵殻を割卵して卵液を除き、得られた卵殻を水洗してから、直径５ｍｍのメッシュを通過するように卵殻を粉砕した。この卵殻２２５０ｇと水２６３０ｍＬを混合し１３００Ｗ、１５ＫＨｚ、４Ａの超音波を、４分間照射し、卵殻膜加水分解物含有溶液を得た。 Hereinafter, the method of the present invention will be described more specifically with reference to examples. The eggshell was broken to remove the egg liquid, and the resulting eggshell was washed with water, and then the eggshell was crushed so as to pass through a mesh having a diameter of 5 mm. 2250 g of this eggshell and 2630 mL of water were mixed and irradiated with ultrasonic waves of 1300 W, 15 KHz, 4 A for 4 minutes to obtain an eggshell membrane hydrolyzate-containing solution.
試験例＜分解物の組成の調査＞本発明により得られた、卵殻膜加水分解物含有溶液中にコラーゲンが含まれていることを確認するために、実施例で得られた卵殻膜加水分解物含有溶液（本発明品）を使用し、下記の条件で電気泳動（SDS-PAGE）を行った。コラゲナーゼタイプＩ（和光純薬）0.01mgをバッファー（Tris ５００ｍM, NaCl ２M, CaCl ２１mM, NaN3 0.2%）0.1mLに溶解し、卵殻膜加水分解物含有溶液中1mLと混合し、３７℃で一晩反応させ、電気泳動を行いコラーゲンが分解されていることを確認した。その結果は図１に示すとおりである。
その結果は図１に示すとおりである。タンパク質の分子量マーカーはBIO RAD protein standard IIを用いた、分子量は高分子側からそれぞれ97400, 66200, 55000, 42700, 40000 kDaである。
Test Example <Investigation of Composition of Degradation Product> In order to confirm that collagen is contained in the eggshell membrane hydrolyzate-containing solution obtained by the present invention, the eggshell membrane hydrolyzate obtained in the examples Using the containing solution (product of the present invention), electrophoresis (SDS-PAGE) was performed under the following conditions. Collagenase type I (Wako Pure Chemical Industries) 0.01 mg is dissolved in 0.1 mL of buffer (Tris 500 mM, NaCl 2 M, CaCl 21 mM, NaN 3 0.2%), mixed with 1 mL in a solution containing eggshell membrane hydrolysate, and mixed at 37 ° C. The reaction was performed overnight and electrophoresis was performed to confirm that the collagen was decomposed. The result is as shown in FIG.
The result is as shown in FIG. The molecular weight marker of the protein was BIO RAD protein standard II, and the molecular weights were 97400, 66200, 55000, 42700, and 40000 kDa from the polymer side, respectively.
Claims (1)
Extracting the hydrolyzed protein containing collagen from the eggshell membrane by irradiating the eggshell membrane with ultrasonic waves in a solution, of the eggshell membrane hydrolyzed protein containing collagen from the eggshell membrane using ultrasound Extraction method.
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| JP2004358460A JP2006158354A (en) | 2004-12-10 | 2004-12-10 | Method for extracting eggshell membrane hydrolysis protein containing collagen from eggshell membrane using ultrasonic wave |
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| JP2004358460A JP2006158354A (en) | 2004-12-10 | 2004-12-10 | Method for extracting eggshell membrane hydrolysis protein containing collagen from eggshell membrane using ultrasonic wave |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100029564A1 (en) * | 2007-10-17 | 2010-02-04 | Biova, L.L.C. | Novel process of solubilizing protein from a proteinaceous material and compositions thereof |
| JP2011184269A (en) * | 2010-03-10 | 2011-09-22 | Wada Kigata Seisakusho:Kk | Method for removing protein from calcareous material of biological origin and resin composition containing solid calcium carbonate obtained by the method |
| CN102302083A (en) * | 2011-08-08 | 2012-01-04 | 中华全国供销合作总社杭州茶叶研究院 | Method for extracting tea protein product |
| US8173174B2 (en) * | 2007-10-17 | 2012-05-08 | Biova, L.L.C. | Solubilized protein composition obtained from avian eggshell membrane |
| CN103275205A (en) * | 2013-05-08 | 2013-09-04 | 江苏福旦生物医药有限公司 | Water-soluble egg membrane bioactive peptide production method |
| US20140348939A1 (en) * | 2013-05-24 | 2014-11-27 | Robert C. Blaine | Wound care product with egg shell membrane |
| CN107043798A (en) * | 2016-03-02 | 2017-08-15 | 世界家庭用具制品厂有限公司 | Eggshell recovery method and application of recovered material |
| JP2017171598A (en) * | 2016-03-23 | 2017-09-28 | 富士フイルム株式会社 | Endo180 enhancers and cosmetics |
-
2004
- 2004-12-10 JP JP2004358460A patent/JP2006158354A/en active Pending
Cited By (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US8425943B2 (en) | 2007-10-17 | 2013-04-23 | Biova, L.L.C. | Method of treatment using solubilized protein composition obtained from eggshell membrane |
| US20100029564A1 (en) * | 2007-10-17 | 2010-02-04 | Biova, L.L.C. | Novel process of solubilizing protein from a proteinaceous material and compositions thereof |
| US8173174B2 (en) * | 2007-10-17 | 2012-05-08 | Biova, L.L.C. | Solubilized protein composition obtained from avian eggshell membrane |
| US8197853B2 (en) * | 2007-10-17 | 2012-06-12 | Biova, L.L.C. | Process for solubilizing protein from a proteinaceous material and compositions thereof |
| US8211477B2 (en) * | 2007-10-17 | 2012-07-03 | Biova, L.L.C. | Solubilized protein composition obtained from eggshell membrane |
| JP2011184269A (en) * | 2010-03-10 | 2011-09-22 | Wada Kigata Seisakusho:Kk | Method for removing protein from calcareous material of biological origin and resin composition containing solid calcium carbonate obtained by the method |
| CN102302083A (en) * | 2011-08-08 | 2012-01-04 | 中华全国供销合作总社杭州茶叶研究院 | Method for extracting tea protein product |
| CN102302083B (en) * | 2011-08-08 | 2013-09-18 | 中华全国供销合作总社杭州茶叶研究院 | Method for extracting tea protein product |
| CN103275205B (en) * | 2013-05-08 | 2014-12-03 | 江苏福旦生物医药有限公司 | Water-soluble egg membrane bioactive peptide production method |
| CN103275205A (en) * | 2013-05-08 | 2013-09-04 | 江苏福旦生物医药有限公司 | Water-soluble egg membrane bioactive peptide production method |
| US20140348939A1 (en) * | 2013-05-24 | 2014-11-27 | Robert C. Blaine | Wound care product with egg shell membrane |
| US10166260B2 (en) * | 2013-05-24 | 2019-01-01 | Blaine Laboratories, Inc. | Wound care product with egg shell membrane |
| CN107043798A (en) * | 2016-03-02 | 2017-08-15 | 世界家庭用具制品厂有限公司 | Eggshell recovery method and application of recovered material |
| CN107043798B (en) * | 2016-03-02 | 2021-08-31 | 纳米及先进材料研发院有限公司 | A kind of eggshell recovery method and the use of the recovered product |
| JP2017171598A (en) * | 2016-03-23 | 2017-09-28 | 富士フイルム株式会社 | Endo180 enhancers and cosmetics |
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