JP7042544B2 - Method for preparing composition for hair loss treatment - Google Patents
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Description
本発明は、様々な組成物を調製するための方法に関し、日常の化学物質の分野に関する。 The present invention relates to methods for preparing various compositions and to the field of everyday chemicals.
生活の加速、仕事のプレッシャーの増加、食事や生活習慣の変化により、中国の若者と中年男性の脱毛の発生率は、20年前と比較して10倍以上増加している。同時に、女性居住者の脱毛の発生率は年々増加している。しかし、脱毛は病院に行く必要がある病気であるという認識率はまだ非常に低いである。主な理由は、遺伝的要因、生理学的機能障害、仕事と研究の圧力、精神的ストレス、薬物反応、放射線療法、環境汚染、栄養失調などである。 Due to accelerated lifestyles, increased work pressure, and changes in diet and lifestyle, the incidence of hair loss in young and middle-aged men in China has increased more than 10 times compared to 20 years ago. At the same time, the incidence of hair loss in female residents is increasing year by year. However, the recognition rate that hair loss is a disease that requires going to the hospital is still very low. The main reasons are genetic factors, physiological dysfunction, work and research pressure, psychological stress, drug reactions, radiation therapy, environmental pollution, malnutrition, etc.
しかし、最新の研究では、脱毛は体の腸内細菌叢の不均衡に関連していることが示されている。腸内細菌叢の障害は、乳酸菌(Lactobacillus rhamnosus)の増加を招き、次に胃腸管微生物の分泌にたよる可能性が最も高い特定の物質または酵素の欠如につながる。人間の腸内細菌叢は複雑なコミュニティである。健康な成人の腸管内の微生物の総量は約1~1.5kgである。1,000を超える種があり、腸内微生物の数は人間の細胞の数の10倍である。これは「見えない人体スーパーオルガン」と呼ばれる。腸内細菌叢は、消化、代謝、免疫機能に重要な役割を果たしている。腸内細菌叢の不均衡は、肥満、糖尿病、腫瘍、脱毛などのさまざまな代謝異常の重要な原因の1つである可能性がある。したがって、この特許は、本発明の組成物を経口摂取することによって、腸内細菌叢の多様性を補完し、腸内細菌叢を改善することを目的としている。 However, recent studies have shown that hair loss is associated with an imbalance in the body's gut flora. Disorders of the gut flora lead to an increase in lactic acid bacteria (Lactobacillus rhamnosus), which in turn leads to the lack of certain substances or enzymes most likely due to the secretion of gastrointestinal microbes. The human gut flora is a complex community. The total amount of microorganisms in the intestinal tract of a healthy adult is about 1-1.5 kg. There are more than 1,000 species, and the number of gut microbiota is ten times the number of human cells. This is called the "invisible human body super organ". The gut microbiota plays an important role in digestion, metabolism and immune function. Gut microbiota imbalance may be one of the important causes of various metabolic disorders such as obesity, diabetes, tumors and hair loss. Therefore, this patent aims to complement the diversity of the intestinal flora and improve the intestinal flora by oral ingestion of the compositions of the present invention.
本発明の目的は、腸内細菌叢を改善し、それによりヒト脱毛症の技術的問題を解決し、脱毛治療用組成物の調製方法を提供することである。 It is an object of the present invention to improve the intestinal bacterial flora, thereby solving the technical problems of human alopecia, and to provide a method for preparing a composition for treating alopecia.
第1の脱毛治療用組成物の調製方法は、以下の工程に従って行われる。 The method for preparing the first composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒木耳(黒きくらげ、black fungus)を洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、その後、超音波パワーが500Wで超音波を15分間をかけ、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。分子量カットオフ(Molecular weight cutoff)が300kDaのきくらげ多糖(Auricularia polysaccharide)保持液を得るために、温度40℃、圧力0.17barの条件で限外濾過する。
1. 1. Membrane separation of wood ear polysaccharides Black ear (black fungus) is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of black fungus. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, then ultrasonic power is 500 W, ultrasonic waves are applied for 15 minutes, extraction is performed in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifuge for a minute to remove the residue, and prepare the extract into a feed solution with a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. In order to obtain an Auricularia polysaccharide holding solution having a molecular weight cutoff of 300 kDa, filtration is performed under the conditions of a temperature of 40 ° C. and a pressure of 0.17 bar.
2.完全な粒子と均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、30℃で3時間浸し、十分に加熱調理し、調理したものを30℃~40℃に冷却する。300kDaのきくらげ多糖保持(retentate)液とアスペルギルス種菌液を1:3の質量比で接種し、室温28℃に保ち、麹のかき混ぜの1回目は20~24時間、2回目は28時間になるときに行われた。34時間後に麹ができる。麹ができたら麹を洗い、12時間密閉してからタンクに入れ、28℃の恒温室に入れて30日以上発酵させ、取り出して乾燥させて麹菌型―きくらげ多糖300kDaの保持液テンペ発酵物を得る。 2. 2. Select perfect particles and uniform size soybeans, weigh and wash, mix with water at a volume ratio of 1: 3, soak at 30 ° C for 3 hours, cook well, cook and cook at 30 ° C. Cool to ~ 40 ° C. Inoculate 300 kDa Kikurage polysaccharide retention solution and Aspergillus inoculum solution at a mass ratio of 1: 3, keep the room temperature at 28 ° C, and stir the koji for 20 to 24 hours for the first time and 28 hours for the second time. Was done in. Jiuqu is formed after 34 hours. When the jiuqu is made, wash the jiuqu, seal it for 12 hours, put it in a tank, put it in a constant temperature room at 28 ° C to ferment it for 30 days or more, take it out and dry it. obtain.
第2の脱毛治療用組成物の調製方法は、以下の工程に従って行われる。 The method for preparing the second composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、その後、超音波パワーが500Wである超音波を、15分間かける。ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。分子量カットオフが300kDaのきくらげ多糖保持液を得るために、温度40℃、圧力0.17barで限外濾過する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. The dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and then ultrasonic waves having an ultrasonic power of 500 W are applied for 15 minutes. Extraction is performed in a water bath for 3 hours, and the extract is centrifuged at 4500 r / min for 10 minutes to remove the residue, and the extract is prepared as a feed solution having a black syrup polysaccharide concentration of 4 g / L and a pH value of 5.6. Ultrafiltration is performed at a temperature of 40 ° C. and a pressure of 0.17 bar in order to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa.
2.ケカビ(mucor)菌株を滅菌生理食塩水に加え、バクテリア液をピペットで取り、ふすま培地に接種し、20℃で96時間培養し、培養した種麹培養フラスコに500mL滅菌生理食塩水を加えてよく振って均質化し、二重ガーゼで濾過し、500mLの滅菌生理食塩水をフィルター残留物に加え、1回洗浄して濾過し、2回の濾液を混合してケカビ胞子懸濁液を作り、ケカビ胞子懸濁液を4℃の冷蔵庫に入れる。 2. 2. Add the mucor strain to sterile physiological saline, take the bacterial solution with a pipette, inoculate the bran medium, incubate at 20 ° C for 96 hours, and add 500 mL sterile physiological saline to the cultured seed koji culture flask. Shake to homogenize, filter with double gauze, add 500 mL of sterile physiological saline to the filter residue, wash once and filter, mix the two filtrates to make a spore suspension, scab. Place the spore suspension in a refrigerator at 4 ° C.
3.全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、排水し、121℃で30分間、高温オートクレーブでスチームして、冷却する。 3. 3. Whole-grain, uniform-sized soybeans are weighed, washed, mixed with water at a volume ratio of 1: 3, drained, steamed at 121 ° C. for 30 minutes in a high temperature autoclave, and cooled.
4.混合接種:分子量カットオフ300kDaのきくらげ多糖保持液とケカビ胞子懸濁液を1:3の質量比で接種し、25℃で48時間培養した後、48時間熟成処理(maturing treatment)してケカビのタイプ-きくらげ多糖300kDa保持液テンペ発酵物を得る。 4. Mixed inoculation: Inoculate a mucor polysaccharide holding solution with a molecular weight cutoff of 300 kDa and a mucor spore suspension at a mass ratio of 1: 3, inoculate at 25 ° C. for 48 hours, and then aged for 48 hours to treat the mucor. Type-Kikurage polysaccharide 300 kDa holding liquid Tempeh fermented product is obtained.
第3の脱毛治療用組成物の調製方法は、以下の工程に従って行われる。 The method for preparing the third composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。分子量カットオフが300kDaのきくらげ多糖保持液を得るために、温度40℃、圧力0.17barで限外濾過する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. Ultrafiltration is performed at a temperature of 40 ° C. and a pressure of 0.17 bar in order to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa.
2.全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、水に10~18時間浸し、120℃で滅菌して、発酵基質を得る。 2. 2. Whole grain and uniform size soybeans are selected, weighed and washed, mixed with water at a volume ratio of 1: 3, soaked in water for 10-18 hours and sterilized at 120 ° C. to obtain a fermentation substrate.
3.発酵基質中の複合溶液の濃度が2ml/Lになるまで、発酵基質に複合溶液を接種する。前記複合溶液は、1:3の質量比できくらげ多糖300kDa保持液と枯草菌接種溶液と混合し、よく振って、4~8℃で72~120時間培養し、枯草菌タイプ―きくらげ多糖300kDa保持液テンペ発酵物を得る。 3. 3. The fermentation substrate is inoculated with the complex solution until the concentration of the complex solution in the fermentation substrate reaches 2 ml / L. The complex solution is mixed with a Bacillus subtilis 300 kDa retention solution and a Bacillus subtilis inoculation solution at a mass ratio of 1: 3, shaken well, and cultured at 4 to 8 ° C. for 72 to 120 hours to retain Bacillus subtilis type-Bacillus polysaccharide 300 kDa. Obtain a liquid tempeh fermented product.
テンペ(発酵大豆)は、主にタンパク質、遊離アミノ酸、必須脂肪酸、可溶性糖質、無機塩、ビタミンなどを含む栄養素が豊富である。さらに、テンペには大豆イソフラボン、大豆サポニン、褐色色素、テンペ線維素溶解酵素、β-グルコシダーゼ、抗生物質、その他の生理活性物質も含まれている。テンペのこれらの活性物質は、一部は生の大豆に由来し、一部は発酵プロセスに関与するさまざまな微生物や酵素による生の大豆の有機物の分解と再組織化などの複雑な生化学反応に由来する。 Tempeh (fermented soybeans) is rich in nutrients, mainly including proteins, free amino acids, essential fatty acids, soluble sugars, inorganic salts, vitamins and the like. In addition, tempeh also contains soy isoflavones, soy saponins, brown pigments, tempeh fibrinolytic enzymes, β-glucosidases, antibiotics and other physiologically active substances. These active substances in Tempe are partly derived from raw soybeans and are partly complex biochemical reactions such as the decomposition and reorganization of raw soybean organics by various microorganisms and enzymes involved in the fermentation process. Derived from.
黒きくらげに含まれる多糖は、抗潰瘍、老化の遅延、血中脂質の低下、抗肝炎、抗突然変異、抗腫瘍、タンパク質および核酸代謝の促進、体の免疫の促進などの機能を持っている。真菌の多糖とテンペとの共同発酵は、膜分離によって切り詰められて分離され、毛髪固定効果は明らかである。 Polysaccharides contained in black sardines have functions such as anti-ulcer, delayed aging, lowering blood lipids, anti-hepatitis, anti-mutation, anti-tumor, promotion of protein and nucleic acid metabolism, and promotion of body immunity. .. The co-fermentation of fungal polysaccharides with tempeh is truncated and separated by membrane separation, with a clear hair-fixing effect.
1.従来の育毛は、表皮マッサージにショウガとクロベの葉を使用して頭皮の微小循環を促進する。本発明の発酵製品の経口投与は、腸内細菌叢の構造を修復し、微生態学的バランスを回復することができる。食事によって腸内細菌叢を調節することで髪を固定する効果を達成するために、それはまだ空白の領域である。 1. 1. Traditional hair growth uses ginger and kurobe leaves for epidermal massage to promote microcirculation of the scalp. Oral administration of the fermented product of the present invention can repair the structure of the intestinal flora and restore the microecological balance. It is still a blank area to achieve the hair-fixing effect by regulating the intestinal flora by diet.
2.動物実験において、本発明の組み合わせは、優れた毛髪固定効果を示し、将来、大規模な脱毛の問題を解決する新しい方法を見出した。 2. 2. In animal experiments, the combination of the present invention showed excellent hair fixing effect and found a new method to solve the problem of large-scale hair loss in the future.
3.本発明の脱毛治療用組成物は、毛包の成長に対する有意な促進効果および脂漏性脱毛症に対する有意な改善効果を有する。 3. 3. The composition for treating hair loss of the present invention has a significant promoting effect on hair follicle growth and a significant improving effect on seborrheic alopecia.
本発明の技術的解決策は、以下に列挙される特定の実施形態に限定されず、特定の実施形態の任意の組み合わせも含む。 The technical solutions of the present invention are not limited to the particular embodiments listed below, but include any combination of the particular embodiments.
[特定実施例1]
脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Specific Example 1]
The method for preparing the composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。分子量カットオフが300kDaのきくらげ多糖保持液を得るために、温度40℃、圧力0.17barで限外濾過する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. Ultrafiltration is performed at a temperature of 40 ° C. and a pressure of 0.17 bar in order to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa.
2.全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、30℃で3時間浸して(含水率45%~55%)、十分に加熱調理して、調理したものを30℃~40℃までに冷却する。分子量カットオフ300kDaのきくらげ多糖保持液とアスペルギルス種菌液を1:3の質量比で接種し、室温を28℃に保ち、麹を1回目は20~24時間、2回目は28時間になるとかき混ぜた。34時間後に麹ができる。麹ができたら麹を洗い、12時間密閉してからタンクに入れ、28℃の恒温室に入れて30日以上発酵させ、取り出して乾燥させて麹菌型―きくらげ多糖300kDaの保持液テンペ発酵物を得る。 2. 2. Select soybeans of uniform size in whole grains, weigh and wash, mix with water at a volume ratio of 1: 3, soak at 30 ° C. for 3 hours (moisture content 45% -55%), and heat sufficiently. Cook and cool the cooked to 30 ° C-40 ° C. The Kikurage polysaccharide holding solution with a molecular weight cutoff of 300 kDa and the Aspergillus inoculum solution were inoculated at a mass ratio of 1: 3, the room temperature was kept at 28 ° C, and the inoculum was stirred for 20 to 24 hours for the first time and 28 hours for the second time. .. Jiuqu is formed after 34 hours. When the jiuqu is made, wash the jiuqu, seal it for 12 hours, put it in a tank, put it in a constant temperature room at 28 ° C to ferment it for 30 days or more, take it out and dry it. obtain.
本実施形態の工程1における前記ウォーターバス抽出の温度は、70℃~85℃である。 The temperature of the water bath extraction in step 1 of the present embodiment is 70 ° C to 85 ° C.
本実施形態の工程1では、遠心分離された上澄みに蒸留水が添加され、上澄み中の黒きくらげ多糖が4g/Lに希釈される。 In step 1 of the present embodiment, distilled water is added to the centrifuged supernatant, and the black fungus polysaccharide in the supernatant is diluted to 4 g / L.
本実施形態の工程2の前記調理したものは調理済み大豆である。 The cooked product in step 2 of the present embodiment is cooked soybeans.
本実施形態の工程2では、200gごとの調理済み大豆に3mlのきくらげ多糖保持液およびアスペルギルス接種液を接種し、アスペルギルス胞子の濃度は1.0×107~1.0×108胞子/mlである。 In step 2 of the present embodiment, cooked soybeans of every 200 g are inoculated with 3 ml of a Kikurage polysaccharide holding solution and an Aspergillus inoculum, and the concentration of Aspergillus spores is 1.0 × 10 7 to 1.0 × 10 8 spores / ml. Is.
本実施形態の工程2において、「密閉」とは、温度が30±2℃の密閉インキュベーターでの処理を指す。 In step 2 of the present embodiment, "sealing" refers to processing in a closed incubator having a temperature of 30 ± 2 ° C.
[特定実施例2]
本実施形態と実施形態1との違いは、工程2における含水率が50%であることである。その他は実施形態1と同様である。
[Specific Example 2]
The difference between the present embodiment and the first embodiment is that the water content in the step 2 is 50%. Others are the same as those in the first embodiment.
[特定実施例3]
本実施形態は、工程2で調理したものが35℃に冷却されるという点で特定の実施例1または2とは異なる。その他は、実施形態1または2と同様である。
[Specific Example 3]
This embodiment differs from the specific Example 1 or 2 in that the cooked product in step 2 is cooled to 35 ° C. Others are the same as those of the first or second embodiment.
[特定実施例4]
この実施例は、工程2で麹のかき混ぜの1回目が麹作りから22時間目に実行されるという点で、特定の実施例1~3とは異なる。その他は、実施形態1~3と同様である。
[Specific Example 4]
This embodiment differs from the specific Examples 1 to 3 in that the first stirring of the Jiuqu in step 2 is performed 22 hours after the production of the Jiuqu. Others are the same as those of the first to third embodiments.
[特定実施例5]
この実施例に係る脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Specific Example 5]
The method for preparing the composition for hair loss treatment according to this example is carried out according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。分子量カットオフが300kDaのきくらげ多糖保持液を得るために、温度40℃、圧力0.17barで限外濾過する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. Ultrafiltration is performed at a temperature of 40 ° C. and a pressure of 0.17 bar in order to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa.
2.ケカビ菌株を滅菌生理食塩水に加え、バクテリア液をピペットで取り、ふすま培地に接種し、20℃で96時間培養する(三角フラスコのふすまは菌糸と灰色の胞子で覆われており、後で使用するために取り出される)。培養した種麹培養フラスコに滅菌生理食塩水500mLを加えてよく振り、二重のガーゼでろ過し、フィルター残渣に滅菌生理食塩水500mLを加え、1回洗浄してろ過し、2つのろ液を混合してケカビ胞子懸濁液を作成する。当該ケカビ胞子懸濁液を4℃の冷蔵庫に入れる。 2. 2. Add the mucor strain to sterile saline, pipette the bacterial solution, inoculate the bran medium, and incubate at 20 ° C. for 96 hours (the bran in the Erlenmeyer flask is covered with hyphae and gray spores and will be used later. Taken out to do). Add 500 mL of sterile saline to the cultured seed koji culture flask, shake well, filter with double gauze, add 500 mL of sterile saline to the filter residue, wash once and filter, and filter the two filtrates. Mix to make a spore suspension. Place the mucor spore suspension in a refrigerator at 4 ° C.
3.全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、排水し、121℃で30分間、高温オートクレーブでスチームして、冷却する。 3. 3. Whole-grain, uniform-sized soybeans are weighed, washed, mixed with water at a volume ratio of 1: 3, drained, steamed at 121 ° C. for 30 minutes in a high temperature autoclave, and cooled.
4.混合接種
分子量カットオフ300kDaのきくらげ多糖保持液とケカビ胞子懸濁液を1:3の質量比で接種し、25℃で48時間培養した後、48時間熟成処理(maturing treatment)してケカビのタイプ-きくらげ多糖300kDa保持液テンペ発酵物を取得する。
4. Mixed inoculation A molecular weight cutoff of 300 kDa Kikurage polysaccharide holding solution and mucor spore suspension are inoculated at a mass ratio of 1: 3, and after culturing at 25 ° C. for 48 hours, they are aged for 48 hours (muturing treatment) to form a mucor type. -Obtain a mucor polysaccharide 300 kDa holding liquid tempeh fermented product.
本実施形態では、工程1のウォーターバス抽出の温度は、70℃~85℃である。 In the present embodiment, the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
本実施形態の工程1において、遠心分離された上澄みに蒸留水が添加され、上澄み中の黒きくらげ多糖が4g/Lに希釈される。 In step 1 of the present embodiment, distilled water is added to the centrifuged supernatant, and the black fungus polysaccharide in the supernatant is diluted to 4 g / L.
本実施形態の工程2に記載されている工程2では、調理された大豆200gごとに、合計2mlのきくらげ多糖保持液およびケカビ胞子懸濁液が接種される。 In step 2 described in step 2 of the present embodiment, a total of 2 ml of a wood ear polysaccharide holding solution and a mucor spore suspension are inoculated for every 200 g of cooked soybeans.
本実施形態ケカビ胞子懸濁液中の胞子の濃度は、1.0×107~1.0×108胞子/mlである。 The concentration of spores in the mucor spore suspension of the present embodiment is 1.0 × 107 to 1.0 × 10 8 spores / ml.
[特定実施例6]
本特定の実施例に係る脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Specific Example 6]
The method for preparing the composition for hair loss treatment according to the present specific embodiment is carried out according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。分子量カットオフが300kDaのきくらげ多糖保持液を得るために、温度40℃、圧力0.17barで限外濾過する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. Ultrafiltration is performed at a temperature of 40 ° C. and a pressure of 0.17 bar in order to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa.
2.全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、水に10~18時間浸し、120℃で滅菌して、発酵基質を得る。 2. 2. Whole grain and uniform size soybeans are selected, weighed and washed, mixed with water at a volume ratio of 1: 3, soaked in water for 10-18 hours and sterilized at 120 ° C. to obtain a fermentation substrate.
3.発酵基質中の複合溶液の濃度が2ml/Lになるまで、発酵基質に複合溶液を接種する。前記複合溶液は、1:3の質量比できくらげ多糖300kDa保持液と枯草菌接種溶液と混合し、よく振って、4~8℃で72~120時間培養し、枯草菌タイプ―きくらげ多糖300kDa保持液テンペ発酵物を得る。 3. 3. The fermentation substrate is inoculated with the complex solution until the concentration of the complex solution in the fermentation substrate reaches 2 ml / L. The complex solution is mixed with a Bacillus subtilis 300 kDa retention solution and a Bacillus subtilis inoculation solution at a mass ratio of 1: 3, shaken well, and cultured at 4 to 8 ° C. for 72 to 120 hours to retain Bacillus subtilis type-Bacillus polysaccharide 300 kDa. Obtain a liquid tempeh fermented product.
本実施形態では、工程1の前記ウォーターバス抽出の温度は、70℃~85℃である。 In the present embodiment, the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
本実施形態の工程1において、遠心分離された上澄みに蒸留水が添加され、上澄み中の黒きくらげ多糖が4g/Lに希釈される。 In step 1 of the present embodiment, distilled water is added to the centrifuged supernatant, and the black fungus polysaccharide in the supernatant is diluted to 4 g / L.
本実施形態の枯草菌接種液の胞子濃度は、1.0×107~1.0×108胞子/mlである。 The spore concentration of the Bacillus subtilis inoculum of the present embodiment is 1.0 × 107 to 1.0 × 10 8 spores / ml.
以下の実験は、本発明の効果を検証するために使用される。 The following experiments are used to verify the effectiveness of the invention.
[実施例1]
脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Example 1]
The method for preparing the composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。分子量カットオフが300kDaのきくらげ多糖保持液を得るために、温度40℃、圧力0.17barで限外濾過する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. Ultrafiltration is performed at a temperature of 40 ° C. and a pressure of 0.17 bar in order to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa.
2.全粒で均一なサイズの大豆を選択し、計量して洗浄し、大豆と水の体積比1:3で、30℃の水に3時間浸し、排水し、高温オートクレーブで121℃、30分間蒸し、冷却する。分子量カットオフが300kDaのきくらげ多糖保持液を添加し、25℃で48時間培養した後、テンペ予備発酵産物を得る。48時間熟成処理(maturing treatment)して大豆-きくらげ多糖300kDaカットオフ発酵物を取得す。 2. 2. Select soybeans of uniform size for all grains, weigh and wash, soak in water at 30 ° C for 3 hours at a volume ratio of soybean to water 1: 3, drain, and steam at 121 ° C for 30 minutes in a high temperature autoclave. ,Cooling. A Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa is added, and the mixture is cultured at 25 ° C. for 48 hours to obtain a tempeh pre-fermented product. A soybean-Kikurage polysaccharide 300 kDa cut-off fermented product is obtained by aging treatment for 48 hours.
本実施形態では、工程1の前記ウォーターバス抽出の温度は、70℃~85℃である。 In the present embodiment, the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
本実施例の工程1では、遠心分離された上澄みに蒸留水が添加され、上澄み中の黒きくらげ多糖が4g/Lに希釈される。 In step 1 of this example, distilled water is added to the centrifuged supernatant, and the black fungus polysaccharide in the supernatant is diluted to 4 g / L.
[実施例2]
脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Example 2]
The method for preparing the composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。温度40℃、圧力0.17barの条件下での限外濾過では、きくらげ多糖300kDa透過液を選択する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. For ultrafiltration under the conditions of a temperature of 40 ° C. and a pressure of 0.17 bar, a 300 kDa permeate of Kikurage polysaccharide is selected.
2.全粒で均一なサイズの大豆を選択し、計量して洗浄し、大豆と水の体積比1:3で30℃で3時間水に浸し、排水し、高温オートクレーブで121℃、30分間蒸し、冷却する。きくらげ多糖300kDa透過液を添加し、25℃で48時間培養してテンペ予備発酵産物を得る。その後48時間熟成処理(maturing treatment)が行われる。こうして、大豆-きくらげ多糖300kDa透過液発酵物を得る。 2. 2. Soybeans of uniform size are selected, weighed and washed, soaked in water at 30 ° C for 3 hours at a volume ratio of soybean and water 1: 3, drained, and steamed in a high temperature autoclave at 121 ° C for 30 minutes. Cooling. A 300 kDa permeate of Kikurage polysaccharide is added and cultured at 25 ° C. for 48 hours to obtain a tempeh pre-fermented product. After that, a aging treatment for 48 hours is performed. In this way, a fermented soybean-Kikurage polysaccharide 300 kDa permeate is obtained.
本実施形態では、工程1の前記ウォーターバス抽出の温度は、70℃~85℃である。 In the present embodiment, the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
この例の工程1では、遠心分離された上澄みに蒸留水が添加され、上澄み中の黒きくらげ多糖が4g/Lに希釈される。 In step 1 of this example, distilled water is added to the centrifuged supernatant, and the black fungus polysaccharide in the supernatant is diluted to 4 g / L.
[実施例3]
脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Example 3]
The method for preparing the composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。分子量カットオフが300kDaのきくらげ多糖保持液を得るために、温度40℃、圧力0.17barで限外濾過する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. Ultrafiltration is performed at a temperature of 40 ° C. and a pressure of 0.17 bar in order to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa.
2.ケカビ菌株を滅菌生理食塩水に加え、バクテリア液をピペットで取り、ふすま培地に接種し、三角フラスコのふすまが菌糸と灰色の胞子でいっぱいになるまで20℃で96時間培養し、後で使用するために取り出す。培養した種麹培養フラスコに滅菌生理食塩水500mLを加えてよく振り、二重ガーゼでろ過し、フィルター残渣に滅菌生理食塩水500mLを加え、1回洗浄してろ過し、2つのろ液を混合してケカビ胞子懸濁液を作成する。当該ケカビ胞子懸濁液を4℃の冷蔵庫に入れる。 2. 2. Add the mucor strain to sterile saline, pipette the bacterial solution, inoculate the bran medium, incubate at 20 ° C. for 96 hours until the Erlenmeyer flask bran is filled with hyphae and gray spores, and use later. Take out for. Add 500 mL of sterile saline to the cultured seed koji culture flask, shake well, filter with double gauze, add 500 mL of sterile saline to the filter residue, wash once, filter and mix the two filtrates. To make a spore suspension. Place the mucor spore suspension in a refrigerator at 4 ° C.
3.全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の質量比で水と混合し、30℃で3時間浸し、排水し、121℃で30分間、高温オートクレーブでスチームして、冷却する。 3. 3. Select whole soybeans of uniform size, weigh and wash, mix with water at a mass ratio of 1: 3, soak at 30 ° C for 3 hours, drain, steam at 121 ° C for 30 minutes in a high temperature autoclave. And cool.
4.混合接種
分子量カットオフ300kDaのきくらげ多糖保持液とケカビ胞子懸濁液を1:3の質量比で接種し、25℃で48時間培養した後、48時間熟成処理(maturing treatment)してケカビのタイプ-きくらげ多糖300kDa保持液テンペ発酵物を得る。
4. Mixed inoculation A molecular weight cutoff of 300 kDa Kikurage polysaccharide holding solution and mucor spore suspension are inoculated at a mass ratio of 1: 3, and after culturing at 25 ° C. for 48 hours, they are aged for 48 hours (muturing treatment) to form a mucor type. -Kikurage polysaccharide 300 kDa holding liquid Tempeh fermented product is obtained.
本実施形態では、工程1の前記ウォーターバス抽出の温度は、70℃~85℃である。 In the present embodiment, the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
本実施例の工程1では、遠心分離された上澄みに蒸留水が添加され、上澄み中の黒きくらげ多糖が4g/Lに希釈される。 In step 1 of this example, distilled water is added to the centrifuged supernatant, and the black fungus polysaccharide in the supernatant is diluted to 4 g / L.
本実施形態の工程2では、調理された大豆200gごとに、合計2mlのきくらげ多糖保持液およびケカビ胞子懸濁液が接種される。 In step 2 of the present embodiment, a total of 2 ml of a wood ear polysaccharide holding solution and a mucor spore suspension are inoculated for every 200 g of cooked soybeans.
本実施例に係るケカビ胞子懸濁液中の胞子の濃度は、1.0×107~1.0×108胞子/mlである。 The concentration of spores in the mucor spore suspension according to this example is 1.0 × 107 to 1.0 × 10 8 spores / ml.
[実施例4]
脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Example 4]
The method for preparing the composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。温度40℃、圧力0.17barの条件下での限外濾過では、きくらげ多糖300kDa透過液を選択する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. For ultrafiltration under the conditions of a temperature of 40 ° C. and a pressure of 0.17 bar, a 300 kDa permeate of Kikurage polysaccharide is selected.
2.ケカビ菌株を滅菌生理食塩水に加え、バクテリア液をピペットで取り、ふすま培地に接種し、三角フラスコのふすまが菌糸と灰色の胞子でいっぱいになるまで20℃で96時間培養し、後で使用するために取り出す。培養した種麹培養フラスコに滅菌生理食塩水500mLを加えてよく振り、二重ガーゼでろ過し、フィルター残渣に滅菌生理食塩水500mLを加え、1回洗浄してろ過し、2つのろ液を混合してケカビ胞子懸濁液を作成する。当該ケカビ胞子懸濁液を4℃の冷蔵庫に入れ、菌株として保存する。 2. 2. Add the mucor strain to sterile saline, pipette the bacterial solution, inoculate the bran medium, incubate at 20 ° C. for 96 hours until the Erlenmeyer flask bran is filled with hyphae and gray spores, and use later. Take out for. Add 500 mL of sterile saline to the cultured seed koji culture flask, shake well, filter with double gauze, add 500 mL of sterile saline to the filter residue, wash once, filter and mix the two filtrates. To make a spore suspension. The mucor spore suspension is placed in a refrigerator at 4 ° C. and stored as a strain.
3.全粒で均一なサイズの大豆を選択し、計量して洗浄し、大豆と水の体積比1:3で、30℃の水に3時間浸し、排水し、間高温オートクレーブで121℃、30分蒸し、冷却しておく。きくらげ多糖300kDa透過液とケカビ胞子懸濁液を1:3の質量比で接種し、25℃で48時間培養した後、テンペ予備発酵産物を得る。48時間熟成処理(maturing treatment)してケカビのタイプ-きくらげ多糖300kDa透過液テンペ発酵物を得る。 3. 3. Select soybeans of uniform size for all grains, weigh and wash, soak in water at 30 ° C for 3 hours at a volume ratio of soybean to water 1: 3, drain, and use a high-temperature autoclave at 121 ° C for 30 minutes. Steam and cool. A 300 kDa permeate of Kikurage polysaccharide and a mucor spore suspension are inoculated at a mass ratio of 1: 3, and after culturing at 25 ° C. for 48 hours, a tempeh pre-fermented product is obtained. It is aged for 48 hours to obtain a mucor type-Kikurage polysaccharide 300 kDa permeate tempeh fermented product.
本実施形態では、工程1の前記ウォーターバス抽出の温度は、70℃~85℃である。 In the present embodiment, the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
本実施例の工程1では、遠心分離された上澄みに蒸留水が添加され、上澄み中の黒きくらげ多糖が4g/Lに希釈される。 In step 1 of this example, distilled water is added to the centrifuged supernatant, and the black fungus polysaccharide in the supernatant is diluted to 4 g / L.
本実施形態の工程2では、調理された大豆200gごとに、合計2mlのきくらげ多糖保持液およびケカビ胞子懸濁液が接種される。 In step 2 of the present embodiment, a total of 2 ml of a wood ear polysaccharide holding solution and a mucor spore suspension are inoculated for every 200 g of cooked soybeans.
本実施例に係るケカビ胞子懸濁液中の胞子の濃度は、1.0×107~1.0×108胞子/mlである。 The concentration of spores in the mucor spore suspension according to this example is 1.0 × 107 to 1.0 × 10 8 spores / ml.
[実施例5]
脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Example 5]
The method for preparing the composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。分子量カットオフが300kDaのきくらげ多糖保持液を得るために、温度40℃、圧力0.17barで限外濾過する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. Ultrafiltration is performed at a temperature of 40 ° C. and a pressure of 0.17 bar in order to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa.
2.全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、含水率が45%-55%になるようにする。十分に加熱調理して、調理したものを30℃~40℃までに冷却する。分子量カットオフ300kDaのきくらげ多糖保持液とアスペルギルス種菌液を1:3の質量比で接種し、室温を28℃に保ち、麹のかき混ぜを1回目は22時間、2回目は28時間麹を作った。34時間後に麹ができる。麹ができたら麹を洗い、12時間密閉してからタンクに入れる28℃の恒温室に入れて30日以上発酵させ、取り出して乾燥させて麹菌型-きくらげ多糖300kDaの保持液テンペ発酵物を得る。 2. 2. Whole grain and uniform size soybeans are selected, weighed and washed and mixed with water in a 1: 3 volume ratio to a moisture content of 45% -55%. Cook well and cool the cooked to 30 ° C-40 ° C. The Kikurage polysaccharide holding solution with a molecular weight cutoff of 300 kDa and the Aspergillus inoculum solution were inoculated at a mass ratio of 1: 3, the room temperature was kept at 28 ° C, and the inoculum was stirred for 22 hours for the first time and 28 hours for the second time. .. Jiuqu is formed after 34 hours. When the jiuqu is made, wash the jiuqu, seal it for 12 hours, and put it in a tank. ..
本実施形態では、工程1の前記ウォーターバス抽出の温度は、70℃~85℃である。 In the present embodiment, the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
本実施例の工程1では、遠心分離された上澄みに蒸留水が添加され、上澄み中の黒きくらげ多糖が4g/Lに希釈される。 In step 1 of this example, distilled water is added to the centrifuged supernatant, and the black fungus polysaccharide in the supernatant is diluted to 4 g / L.
本実施形態の工程2では、200gごとの調理済み大豆に3mlのきくらげ多糖保持液およびアスペルギルス接種液を接種し、アスペルギルス胞子の濃度は1.0×107~1.0×108胞子/mlである。 In step 2 of the present embodiment, cooked soybeans of every 200 g are inoculated with 3 ml of a Kikurage polysaccharide holding solution and an Aspergillus inoculum, and the concentration of Aspergillus spores is 1.0 × 10 7 to 1.0 × 10 8 spores / ml. Is.
本実施形態の工程2における「密閉」とは、温度が30±2℃の密閉インキュベーターでの処理を指す。 “Sealing” in step 2 of the present embodiment refers to processing in a closed incubator having a temperature of 30 ± 2 ° C.
[実施例6]
脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Example 6]
The method for preparing the composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、その後超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。温度40℃、圧力0.17barの条件下での限外濾過では、きくらげ多糖300kDa透過液を選択する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. The dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and then ultrasonic waves having an ultrasonic power of 500 W are extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifuge for a minute to remove the residue, and prepare the extract into a feed solution with a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. For ultrafiltration under the conditions of a temperature of 40 ° C. and a pressure of 0.17 bar, a 300 kDa permeate of Kikurage polysaccharide is selected.
2.全粒で均一なサイズの大豆を選択し、計量して洗浄し、含水率50%になるまで大豆と水の体積比1:3で、30℃の水に3時間浸し、十分に加熱調理して、調理したものを35℃まで冷却する。きくらげ多糖300kDa透過液とアスペルギルス種菌液を1:3の質量比で接種し、よく混ぜ、室温を28℃に保ち、麹のかき混ぜの1回目は麹を作ってから22時間目、2回目は28時間目に開始される。34時間後に麹ができる。麹が製造された後、麹を洗浄し、12時間密閉してからタンクに入れ、28℃の恒温室で30日以上発酵する。麹菌型-きくらげ多糖300kDa透過液テンペ発酵物を得るためには、取り出して乾燥させる。 2. 2. Select soybeans of uniform size for all grains, weigh and wash, soak in water at 30 ° C for 3 hours with a volume ratio of soybean and water of 1: 3 until the water content reaches 50%, and cook thoroughly. And cool the cooked food to 35 ° C. Inject Kikurage polysaccharide 300 kDa permeate and Aspergillus inoculum at a mass ratio of 1: 3, mix well, keep the room temperature at 28 ° C, stir the inoculum 22 hours after making the inoculum, and 28 in the second. It starts at the hour. Jiuqu is formed after 34 hours. After the jiuqu is produced, the jiuqu is washed, sealed for 12 hours, placed in a tank, and fermented in a constant temperature room at 28 ° C. for 30 days or more. Jiuqu-type-Kikurage polysaccharide 300 kDa permeate Tempeh To obtain a fermented product, take it out and dry it.
本実施形態では、工程1の前記ウォーターバス抽出の温度は、70℃~85℃である。 In the present embodiment, the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
本実施例の工程1では、遠心分離された上澄みに蒸留水が添加され、上澄み中の黒きくらげ多糖が4g/Lに希釈される。 In step 1 of this example, distilled water is added to the centrifuged supernatant, and the black fungus polysaccharide in the supernatant is diluted to 4 g / L.
本実施形態の工程2では、調理された大豆200gごとに、合計3mlのきくらげ多糖保持液およびアスペルギルス種菌が接種される。アスペルギルス胞子の濃度は1.0×107~1.0×108胞子/mlである。 In step 2 of the present embodiment, a total of 3 ml of the Kikurage polysaccharide holding solution and Aspergillus inoculum are inoculated for every 200 g of cooked soybeans. The concentration of Aspergillus spores is 1.0 × 107 to 1.0 × 10 8 spores / ml.
本実施形態の工程2における「密閉」とは、温度が30±2℃の密閉インキュベーターでの処理を指す。 “Sealing” in step 2 of the present embodiment refers to processing in a closed incubator having a temperature of 30 ± 2 ° C.
[実施例7]
脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Example 7]
The method for preparing the composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。分子量カットオフが300kDaのきくらげ多糖保持液を得るために、温度40℃、圧力0.17barで限外濾過する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. Ultrafiltration is performed at a temperature of 40 ° C. and a pressure of 0.17 bar in order to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa.
2.全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の質量比で水と混合し、水に10~18時間浸し、120℃で滅菌して、発酵基質を得る。 2. 2. Whole grain and uniform size soybeans are selected, weighed and washed, mixed with water at a mass ratio of 1: 3, soaked in water for 10-18 hours and sterilized at 120 ° C. to give the fermentation substrate.
3.発酵基質中の複合溶液の濃度が2ml/Lになるまで、発酵基質に複合溶液を接種する。前記複合溶液は、1:3の質量比できくらげ多糖300kDa保持液と枯草菌接種溶液と混合し、よく振って、4~8℃で72~120時間培養し、枯草菌タイプ―きくらげ多糖300kDa保持液テンペ発酵物を得る。 3. 3. The fermentation substrate is inoculated with the complex solution until the concentration of the complex solution in the fermentation substrate reaches 2 ml / L. The complex solution is mixed with a Bacillus subtilis 300 kDa retention solution and a Bacillus subtilis inoculation solution at a mass ratio of 1: 3, shaken well, and cultured at 4 to 8 ° C. for 72 to 120 hours to retain Bacillus subtilis type-Bacillus polysaccharide 300 kDa. Obtain a liquid tempeh fermented product.
本実施形態では、工程1の前記ウォーターバス抽出の温度は、70℃~85℃である。 In the present embodiment, the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
本実施例の工程1では、遠心分離された上澄みに蒸留水が添加され、上澄みの黒きくらげ多糖を4g/Lに希釈する。 In step 1 of this example, distilled water is added to the centrifuged supernatant to dilute the black fungus polysaccharide of the supernatant to 4 g / L.
本実施例に係る枯草菌接種液の胞子濃度は、1.0×107~1.0×108胞子/mlである。 The spore concentration of the Bacillus subtilis inoculum according to this example is 1.0 × 107 to 1.0 × 10 8 spores / ml.
[実施例8]
脱毛治療用組成物の調製方法は、以下の工程に従って行われる。
[Example 8]
The method for preparing the composition for treating hair loss is performed according to the following steps.
1.きくらげ多糖の膜分離
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成する。黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製する。温度40℃、圧力0.17barの条件下での限外濾過では、きくらげ多糖300kDa透過液を選択する。
1. 1. Membrane Separation of Kikurage Polysaccharide The black ear is washed, impurities are removed, crushed and passed through a 40 mesh sieve to produce a dry powder of the black ear. Dry powder of black kikurage is added to distilled water at a mass ratio of 140: 1, and an ultrasonic wave having an ultrasonic power of 500 W is extracted over 15 minutes in a water bath for 3 hours, and the extract is extracted at 4500 r / min for 10 minutes. Centrifugation is performed to remove the residue, and the extract is prepared as a feed solution having a black sardine polysaccharide concentration of 4 g / L and a pH value of 5.6. For ultrafiltration under the conditions of a temperature of 40 ° C. and a pressure of 0.17 bar, a 300 kDa permeate of Kikurage polysaccharide is selected.
2.全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の質量比で水と混合し、水に10~18時間浸し、120℃で滅菌して、発酵基質を得る。 2. 2. Whole grain and uniform size soybeans are selected, weighed and washed, mixed with water at a mass ratio of 1: 3, soaked in water for 10-18 hours and sterilized at 120 ° C. to give the fermentation substrate.
3.発酵基質中の複合溶液の濃度が2ml/Lになるまで、発酵基質に複合溶液を接種する。前記複合溶液は、1:3の質量比できくらげ多糖300kDa保持液と枯草菌接種溶液と混合し、よく振って、4~8℃で72~120時間培養し、枯草菌―きくらげ多糖300kDa透過液テンペ発酵物を得る。 3. 3. The fermentation substrate is inoculated with the complex solution until the concentration of the complex solution in the fermentation substrate reaches 2 ml / L. The complex solution is mixed with a Bacillus subtilis 300 kDa holding solution and a Bacillus subtilis inoculation solution at a mass ratio of 1: 3, shaken well, and cultured at 4 to 8 ° C. for 72 to 120 hours. Obtain a tempeh fermented product.
本実施形態では、工程1の前記ウォーターバス抽出の温度は、70℃~85℃である。 In the present embodiment, the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
本実施例の工程1では、遠心分離された上澄みに蒸留水が添加され、上澄み中の黒きくらげ多糖が4g/Lに希釈される。 In step 1 of this example, distilled water is added to the centrifuged supernatant, and the black fungus polysaccharide in the supernatant is diluted to 4 g / L.
本実施例に係る枯草菌接種液の胞子濃度は、1.0×107~1.0×108胞子/mlである。 The spore concentration of the Bacillus subtilis inoculum according to this example is 1.0 × 107 to 1.0 × 10 8 spores / ml.
[実験1]脂漏性マウスの脱毛に対する本発明の経口組成物の阻害実験
1.実験材料
テストステロンプロピオン酸注射液25mg/1ml、寧波第2ホルモン工場から購入、生産バッチ番号:11025である。
サラダ油90ml/瓶、アロワナ株式会社
成体オスのB6CBAF1/Jマウス、体重18~22g、クリーングレード、北京維通利華実験動物技術株式会社(Beijing Weitong Lihua Laboratory Animal Technology CO. Ltd.) [ライセンス番号:SCXK(京)2017-0011]から購入、室温23℃、湿度50-60%、1日2回、毎回1時間換気マウスの自然な生活パターンによれば、B6CBAF1/Jマウスは自由に動き、自由に飲んで食べられる。
[Experiment 1] Inhibition experiment of the oral composition of the present invention for hair loss in seborrheic mice 1. Experimental material Testosterone propionic acid injection 25 mg / 1 ml, purchased from Ningbo No. 2 Hormone Factory, production batch number: 11025.
Salad oil 90 ml / bottle, Arowana Co., Ltd. Adult male B6CBAF1 / J mouse, weight 18-22 g, clean grade, Beijing Weitong Lihua Laboratory Animal Technology CO. Ltd. [License number: SC] (K computer) 2017-0011], room temperature 23 ° C, humidity 50-60%, twice a day, 1 hour each time According to the natural life pattern of the mice, the B6CBAF1 / J mice can move freely and freely. You can drink and eat.
2.実験方法
2.1実験グループ化
100匹のオスのマウスから、10匹のB6CBAF1/Jマウスが無作為に実験対照群として選択され、残りの90匹のB6CBAF1/Jマウスが実験用マウスとして使用され、別々のケージで飼育された。すなわち、脱毛モデル群と口腔用組成物例1~8群である。
2. 2. Experimental method 2.1 Experimental grouping From 100 male mice, 10 B6CBAF1 / J mice were randomly selected as the experimental control group, and the remaining 90 B6CBAF1 / J mice were used as experimental mice. , Raised in separate cages. That is, the hair loss model group and the oral composition examples 1 to 8 groups.
2.2動物モデルの作成と投与方法
脱毛動物モデルは注射によって作成された。注射部位は脊椎の背中と平行にマウスの背中の皮下にある。実験対照群の場合生理食塩水を注射し、他の9群にはプロピオン酸テストステロンを注射した。注射用量は5mg/kg・dである。注射時間は4週間である。
2.2 Preparation and administration method of animal model The depilatory animal model was created by injection. The injection site is subcutaneously on the back of the mouse, parallel to the back of the spine. In the case of the experimental control group, saline was injected, and the other 9 groups were injected with testosterone propionate. The injection dose is 5 mg / kg · d. The injection time is 4 weeks.
脱毛のB6CBAF1/Jマウスモデルが確立されてから4週目に薬物治療を開始し、対照群を除いて、脱毛モデル群に各実施形態の組成物を6週間強制経口投与した。脱毛部位でのマウスの発毛状態を観察し、マウスの脱毛の重量を計算する。病理学的観察と組み合わせて、モデル群と比較して、顕微鏡下での硬毛/軟毛の比率が平均として計算される。 Drug treatment was started 4 weeks after the B6CBAF1 / J mouse model of hair loss was established, and the composition of each embodiment was orally administered to the hair loss model group for 6 weeks except for the control group. Observe the hair growth state of the mouse at the hair loss site and calculate the weight of the mouse hair loss. In combination with pathological observations, the ratio of hair / vellus hair under a microscope is calculated as an average compared to the model group.
3.データの統計分析
SPSS 20.0ソフトウェアをデータ処理に使用した。群間の差異は、一元配置分散分析によって分析され、p<0.05を有意差とし、統計的に有意であると見なされた。
3. 3. Statistical analysis of data SPSS 20.0 software was used for data processing. Differences between groups were analyzed by one-way ANOVA and were considered statistically significant with p <0.05 as the significant difference.
4.実験結果
(1)脱毛重量の決定
表1から、実験対照群と比較して、他の実験用マウスでは脱毛の程度が異なることがわかる。実施例3、5、7の有意差(p<0.05)を除いて、他の群では極端な有意差(p<0.01)脱毛が見られる。モデル群と比較して、例3、5、および7群のマウスの脱毛現象は改善されている(p<0.01)。他の実施例群では有意差はなく、脱毛には改善傾向が見えない。
4. Experimental results (1) Determination of hair loss weight From Table 1, it can be seen that the degree of hair loss differs between the other experimental mice as compared with the experimental control group. Except for the significant difference (p <0.05) in Examples 3, 5 and 7, extreme significant difference (p <0.01) hair loss is seen in the other groups. Compared with the model group, the hair loss phenomenon of the mice of Examples 3, 5, and 7 was improved (p <0.01). There was no significant difference in the other example groups, and no improvement tendency was seen in hair loss.
(2)マウスの硬毛/軟毛比の決定
表2から、実験対照群と比較して、モデル群の硬毛/軟毛の比率が大幅に減少することがわかり(p<0.01)、モデル群と比較して、実施例3、5、7は硬毛/軟毛の比率を大幅に改善できる(p<0.01)。比率、他の実験例における硬毛/軟毛の比率は変化しながった。
(2) Determination of mouse hair / vellus hair ratio From Table 2, it was found that the ratio of hair / vellus hair in the model group was significantly reduced compared to the experimental control group (p <0.01), and the model. Compared with the group, Examples 3, 5 and 7 can significantly improve the hair / vellus hair ratio (p <0.01). The ratio, the ratio of hair / vellus hair in other experimental examples, did not change.
(3)本発明の口腔用組成物の実施例3、5、および7は、脂漏性脱毛症に対して有意な改善効果を有し、大豆が菌株による発酵後に明らかな毛髪固定効果を発揮できることを示している。 (3) Examples 3, 5, and 7 of the oral composition of the present invention have a significant improving effect on seborrheic alopecia, and soybean exerts a clear hair fixing effect after fermentation by a strain. It shows that it can be done.
[実験2]発毛期マウスの毛包数に対する経口組成物の影響
1.実験材料
ロジン(Rosin)は深セン吉田ケミカル株式会社(Shenzhen Jitian Chemical Co.,Ltd.)から購入した。パラフィンワックスはアオウェイプラスチックワックス製品(Aowei Glue Wax Products)から購入した。成体のメスのC57BL/6マウス、体重18~22g、クリーングレード、肌の色はピンク、髪の毛は中止期間(interruption)であるマウスである。北京維通利華実験動物技術株式会社(Beijing Weitong Lihua Laboratory Animal Technology Co.Ltd.)[ライセンス番号:SCXK(京)2017-0149]から購入され、室温23℃、湿度50~60%、1日2回、毎回1時間換気マウスの自然な生活パターンによれば、マウスは自由に動き、自由に飲んで食べられる。
[Experiment 2] Effect of oral composition on the number of hair follicles in hair-growth mice 1. The experimental material rosin was purchased from Shenzhen Jitian Chemical Co., Ltd. Paraffin wax was purchased from Aowei Plastic Wax Products. Adult female C57BL / 6 mice, body weight 18-22 g, clean grade, skin color pink, hair is interruption mice. Purchased from Beijing Weitong Lihua Laboratory Co. Ltd. [License No .: SCXK (Kyo) 2017-0149], room temperature 23 ° C, humidity 50-60%, 2 days a day Ventilation for 1 hour each time According to the natural life pattern of mice, mice are free to move, drink and eat freely.
2.実験方法
2.1実験群化
90匹のメスのC57BL/6マウスを無作為に9つの群、実験対照群、および経口組成物例1-8群に分け、別々のケージで飼育した。
2. 2. Experimental method 2.1 Experimental grouping 90 female C57BL / 6 mice were randomly divided into 9 groups, an experimental control group, and an oral composition example 1-8 group, and kept in separate cages.
2.2動物モデルの作成と投与方法
ロジンとパラフィンワックスを1:1で混合した後、加熱して溶かし、マウスの背中に約2cm×3cmの面積で均一に塗布する。冷却して固化した後、背中の毛を取り除く(発毛が中止期間(interruption)でいることを確認する)。強制経口投与により蒸留水を与えられた対照群を除いて、脱毛の翌日、他の群は1日1回、強制経口投与により2ml/100gの強制経口投与量で20日間連続投与された。20日目に、マウスの各群の背中の同じ位置にある皮膚組織ブロックを取り、10%ホルマリンで固定し、切片をパラフィンワックスで包埋する。切片の厚さは約4~5ミクロン、HE染色、高倍率視野での毛包の数(×400)であり、平均値を取り、統計処理を実行する。
2.2 Preparation of animal model and administration method After mixing rosin and paraffin wax at a ratio of 1: 1, heat and melt them, and apply them evenly to the back of the mouse in an area of about 2 cm × 3 cm. After cooling and solidifying, remove the hair on the back (make sure the hair growth is in the interruption period). Except for the control group to which distilled water was given by gavage, the other groups were administered once daily for 20 consecutive days at a gavage dose of 2 ml / 100 g by gavage. On day 20, the co-located skin tissue blocks on the backs of each group of mice are removed, fixed with 10% formalin and the sections are embedded with paraffin wax. Section thickness is about 4-5 microns, HE stain, number of hair follicles in high-power field (x400), averaged and statistically processed.
3.データの統計分析
SPSS20.0ソフトウェアをデータ処理に使用した。群間の差異は、一元配置分散分析によって分析され、p<0.05を有意差とし、統計的に有意であると見なされた。
3. 3. Statistical analysis of data SPSS20.0 software was used for data processing. Differences between groups were analyzed by one-way ANOVA and were considered statistically significant with p <0.05 as the significant difference.
4.実験結果
(1)毛包密度の影響
実験20日目の各群の毛包数を表3に示す。実験対照群と比較して、実施例3、5、および7は実験対照群マウスの毛包の数を増やすことができ(p<0.01)、他の群の実施例はC57BL/6マウスの毛包の数に影響を与えない(p>0.05)。当該結果は、発酵大豆は、C57BL/6マウスの毛包に成長期に入り、成長期間を延長し、退化期への移行を遅らせ、毛包の成長を促進することが示唆されている。
4. Experimental results (1) Effect of hair follicle density Table 3 shows the number of hair follicles in each group on the 20th day of the experiment. Compared to the experimental control group, Examples 3, 5, and 7 were able to increase the number of hair follicles in the experimental control group mice (p <0.01), and the examples in the other groups were C57BL / 6 mice. Does not affect the number of hair follicles (p> 0.05). The results suggest that fermented soybeans enter the hair follicle of C57BL / 6 mice, prolong the growth period, delay the transition to the degeneration phase, and promote the growth of the hair follicle.
(付記)
(付記1)
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成し、黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製し、分子量カットオフが300kDaのきくらげ多糖保持液を得るために、温度40℃、圧力0.17barで限外濾過する、きくらげ多糖の膜分離の工程1と、
全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、十分に加熱調理して、加熱したものを30℃~40℃までに冷却し、分子量カットオフ300kDaのきくらげ多糖保持液とアスペルギルス種菌液を1:3の質量比で接種し、室温を28℃に保ち、麹作りが20~24時間になると1回目の麹のかき混ぜが行われ、麹作りが28時間になると2回目の麹のかき混ぜが行われ、麹作りが34時間になると麹が製造でき、麹が製造された後、麹を洗浄し、12時間密閉してからタンクに入れ、28℃の恒温室に入れて30日以上発酵させ、取り出して乾燥させて麹菌型-きくらげ多糖300kDaの保持液テンペ発酵物を得る工程2と、
を含むことを特徴とする脱毛治療用組成物の調製方法。
(Additional note)
(Appendix 1)
The black sardines are washed, decontaminated, crushed and passed through a 40 mesh sieve to produce a dry black sardine powder, the dry black sardines are added to distilled water at a mass ratio of 140: 1 and ultrasonic. An ultrasonic wave having a power of 500 W was extracted over 15 minutes in a water bath for 3 hours, and the extract was centrifuged at 4500 r / min for 10 minutes to remove the residue. , Prepared in a feed solution with a pH value of 5.6, and subjected to ultrafiltration at a temperature of 40 ° C. and a pressure of 0.17 bar to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa. Step 1 of membrane separation of Kikurage polysaccharide. When,
Select whole-grain, uniform-sized soybeans, weigh and wash, mix with water at a volume ratio of 1: 3, cook well, and cool the heated to 30 ° C-40 ° C. Inject the Kikurage polysaccharide holding solution with a molecular weight cutoff of 300 kDa and the Aspergillus inoculum at a mass ratio of 1: 3, keep the room temperature at 28 ° C, and stir the Jiuqu for the first time when the Jiuqu is made for 20 to 24 hours. , When the koji making is 28 hours, the second stirring of the koji is performed, and when the koji making is 34 hours, the koji can be produced. Put it in a constant temperature room at 28 ° C, ferment it for 30 days or more, take it out and dry it to obtain a Jiuqu-type-Kikurage polysaccharide 300 kDa holding liquid tempeh fermented product.
A method for preparing a composition for treating hair loss, which comprises.
(付記2)
工程2において、30℃で3時間浸して、含水率が50%であることを特徴とする付記1に記載の脱毛治療用組成物の調製方法。
(Appendix 2)
The method for preparing a composition for hair loss treatment according to Appendix 1, wherein in step 2, the composition is immersed at 30 ° C. for 3 hours and has a water content of 50%.
(付記3)
工程2において、調理したものを35℃に冷却することを特徴とする付記1に記載の脱毛治療用組成物の調製方法。
(Appendix 3)
The method for preparing a composition for hair loss treatment according to Appendix 1, wherein the cooked product is cooled to 35 ° C. in step 2.
(付記4)
工程2において、麹作りから22時間目になると1回目の麹のかき混ぜが行われることを特徴とする付記1に記載の脱毛治療用組成物の調製方法。
(Appendix 4)
The method for preparing a composition for hair loss treatment according to Appendix 1, wherein in step 2, the first stirring of the jiuqu is performed 22 hours after the production of the jiuqu.
(付記5)
工程2の200gごとの調理済み大豆に3mlのきくらげ多糖保持液およびアスペルギルス接種液を接種し、アスペルギルス胞子の濃度は1.0×107~1.0×108胞子/mlであることを特徴とする付記1に記載の脱毛治療用組成物の調製方法。
(Appendix 5)
3 ml of Kikurage polysaccharide holding solution and Aspergillus inoculum are inoculated into the cooked soybeans every 200 g in Step 2, and the concentration of Aspergillus spores is 1.0 × 10 7 to 1.0 × 10 8 spores / ml. The method for preparing the composition for hair loss treatment according to Appendix 1.
(付記6)
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成し、黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製し、温度40℃、圧力0.17barの条件下で限外濾過して、分子量カットオフ300kDaのきくらげ多糖保持液を得る、きくらげ多糖の膜分離工程1と、
ケカビ菌株を滅菌生理食塩水に加え、バクテリア液をピペットで取り、ふすま培地に接種し、20℃で96時間培養し、培養した種麹培養フラスコに滅菌生理食塩水500mLを加えてよく振り、二重ガーゼでろ過し、フィルター残渣に滅菌生理食塩水500mLを加え、1回洗浄してろ過し、2つのろ液を混合してケカビ胞子懸濁液を作成し、当該ケカビ胞子懸濁液を4℃の冷蔵庫に入れる工程2と、
全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、排水し、121℃で30分間、高温オートクレーブでスチームして、冷却する工程3と、
分子量カットオフ300kDaのきくらげ多糖保持液とケカビ胞子懸濁液を1:3の質量比で接種し、25℃で48時間培養した後、48時間熟成処理してケカビのタイプーきくらげ多糖30000Daテンペ発酵物を得る、混合接種工程4と、
に従って行われることを特徴とする脱毛治療用組成物の調製方法。
(Appendix 6)
The black sardines are washed, decontaminated, crushed and passed through a 40 mesh sieve to produce a dry black sardine powder, the dry black sardines are added to distilled water at a mass ratio of 140: 1 and ultrasonic. An ultrasonic wave having a power of 500 W was extracted over 15 minutes in a water bath for 3 hours, and the extract was centrifuged at 4500 r / min for 10 minutes to remove the residue. , Prepared in a feed solution with a pH value of 5.6 and subjected to ultrafiltration under the conditions of a temperature of 40 ° C. and a pressure of 0.17 bar to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa. When,
Add the Kekabi strain to sterile physiological saline, take the bacterial solution with a pipette, inoculate it into bran medium, incubate at 20 ° C for 96 hours, add 500 mL of sterile physiological saline to the cultured seed koji culture flask, shake well, and shake well. Filter with heavy gauze, add 500 mL of sterile physiological saline to the filter residue, wash once and filter, mix the two filtrates to make a kekabi spore suspension, and prepare the kekabi spore suspension 4 Step 2 of putting in a refrigerator at ℃ and
Step 3 of selecting soybeans of uniform size in whole grains, weighing, washing, mixing with water at a volume ratio of 1: 3, draining, steaming at 121 ° C. for 30 minutes in a high temperature autoclave, and cooling. When,
Inoculate the mucor polysaccharide holding solution with a molecular weight cutoff of 300 kDa and the mucor spore suspension at a mass ratio of 1: 3, inoculate at 25 ° C for 48 hours, and then aged for 48 hours to treat the mucor type-mucor polysaccharide 30000 Da tempeh fermented product. In the mixed inoculation step 4 and
A method for preparing a composition for treating hair loss, which is performed according to the above.
(付記7)
工程2で調理された大豆200gごとに、合計2mlのきくらげ多糖保持液およびケカビ胞子懸濁液が接種されることを特徴とする付記6に記載の脱毛治療用組成物の調製方法。
(Appendix 7)
The method for preparing a composition for hair loss treatment according to Appendix 6, wherein a total of 2 ml of a wood ear polysaccharide holding solution and a mucor spore suspension is inoculated for every 200 g of soybean cooked in step 2.
(付記8)
黒きくらげを洗浄し、不純物を取り除き、粉砕し、40メッシュのふるいに通して、黒きくらげの乾燥粉末を生成し、黒きくらげの乾燥粉末を蒸留水に質量比140:1で添加し、超音波パワーが500Wである超音波を15分間かけて、ウォーターバスで3時間抽出し、抽出物を4500r/minで10分間遠心分離して残留物を除去し、抽出物を黒きくらげ多糖濃度4g/L、pH値5.6のフィード溶液に調製し、温度40℃、圧力0.17barの条件下での限外濾過では、分子量カットオフ30000Daのきくらげ多糖保持液を得る、きくらげ多糖の膜分離工程1と、
全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、水に10~18時間浸し、120℃で滅菌して、発酵基質を得る工程2と、
発酵基質中の複合溶液の濃度が2ml/Lになるまで、発酵基質に複合溶液を接種し、前記複合溶液は、1:3の質量比できくらげ多糖300kDa保持液と枯草菌接種溶液と混合し、よく振って、4~8℃で72~120時間培養し、枯草菌~きくらげ多糖300kDa保持液テンペ発酵物を得る工程3と、
に従って行われることを特徴とする脱毛治療用組成物の調製方法。
(Appendix 8)
The black sardines are washed, decontaminated, crushed and passed through a 40 mesh sieve to produce a dry black sardine powder, the dry black sardines are added to distilled water at a mass ratio of 140: 1 and ultrasonic. An ultrasonic wave having a power of 500 W was extracted over 15 minutes in a water bath for 3 hours, and the extract was centrifuged at 4500 r / min for 10 minutes to remove the residue. , Prepared in a feed solution with a pH value of 5.6, and obtained a Kikurage polysaccharide holding solution having a molecular weight cutoff of 30,000 Da by ultrafiltration under the conditions of a temperature of 40 ° C. and a pressure of 0.17 bar. When,
A step of selecting soybeans of uniform size in whole grains, weighing and washing, mixing with water at a volume ratio of 1: 3, soaking in water for 10 to 18 hours, and sterilizing at 120 ° C. to obtain a fermentation substrate. 2 and
The fermentation substrate is inoculated with the complex solution until the concentration of the complex solution in the fermentation substrate reaches 2 ml / L, and the complex solution is mixed with the syrup polysaccharide 300 kDa holding solution and the bacillus inoculum solution at a mass ratio of 1: 3. 3. Shake well and incubate at 4-8 ° C. for 72-120 hours to obtain a fermented product of Bacillus subtilis-Kikurage polysaccharide 300 kDa holding solution Tempe.
A method for preparing a composition for treating hair loss, which is performed according to the above.
(付記9)
枯草菌接種液の胞子濃度は、1.0×107~1.0×108胞子/mlであることを特徴とする付記8に記載の脱毛治療用組成物の調製方法。
(Appendix 9)
The method for preparing a composition for treating hair loss according to Appendix 8, wherein the spore concentration of the Bacillus subtilis inoculum is 1.0 × 107 to 1.0 × 10 8 spores / ml.
(付記10)
工程1における前記ウォーターバス抽出の温度は、70℃~85℃であることを特徴とする付記1、6または8に記載の脱毛治療用組成物の調製方法。
(Appendix 10)
The method for preparing a composition for hair loss treatment according to Appendix 1, 6 or 8, wherein the temperature of the water bath extraction in step 1 is 70 ° C to 85 ° C.
Claims (7)
全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、十分に加熱調理して、加熱したものを30℃~40℃までに冷却し、分子量カットオフ300kDaのきくらげ多糖保持液とアスペルギルス種菌液を1:3の質量比で接種し、室温を28℃に保ち、麹作りが20~24時間になると1回目の麹のかき混ぜが行われ、麹作りが28時間になると2回目の麹のかき混ぜが行われ、麹作りが34時間になると麹が製造でき、麹が製造された後、麹を洗浄し、12時間密閉してからタンクに入れ、28℃の恒温室に入れて30日以上発酵させ、取り出して乾燥させて麹菌型-きくらげ多糖300kDaの保持液テンペ発酵物を得る工程2と、
を含むことを特徴とする脱毛治療用組成物の調製方法。 The black sardines are washed, decontaminated, crushed and passed through a 40 mesh sieve to produce a dry black sardine powder, the dry black sardines are added to distilled water at a mass ratio of 140: 1 and ultrasonic. An ultrasonic wave having a power of 500 W was extracted over 15 minutes in a water bath for 3 hours, and the extract was centrifuged at 4500 r / min for 10 minutes to remove the residue. , Prepared in a feed solution with a pH value of 5.6, and subjected to ultrafiltration at a temperature of 40 ° C. and a pressure of 0.17 bar to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa. Step 1 of membrane separation of Kikurage polysaccharide. When,
Select whole-grain, uniform-sized soybeans, weigh and wash, mix with water at a volume ratio of 1: 3, cook well, and cool the heated to 30 ° C-40 ° C. Inject the Kikurage polysaccharide holding solution with a molecular weight cutoff of 300 kDa and the Aspergillus inoculum at a mass ratio of 1: 3, keep the room temperature at 28 ° C, and stir the Jiuqu for the first time when the Jiuqu is made for 20 to 24 hours. , When the koji making is 28 hours, the second stirring of the koji is performed, and when the koji making is 34 hours, the koji can be produced. Put it in a constant temperature room at 28 ° C, ferment it for 30 days or more, take it out and dry it to obtain a Jiuqu-type-Kikurage polysaccharide 300 kDa holding liquid tempeh fermented product.
A method for preparing a composition for treating hair loss, which comprises.
ケカビ菌株を滅菌生理食塩水に加え、バクテリア液をピペットで取り、ふすま培地に接種し、20℃で96時間培養し、培養した種麹培養フラスコに滅菌生理食塩水500mLを加えてよく振り、二重ガーゼでろ過し、フィルター残渣に滅菌生理食塩水500mLを加え、1回洗浄してろ過し、2つのろ液を混合してケカビ胞子懸濁液を作成し、当該ケカビ胞子懸濁液を4℃の冷蔵庫に入れる工程2と、
全粒で均一なサイズの大豆を選択し、計量して洗浄し、1:3の体積比で水と混合し、排水し、121℃で30分間、高温オートクレーブでスチームして、冷却する工程3と、
前記工程3を経た大豆に、分子量カットオフ300kDaのきくらげ多糖保持液とケカビ胞子懸濁液を1:3の質量比で接種し、25℃で48時間培養した後、48時間熟成処理してケカビのタイプ-きくらげ多糖300kDaテンペ発酵物を得る、混合接種工程4と、
に従って行われることを特徴とする脱毛治療用組成物の調製方法。 The black sardines are washed, decontaminated, crushed and passed through a 40 mesh sieve to produce a dry black sardine powder, the dry black sardines are added to distilled water at a mass ratio of 140: 1 and ultrasonic. An ultrasonic wave having a power of 500 W was extracted over 15 minutes in a water bath for 3 hours, and the extract was centrifuged at 4500 r / min for 10 minutes to remove the residue. , Prepared in a feed solution with a pH value of 5.6 and subjected to ultrafiltration under the conditions of a temperature of 40 ° C. and a pressure of 0.17 bar to obtain a Kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa. When,
Add the Kekabi strain to sterile physiological saline, take the bacterial solution with a pipette, inoculate it into bran medium, incubate at 20 ° C for 96 hours, add 500 mL of sterile physiological saline to the cultured seed koji culture flask, shake well, and shake well. Filter with heavy gauze, add 500 mL of sterile physiological saline to the filter residue, wash once and filter, mix the two filtrates to make a kekabi spore suspension, and prepare the kekabi spore suspension 4 Step 2 of putting in a refrigerator at ℃ and
Step 3 of selecting soybeans of uniform size in whole grains, weighing, washing, mixing with water at a volume ratio of 1: 3, draining, steaming at 121 ° C. for 30 minutes in a high temperature autoclave, and cooling. When,
Soybeans that have undergone the above step 3 are inoculated with a kikurage polysaccharide holding solution having a molecular weight cutoff of 300 kDa and a mucor spore suspension at a mass ratio of 1: 3, cultured at 25 ° C. for 48 hours, and then aged for 48 hours to produce mucor. Type - Kikurage polysaccharide 300k Da Tempeh fermented product, mixed inoculation step 4 and
A method for preparing a composition for treating hair loss, which is performed according to the above.
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