TW202339782A - Method for preparing ferment of chinese herbal medicine and skin complexion improved composition - Google Patents
Method for preparing ferment of chinese herbal medicine and skin complexion improved composition Download PDFInfo
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- TW202339782A TW202339782A TW111120415A TW111120415A TW202339782A TW 202339782 A TW202339782 A TW 202339782A TW 111120415 A TW111120415 A TW 111120415A TW 111120415 A TW111120415 A TW 111120415A TW 202339782 A TW202339782 A TW 202339782A
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Abstract
Description
本發明是關於一種發酵物的製備方法,特別是關於一種中草藥發酵物的製備方法以及包含中草藥發酵物之改善皮膚狀態的組合物。The present invention relates to a method for preparing a fermented product, in particular to a method for preparing a Chinese herbal fermented product and a composition containing the Chinese herbal fermented product for improving skin condition.
根據古代藥典記載,將多種不同的中藥材混合後塗抹於臉上可具有去除粉刺、黑斑、面皰及細毛的效果,然而,傳統中藥在取用上較為困難,費用較為可觀,且藥材內的活性成分取得亦較為不易。再者,常規的中藥係以煎煮方式萃取其有效成分,藥材煎煮物的顏色較深且觸感不佳,且萃取藥材中的活性成分時所需耗用的資源甚多,萃取效率亦不甚理想,致使運用中藥材之美容品、化妝品並不普及,目前也鮮少有針對中藥古方應用於化妝品的功效研究。According to ancient pharmacopoeia records, mixing a variety of different Chinese medicinal materials and applying them on the face can have the effect of removing acne, dark spots, blisters and fine hairs. However, traditional Chinese medicine is more difficult to obtain, the cost is considerable, and the ingredients in the medicinal materials It is also difficult to obtain active ingredients. Furthermore, traditional Chinese medicine extracts its active ingredients by decoction. The decoction of medicinal materials is darker in color and has a poor texture. In addition, a lot of resources are consumed to extract the active ingredients in the medicinal materials, and the extraction efficiency is also low. This is so ideal that beauty products and cosmetics using Chinese medicinal materials are not popular. Currently, there are few studies on the efficacy of ancient Chinese medicine prescriptions in cosmetics.
因此,如何提供一種能將中藥材有效地應用於美容護膚之相關領域的策略,實為相關學界及業界所致力發展的目標。Therefore, how to provide a strategy that can effectively apply Chinese medicinal materials in related fields of beauty and skin care is actually the development goal of the relevant academic circles and industry.
本發明之一態樣在於提供一種中草藥發酵物的製備方法,包含下述步驟。提供一發酵基質,其中所述之發酵基質包含一中草藥原料與水,所述之中草藥原料由黃耆、白朮、白蘞、黃精、川芎、杏仁、防風所組成。提供一發酵菌液,其中所述之發酵菌液包含一酵母菌。進行一發酵步驟,其係混合發酵基質與發酵菌液,以得一混合物,並將所述之混合物於20°C至50°C培養10小時至50小時,以得一發酵後產物。去除發酵後產物之一固體成分,以得一中草藥發酵物。One aspect of the present invention is to provide a preparation method of Chinese herbal medicine fermentation product, which includes the following steps. A fermentation substrate is provided, wherein the fermentation substrate includes a Chinese herbal medicine raw material and water, and the Chinese herbal medicine raw material is composed of Astragalus baicalensis, Atractylodes rhizome, Atractylodes baicalensis, Polygonatum sibiricum, Ligusticum chuanxiong, almond, and Fangfeng. A fermentation bacteria liquid is provided, wherein the fermentation bacteria liquid contains a yeast. A fermentation step is performed, which involves mixing the fermentation substrate and the fermentation bacterial liquid to obtain a mixture, and culturing the mixture at 20°C to 50°C for 10 hours to 50 hours to obtain a fermentation product. One solid component of the fermentation product is removed to obtain a Chinese herbal medicine fermentation product.
依據前述實施方式之中草藥發酵物的製備方法,其中所述之中草藥原料可呈一乾粉型式,且基於中草藥原料為100 wt%,黃耆的含量可為10~20 wt%、白朮的含量可為10~20 wt%、白蘞的含量可為10~20 wt%、黃精的含量可為15~30 wt%、川芎的含量可為15~30 wt%、杏仁的含量可為8~15 wt%,防風的含量可為8~15 wt%。According to the preparation method of Chinese herbal medicine fermentation product in the aforementioned embodiment, the Chinese herbal medicine raw material can be in the form of a dry powder, and based on 100 wt% of the Chinese herbal medicine raw material, the content of Scutellaria baicalensis can be 10~20 wt%, and the content of Atractylodes rhizome can be 10~20 wt%. 10~20 wt%, the content of Baiji can be 10~20 wt%, the content of Polygonatum can be 15~30 wt%, the content of Ligusticum Chuanxiong can be 15~30 wt%, and the content of almond can be 8~15 wt %, the content of windproof can be 8~15 wt%.
依據前述實施方式之中草藥發酵物的製備方法,其中酵母菌可為釀酒酵母菌 ( Saccharomyces cerevisiae)。 According to the preparation method of Chinese herbal medicine fermentation product in the foregoing embodiment, the yeast may be Saccharomyces cerevisiae .
依據前述實施方式之中草藥發酵物的製備方法,其中於所述之發酵基質中,每100 mL的水中可含有1.15~20 g的中草藥原料。According to the preparation method of Chinese herbal medicine fermentation product in the aforementioned embodiment, the fermentation matrix may contain 1.15 to 20 g of Chinese herbal medicine raw materials per 100 mL of water.
依據前述實施方式之中草藥發酵物的製備方法,其中所述之發酵菌液的濃度可為10 5CFU/mL至10 8CFU/mL,且於所述之混合物中,每10 g的中草藥原料可含有5~32 mL的發酵菌液。 According to the preparation method of Chinese herbal medicine fermentation product in the above embodiment, the concentration of the fermentation bacterial liquid can be 10 5 CFU/mL to 10 8 CFU/mL, and in the mixture, every 10 g of Chinese herbal medicine raw materials can be Contains 5~32 mL of fermentation bacteria liquid.
依據前述實施方式之中草藥發酵物的製備方法,其中所述之中草藥發酵物的pH值可為4.5至6.8。According to the preparation method of the Chinese herbal medicine fermentation product in the aforementioned embodiment, the pH value of the Chinese herbal medicine fermentation product may be from 4.5 to 6.8.
依據前述實施方式之中草藥發酵物的製備方法,可更包含一滅菌步驟,其係對發酵後產物進行一滅菌處理,且所述之固體成分可利用一離心方式、一過濾方式或上述方式之組合而去除。According to the above embodiment, the preparation method of Chinese herbal medicine fermentation product may further include a sterilization step, which is to perform a sterilization treatment on the fermentation product, and the solid component may be centrifuged, filtered or a combination of the above methods And remove.
本發明之另一態樣是在於提供一種改善皮膚狀態的組合物,包含一有效劑量之一中草藥發酵物,其中所述之中草藥發酵物係利用如前段所述之中草藥發酵物的製備方法所製得。Another aspect of the present invention is to provide a composition for improving skin condition, including an effective dose of a Chinese herbal medicine fermentation product, wherein the Chinese herbal medicine fermentation product is prepared by using the preparation method of a Chinese herbal medicine fermentation product as described in the previous paragraph. have to.
依據前述實施方式之改善皮膚狀態的組合物,其中所述之改善皮膚狀態的組合物可具有一抗氧化活性。According to the composition for improving skin condition according to the aforementioned embodiment, the composition for improving skin condition may have an antioxidant activity.
依據前述實施方式之改善皮膚狀態的組合物,其中所述之改善皮膚狀態的組合物可用以提高皮膚之一保濕能力及/或調理皮膚之一皮脂分泌能力。According to the composition for improving skin condition according to the aforementioned embodiment, the composition for improving skin condition can be used to improve the moisturizing ability of the skin and/or regulate the sebum secretion ability of the skin.
藉此,本發明之中草藥發酵物的製備方法採用酵母菌對由黃耆、白朮、白蘞、黃精、川芎、杏仁、防風所組成之中草藥原料進行發酵,可有效地萃取黃耆、白朮、白蘞、黃精、川芎、杏仁、防風的有效成分並保留其活性,避免傳統的萃取方法所造成的活性成分流失及有機溶劑的危害,進而使本發明之中草藥發酵物的製備方法所製得之中草藥發酵物可應用於改善皮膚狀態的組合物或其他化妝品中,並具有抗氧化、保濕及調理皮膚之皮脂分泌能力等功效,使本發明之中草藥發酵物的製備方法及改善皮膚狀態的組合物具有優異的市場應用潛力。Thereby, the preparation method of the Chinese herbal medicine fermentation product of the present invention uses yeast to ferment the Chinese herbal medicine raw materials composed of Astragalus, Atractylodes, Bletilla, Polygonatum, Chuanxiong, Almond, and Fangfeng, which can effectively extract Astragalus, Atractylodes, The active ingredients of Bletilla chinensis, Polygonatum sibiricum, Ligusticum chuanxiong, almond and Fangfeng retain their activity, thereby avoiding the loss of active ingredients and the harm of organic solvents caused by traditional extraction methods, thereby making the preparation method of the Chinese herbal medicine fermentation product of the present invention Chinese herbal fermentation can be used in compositions or other cosmetics to improve skin condition, and has the effects of antioxidant, moisturizing and regulating sebum secretion of the skin. The preparation method of Chinese herbal fermentation and the combination of improving skin condition of the present invention The material has excellent market application potential.
下述將更詳細討論本發明各實施方式。然而,此實施方式可為各種發明概念的應用,可被具體實行在各種不同的特定範圍內。特定的實施方式是僅以說明為目的,且不受限於揭露的範圍。Various embodiments of the invention are discussed in greater detail below. However, the embodiments are applicable to various inventive concepts and may be embodied in various specific scopes. The specific embodiments are provided for illustrative purposes only and do not limit the scope of the disclosure.
[本發明之中草藥發酵物的製備方法][Preparation method of Chinese herbal medicine fermentation product of the present invention]
請參照第1圖,其係繪示本發明之中草藥發酵物的製備方法100的步驟流程圖。中草藥發酵物的製備方法100包含步驟110、步驟120、步驟130以及步驟140。Please refer to Figure 1, which is a step flow chart of the
步驟110為提供一發酵基質,其中發酵基質包含一中草藥原料與水,所述之中草藥原料由黃耆 (
Astragalus membranaceus)、白朮 (
Atractylodes macrocephala)、白蘞 (
Ampelopsis japonica)、黃精 (
Polygonatum sibiricum)、川芎 (
Ligusticum striatum)、杏仁 (
Prunus Amygdalus)、防風 (
Saposhnikovia divaricata)所組成。詳細而言,本發明之中草藥原料的成分靈感來自於四庫全書之《普濟方》的藥方「澤潤芳」,其中黃耆又稱黃芪,其具有補氣升陽、益衛固表、利水消腫、托瘡生肌等功效;白朮具有補氣健脾、燥濕利水、止汗、安胎等功效;白蘞具有清熱解毒、消癰散結、生肌止痛等功效;黃精具有滋腎潤肺、補脾益氣等功效;川芎具有活血行氣、祛風止痛等功效;杏仁又稱苦扁桃籽,其具有止咳平喘、潤腸通便等功效;而防風則具有發表散風、勝濕止痛、止痙、止瀉等功效。
再者,本發明之中草藥原料可呈一乾粉型式,且基於中草藥原料為100 wt%,黃耆的含量可為10~20 wt%、白朮的含量為10~20 wt%、白蘞的含量可為10~20 wt%、黃精的含量可為15~30 wt%、川芎的含量可為15~30 wt%、杏仁的含量可為8~15 wt%,而防風的含量可為8~15 wt%。詳細而言,本發明之中草藥原料在進行後續步驟前可先進行乾燥及研磨步驟,使其呈現乾粉的型式而增加反應面積,其中乾燥的方法可為曬乾法、陰乾法、烘乾法、真空凍乾法、遠紅外光乾燥法或微波乾燥法,而研磨的方法則可利用研缽、研杵或粉碎機等研磨器具進行,但本發明並不以此為限。具體而言,本發明之乾燥方法可以70°C至80°C進行烘乾,以獲得粒徑為30目至80目的乾粉。較佳地,乾粉的粒徑可為70目,以利於後續操作。Furthermore, the Chinese herbal medicine raw material of the present invention can be in the form of a dry powder, and based on 100 wt% of the Chinese herbal medicine raw material, the content of Scutellaria baicalensis can be 10~20 wt%, the content of Atractylodes rhizome can be 10~20 wt%, and the content of Atractylodes atractylodes can be 10~20 wt%. The content of polygonatum can be 10~20 wt%, the content of Polygonatum can be 15~30 wt%, the content of Ligusticum chuanxiong can be 15~30 wt%, the content of almond can be 8~15 wt%, and the content of Fangfeng can be 8~15 wt%. In detail, the Chinese herbal medicine raw materials of the present invention can be dried and ground before subsequent steps, so that they can be in the form of dry powder to increase the reaction area. The drying method can be sun drying, shade drying, oven drying, Vacuum freeze-drying method, far-infrared light drying method or microwave drying method, and the grinding method can be carried out by using grinding equipment such as mortar, pestle or pulverizer, but the present invention is not limited thereto. Specifically, the drying method of the present invention can dry at 70°C to 80°C to obtain dry powder with a particle size of 30 mesh to 80 mesh. Preferably, the particle size of the dry powder can be 70 mesh to facilitate subsequent operations.
再者,在所述之發酵基質中,每100 mL的水中可含有1.15~20 g的中草藥原料。或者,在所述之發酵基質中,每100 mL的水中可含有5 g的中草藥原料。或者,在所述之發酵基質中,每300 mL的水中可含有3~20 g的中草藥原料。Furthermore, the fermentation matrix may contain 1.15 to 20 g of Chinese herbal medicine raw materials per 100 mL of water. Alternatively, the fermentation matrix may contain 5 g of Chinese herbal medicine raw materials per 100 mL of water. Alternatively, the fermentation matrix may contain 3 to 20 g of Chinese herbal medicine raw materials per 300 mL of water.
步驟120為提供一發酵菌液,其中所述之發酵菌液包含一酵母菌。詳細而言,酵母菌可代謝發酵基質中的蛋白質、脂質及澱粉,從而獲得富含多醣、胺基酸或其他特定成分及特定比例的發酵物。再者,本發明所使用的酵母菌可為但不限於任何品系之釀酒酵母菌(
Saccharomyces cerevisiae)或其他品種之酵母菌。較佳地,本發明所使用的酵母菌可為寄存於臺灣生物資源保存及研究中心 (Bioresource Collection and Research Center,BCRC) (地址:臺灣新竹食品路331號),且寄存編號為BCRC 21815的釀酒酵母菌 (以下簡稱為「釀酒酵母菌21815」)。釀酒酵母菌21815的菌落為點狀,菌落表面邊緣光滑,菌落顏色呈不透明之乳白色,且酵母菌以單細胞形式存在。較佳地,發酵菌液的濃度可為10
5CFU/mL至10
8CFU/mL,但本發明並不以此為限。
步驟130為進行一發酵步驟,其係混合所述之發酵基質與所述之發酵菌液,以得一混合物,並將混合物於20°C至50°C培養10小時至50小時,以得一發酵後產物。詳細而言,在上述溫度範圍下,酵母菌具有最佳的發酵效率。若發酵時間過短,將使中草藥原料的有效成分無法充分釋出;反之,若發酵時間過長,則可能使其他雜菌滋生而影響發酵的品質。較佳地,混合物可於25°C培養40小時,但本發明並不以此為限。Step 130 is to perform a fermentation step, which involves mixing the fermentation substrate and the fermentation bacterial liquid to obtain a mixture, and cultivating the mixture at 20°C to 50°C for 10 hours to 50 hours to obtain a mixture. Products after fermentation. In detail, yeast has the best fermentation efficiency in the above temperature range. If the fermentation time is too short, the active ingredients of the Chinese herbal medicine raw materials will not be fully released; conversely, if the fermentation time is too long, other bacteria may breed and affect the quality of the fermentation. Preferably, the mixture can be cultured at 25°C for 40 hours, but the present invention is not limited thereto.
再者,於所述之混合物中,每10 g的中草藥原料可含有5~32 mL的發酵菌液,但本發明並不以此為限。或者,於所述之混合物中,每10 g的中草藥原料可含有28 mL的發酵菌液。或者,於所述之混合物中,發酵菌液與中草藥原料的混合比例可為5~10 mL:2~20 g。Furthermore, in the mixture, every 10 g of Chinese herbal medicine raw materials may contain 5 to 32 mL of fermentation bacterial liquid, but the present invention is not limited to this. Alternatively, in the mixture, every 10 g of Chinese herbal medicine raw materials may contain 28 mL of fermentation bacteria liquid. Alternatively, in the mixture, the mixing ratio of fermentation bacterial liquid and Chinese herbal medicine raw materials can be 5~10 mL:2~20 g.
步驟140為去除發酵後產物之一固體成分,以得一中草藥發酵物。詳細而言,在經過步驟110至步驟130的處理後,黃耆、白朮、白蘞、黃精、川芎、杏仁與防風的有效成分已全數進入發酵後產物中,而在將發酵後產物中的固體成分去除後,餘下之液體成分即為本發明之中草藥發酵物,其中中草藥發酵物的pH值可為4.5至6.8,黏度可為100 cP至800 cP,並可含有1.0 wt%至5.0 wt%的可溶性固形物,且每毫升的中草藥發酵物可包含1.0~5.0 mg的蛋白質、5.0~10.0 mg的總多醣、1.0~1.8 mg總黃酮及0.1~0.6 mg的總酚。
另外,本發明之中草藥發酵物的製備方法100可更包含一滅菌步驟,其係對發酵後產物進行一滅菌處理。詳細而言,固體成分可利用離心方式、過濾方式或上述方式之組合而去除,滅菌步驟可為高壓蒸汽處理、乾熱處理或紫外線處理,但本發明並不以此為限。具體而言,發酵後產物可利用離心轉速為4000 rpm至12000 rpm的條件離心15分鐘至30分鐘後收集上清液,所述之上清液即為本發明之中草藥發酵物。或者,發酵後產物可利用離心轉速為4500 rpm的條件離心20分鐘。In addition, the
藉此,本發明之中草藥發酵物的製備方法100採用酵母菌對本發明之中草藥原料進行發酵,不僅可有效地萃取黃耆、白朮、白蘞、黃精、川芎、杏仁與防風的有效成分並保留其活性,更可縮短發酵的時間而避免長時間發酵造成的雜菌污染問題。再者,本發明之中草藥發酵物的製備方法100在萃取過程中不添加任何有機試劑,可避免傳統的萃取方法所造成的活性成分流失及有機溶劑殘留的危害,且萃取而得之中草藥發酵物沒有現有之中草藥萃取物之色澤過深的問題,進而使本發明之中草藥發酵物的製備方法100所製得之中草藥發酵物具有良好的生物安全性與市場美感,而具有優異的市場應用潛力。Thereby, the
[本發明之改善皮膚狀態的組合物][Composition for improving skin condition of the present invention]
本發明之改善皮膚狀態的組合物包含一有效劑量之本發明之中草藥發酵物。中草藥發酵物係以本發明之中草藥發酵物的製備方法所製得並具有抗氧化活性與提高皮膚之保濕能力、調理皮膚之皮脂分泌能力的效果,是以本發明之改善皮膚狀態的組合物具有優異的抗氧化、保濕、控油等功效而具有絕佳的市場應用潛力。The composition for improving skin condition of the present invention contains an effective dose of the fermented Chinese herbal medicine of the present invention. The fermented Chinese herbal medicine is prepared by the preparation method of the fermented Chinese herbal medicine of the present invention and has antioxidant activity and the effect of improving the moisturizing ability of the skin and regulating the sebum secretion of the skin. Therefore, the composition for improving the skin condition of the present invention has It has excellent antioxidant, moisturizing, oil control and other functions and has excellent market application potential.
另外,本發明之改善皮膚狀態的組合物之劑型可為液劑、乳劑、霜劑、安瓿、膠囊、丸劑、錠劑、粉末、顆粒或凝膠,並可應用於乳液、化妝水、精華液、隔離霜、洗面乳、面膜或防曬乳上,但本發明並不以此為限。In addition, the dosage form of the composition for improving skin condition of the present invention can be liquid, emulsion, cream, ampoule, capsule, pill, lozenge, powder, granule or gel, and can be applied to lotion, lotion, essence. , isolation cream, facial cleanser, facial mask or sunscreen, but the invention is not limited thereto.
[實施例與比較例][Examples and Comparative Examples]
一、本發明之中草藥發酵物1. Chinese herbal fermentation product of the present invention
以下將以實施例1至實施例4的中草藥發酵物說明本發明之中草藥發酵物的製備方法所製得之中草藥發酵物的性質,並進一步分析其抗氧化活性以及應用於皮膚保濕、調理皮膚之皮脂分泌能力等效果,而實施例1至實施例4的中草藥發酵物的製備原料請參表一。
實施例1至實施例4的中草藥發酵物是以下述步驟製得。首先,將自中藥房購得之黃耆、白朮、白蘞、黃精、川芎、杏仁與防風於75±5°C的烘箱中分別進行烘乾處理,再利用粉碎機進行研磨而得大小為50目的中草藥乾粉,並將15 g的中草藥乾粉及300 mL的水混合成懸浮液後作為本發明之發酵基質,以和包含酵母菌的發酵菌液混合進行發酵。The Chinese herbal medicine fermentation products of Examples 1 to 4 were prepared by the following steps. First, astragalus, Atractylodes, Atractylodes, Polygonatum, Chuanxiong, almonds and Fangfeng purchased from a Chinese medicine pharmacy were dried in an oven at 75±5°C, and then ground using a grinder to obtain a size of 50-mesh Chinese herbal medicine dry powder, and 15 g of Chinese herbal medicine dry powder and 300 mL of water are mixed into a suspension as the fermentation substrate of the present invention, and mixed with a fermentation bacterial liquid containing yeast for fermentation.
在將酵母菌用於進行發酵之前將先進行一預處理步驟。在預處理步驟方面,首先將釀酒酵母菌21815接種於馬鈴薯葡萄糖瓊脂培養基 (potato dextrose agar, PDA)中並於30°C的培養箱中活化,而後挑選單一菌落進行後續的擴增,並以馬鈴薯葡萄糖瓊脂培養水溶液 (potato dextrose water, PDW)於20°C至35°C的條件進行液態培養至OD值 = 0.5~1.0,此時的酵母菌將處於對數期而具有較佳的活性,即可用於後續之發酵。A pretreatment step is performed before the yeast is used for fermentation. In terms of pretreatment steps, Saccharomyces cerevisiae 21815 was first inoculated into potato dextrose agar (PDA) and activated in an incubator at 30°C. Then a single colony was selected for subsequent amplification, and potato was used Liquid culture the glucose agar culture solution (potato dextrose water, PDW) at 20°C to 35°C until the OD value = 0.5~1.0. At this time, the yeast will be in the logarithmic phase and have better activity, and it can be used for subsequent fermentation.
接著,將5 mL擴增培養後的發酵菌液接種至發酵基質中並均勻混合後,於25°C的培養箱中培養40小時,而後將所得之發酵後產物以121°C、1.1 kg/cm 2的高壓蒸氣進行滅菌,再將滅菌後的發酵後產物以4500 rpm、離心半徑為9公分的條件離心20分鐘後收集上清液,此上清液即為本發明之中草藥發酵物。 Next, 5 mL of the amplified fermentation culture was inoculated into the fermentation matrix and mixed evenly, then cultured in an incubator at 25°C for 40 hours, and then the obtained fermentation product was heated at 121°C, 1.1 kg/ cm 2 of high-pressure steam for sterilization, and then the sterilized fermentation product was centrifuged for 20 minutes at 4500 rpm and a centrifugal radius of 9 cm, and then the supernatant was collected. This supernatant is the Chinese herbal medicine fermentation product of the present invention.
實施例1至實施例4的中草藥發酵物在外觀上呈現無色至棕色的液體且觸感佳,而利用習知的酸鹼度或黏度測定法 (如酸鹼度計定法及旋轉黏度測定計)測得其pH值為4.5至6.8,黏度為100 cP至800 cP並含有0.9~2.0 wt%的可溶性固形物,且無致病菌檢出,具有高度的生物安全性而符合化妝品的品質要求。The Chinese herbal medicine fermentation products of Examples 1 to 4 appear as colorless to brown liquids with good touch, and their pH is measured using conventional pH or viscometer methods (such as pH meter and rotational viscometer). The value is 4.5 to 6.8, the viscosity is 100 cP to 800 cP and contains 0.9~2.0 wt% soluble solids, and no pathogenic bacteria are detected. It has a high degree of biological safety and meets the quality requirements of cosmetics.
二、本發明之中草藥發酵物的成分分析2. Component analysis of the fermented Chinese herbal medicine of the present invention
本試驗是利用高效液相層析 (HPLC)並參照試驗方法GB/T 30483-2013來分析實施例1至實施例4的中草藥發酵物的成分。而由高效液相層析的結果顯示,每毫升的實施例1至實施例4的中草藥發酵物皆含有1.0~5.0 mg的蛋白質、5.0~10.0 mg的總多醣、1.0~1.8 mg總黃酮及0.1~0.6 mg的總酚。再者,由於實施例1至實施例4的中草藥發酵物是以相同的製造方法所製得,其差異僅在於中草藥原料中的各組份比例與發酵菌液的濃度不同,且實施例1至實施例4的中草藥發酵物的蛋白質、總多醣、總黃酮及總酚含量相仿,故以下僅以實施例1的中草藥發酵物進行後續之不同有效成分之含量及其效果的分析,以說明本發明之中草藥發酵物應用於改善皮膚狀態的組合物的效果,特此敘明。This test uses high performance liquid chromatography (HPLC) and refers to the test method GB/T 30483-2013 to analyze the components of the Chinese herbal medicine fermentation products of Examples 1 to 4. The results of high performance liquid chromatography show that each milliliter of the Chinese herbal medicine fermentation products of Examples 1 to 4 contains 1.0~5.0 mg of protein, 5.0~10.0 mg of total polysaccharides, 1.0~1.8 mg of total flavonoids and 0.1 ~0.6 mg of total phenols. Furthermore, since the Chinese herbal medicine fermentation products of Examples 1 to 4 are produced by the same manufacturing method, the only difference lies in the proportion of each component in the Chinese herbal medicine raw materials and the concentration of the fermentation bacteria liquid, and the fermentation products of Examples 1 to 4 are different. The Chinese herbal medicine fermentation product of Example 4 has similar protein, total polysaccharide, total flavonoids and total phenolic contents. Therefore, the following only uses the Chinese herbal medicine fermentation product of Example 1 to perform subsequent analysis of the contents of different active ingredients and their effects to illustrate the present invention. The effect of using fermented Chinese herbal medicine in a composition for improving skin condition is hereby described.
另外,以下之試驗可同時包含比較例1與比較例2。比較例1是以本發明之中草藥發酵物的製備方法進行發酵物的製備 (以下稱為「比較例1的中草藥發酵物」),其差異僅在於比較例1的中草藥原料中各成分的比例與實施例1並不相同,而比較例1的中草藥原料中各成分的比例請參表二。比較例2係以傳統的煎煮方式處理本發明之中草藥原料而未經過任何發酵步驟(以下稱為「比較例2的中草藥水萃物」),且比較例2的中草藥原料成分比例與實施例1相同,以進一步分析以本發明之中草藥發酵物的製備方法所製得之中草藥發酵物的有效成分含量。
1. 本發明之中草藥發酵物的蛋白質含量分析1. Analysis of protein content of Chinese herbal medicine fermentation products of the present invention
在經過高效液相層析分析實施例1至實施例4的中草藥發酵物的成分後,本試驗進一步分析實施例1與比較例1在蛋白質含量方面的差異。After analyzing the components of the Chinese herbal medicine fermentation products of Examples 1 to 4 through high-performance liquid chromatography, this test further analyzed the difference in protein content between Example 1 and Comparative Example 1.
在實驗方面,首先精密量取0.0 mL、0.2 mL、0.4 mL、0.6 mL、0.8 mL、1.0 mL的牛血清蛋白標準溶液分別置於10 mL的試管中並分別加水補至1.0 mL,並於試管中加入1.0 mL的鹼性銅試液搖勻後於室溫放置10分鐘。接著,加入4.0 mL之福林酚試液 (2 mol/L 酸濃度)混勻並於室溫放置30分鐘後,使用分光光度計測量於波長650 nm的吸光值,並依據所得數值繪製標準曲線。接著,量取1 mL之實施例1的中草藥發酵物與比較例1的中草藥發酵物分別置於10 mL的試管中,依照製備標準曲線時的實驗方式進行實驗並測定波長650 nm的吸光值,並根據標準曲線換算為樣品中蛋白質的濃度。In terms of experiments, first accurately measure 0.0 mL, 0.2 mL, 0.4 mL, 0.6 mL, 0.8 mL, and 1.0 mL of bovine serum albumin standard solutions into 10 mL test tubes, add water to make up to 1.0 mL, and place in the test tubes. Add 1.0 mL of alkaline copper test solution to the solution, shake well, and place at room temperature for 10 minutes. Then, add 4.0 mL of folinol test solution (2 mol/L acid concentration), mix well and let it stand at room temperature for 30 minutes. Use a spectrophotometer to measure the absorbance value at a wavelength of 650 nm, and draw a standard curve based on the obtained value. Next, measure 1 mL of the Chinese herbal medicine fermentation product of Example 1 and the Chinese herbal medicine fermentation product of Comparative Example 1 and place them in 10 mL test tubes respectively. Conduct the experiment according to the experimental method for preparing the standard curve and measure the absorbance value at a wavelength of 650 nm. And converted to the concentration of protein in the sample according to the standard curve.
請參照表三,其係實施例1的中草藥發酵物與比較例1的中草藥發酵物的蛋白質含量分析結果。由表三的內容可知,每毫升之實施例1的中草藥發酵物含有1.706 mg的蛋白質,相較於比較例1的中草藥發酵物的蛋白質含量為1.404 mg為佳。
2. 本發明之中草藥發酵物的多醣含量分析2. Analysis of polysaccharide content of Chinese herbal medicine fermentation products of the present invention
在經過高效液相層析分析實施例1至實施例4的中草藥發酵物的成分後,本試驗進一步分析實施例1與比較例1在多醣含量方面的差異。After analyzing the components of the Chinese herbal medicine fermentation products of Examples 1 to 4 through high-performance liquid chromatography, this test further analyzed the difference in polysaccharide content between Example 1 and Comparative Example 1.
在實驗方面,首先精密量取0.2 mL、0.4 mL、0.6 mL、0.8 mL、1.0mL的葡萄糖標準溶液 (90 μg/mL)分別置10 mL的試管中並分別加水補至1.0 mL,並於試管中加入1 mL的5%苯酚溶液搖勻後,再加入5 mL的濃硫酸搖勻後於沸水中水浴20分鐘。接著,將樣品冰浴冷卻5分鐘後使用分光光度計測量於波長488 nm的吸光值,並依據所得數值繪製標準曲線。接著,量取1 mL之實施例1的中草藥發酵物與比較例1的中草藥發酵物分別置於10 mL的試管中,進一步依照製備標準曲線時的實驗方式進行實驗而測定波長488 nm的吸光值,並根據前述之標準曲線以及式(I)換算為樣品中多醣的含量,式(I)如下。
請參照表四,其係實施例1的中草藥發酵物與比較例1的中草藥發酵物的多醣含量分析結果。由表四的內容可知,每毫升之實施例1的中草藥發酵物含有7.069 mg的多醣,相較於比較例1的中草藥發酵物的多醣含量為4.629 mg為佳。
3. 本發明之中草藥發酵物的總酚含量分析3. Analysis of total phenolic content of Chinese herbal medicine fermentation products of the present invention
在經過高效液相層析分析實施例1至實施例4的中草藥發酵物的成分後,本試驗進一步分析實施例1與比較例1在總酚含量方面的差異。After analyzing the components of the Chinese herbal medicine fermentation products of Examples 1 to 4 through high-performance liquid chromatography, this test further analyzed the difference in total phenolic content between Example 1 and Comparative Example 1.
在實驗方面,首先以去離子水進行稀釋而配製濃度為10 μg/mL、20 μg/mL、30 μg/mL、40 μg/mL、50 μg/mL的沒食子酸溶液,以作為標準曲線之用,並另配置7.5%的Na 2CO 3溶液作為實驗試劑。接著,取1 mL的實施例1的中草藥發酵物與比較例1的中草藥發酵物分別與5 mL的10 mol/L%的福林酚試劑混合均勻後,再加入4 mL的7.5% Na 2CO 3混合均勻,於室溫下避光靜置1小時後,使用分光光度計測定其於波長765 nm的吸光值,並將所得數據依標準曲線換算成沒食子酸的當量,而後依照式(II)計算總酚含量,式(II)如下: X = C × 10 × n 式(II); 其中X為樣品中總酚的含量 (mg/L),C為由標準曲線計算而得之待測液中總酚的含量 (mg/L),而n則為樣品的稀釋倍數。 In terms of experiments, first dilute with deionized water to prepare gallic acid solutions with concentrations of 10 μg/mL, 20 μg/mL, 30 μg/mL, 40 μg/mL, and 50 μg/mL as a standard curve. For this purpose, a 7.5% Na 2 CO 3 solution is also prepared as an experimental reagent. Next, take 1 mL of the Chinese herbal medicine fermentation product of Example 1 and the Chinese herbal medicine fermentation product of Comparative Example 1 and mix them evenly with 5 mL of 10 mol/L% folinol reagent, and then add 4 mL of 7.5% Na 2 CO 3. Mix evenly and let it stand for 1 hour in the dark at room temperature. Use a spectrophotometer to measure the absorbance value at a wavelength of 765 nm. Convert the obtained data into the equivalent of gallic acid according to the standard curve, and then use the formula ( II) Calculate the total phenolic content, the formula (II) is as follows: X = C × 10 × n Formula (II); where The total phenol content in the test solution (mg/L), and n is the dilution factor of the sample.
請參照表五,其係實施例1的中草藥發酵物與比較例1的中草藥發酵物的總酚含量分析結果。由表五的內容可知,每毫升之實施例1的中草藥發酵物含有0.422 mg的總酚,相較於比較例1的中草藥發酵物的總酚含量為0.235 mg為佳。
4. 本發明之中草藥發酵物的胺基酸含量分析4. Analysis of amino acid content of Chinese herbal medicine fermentation products of the present invention
在經過高效液相層析分析實施例1至實施例4的中草藥發酵物的成分後,本試驗進一步分析實施例1與比較例1在胺基酸含量方面的差異。After analyzing the components of the Chinese herbal medicine fermentation products of Examples 1 to 4 through high-performance liquid chromatography, this test further analyzed the difference in amino acid content between Example 1 and Comparative Example 1.
在實驗方面,首先精密量取0.0 mL、0.2 mL、0.4 mL、0.6 mL、0.8 mL、1.0 mL的胺基酸標準溶液 (相當於0 μg、40 μg、80 μg、120 μg、160μg、200 μg的胺基酸)分別置於25 mL的試管中並加水補至4 mL,再分別加入1 mL之2%茚三酮溶液和磷酸鹽緩衝液 (pH = 8.04)搖勻後於沸水中水浴15分鐘。接著,將樣品冰浴冷卻至室溫並靜置15分鐘後,使用分光光度計測量於波長570 nm的吸光值,並依據所得數值繪製標準曲線。接著,量取1 mL之實施例1的中草藥發酵物與比較例1的中草藥發酵物分別置於25 mL的試管中,依照製備標準曲線時的實驗方式進行實驗而測定波長570 nm的吸光值,並根據標準曲線以及式(III)換算為樣品中胺基酸的含量,式(III)如下。
請參照表六,其係實施例1的中草藥發酵物與比較例1的中草藥發酵物的胺基酸含量分析結果。由表六的內容可知,每毫升之實施例1的中草藥發酵物含有0.671 mg的胺基酸,相較於比較例1的中草藥發酵物的胺基酸含量為0.293 mg為佳。
5. 本發明之中草藥發酵物的總黃酮含量分析5. Analysis of total flavonoid content of Chinese herbal medicine fermentation products of the present invention
在經過高效液相層析分析實施例1至實施例4的中草藥發酵物的成分後,本試驗進一步分析實施例1與比較例1在總黃酮含量方面的差異。After analyzing the components of the Chinese herbal medicine fermentation products of Examples 1 to 4 through high-performance liquid chromatography, this test further analyzed the difference in total flavonoid content between Example 1 and Comparative Example 1.
在實驗方面,首先精密吸取0.0 mL、1.0 mL、2.0 mL、3.0 mL、4.0 mL、5.0 mL、6.0 mL的蘆丁標準儲備液 (純度 ≥ 90.0%)分別置於25 mL容量瓶中並加水至6 mL,接著依序加入1 mL的5%亞硝酸鈉溶液 (靜置反應6分鐘)、1 mL的10%硝酸鋁溶液 (靜置反應6分鐘)與10 mL的4.3%氫氧化鈉試液 (靜置反應15分鐘)後,以配製濃度為0.0 μg/mL、8.0 μg/mL、16 μg/mL、24 μg/mL、32 μg/mL、40 μg/mL、48 μg/mL的蘆丁溶液,以作為標準曲線之用,並另配製10%的硝酸鋁溶液作為實驗試劑。接著,分別取2 mL的實施例1的中草藥發酵物與比較例1的中草藥發酵物並加水至6 mL,接著依序加入1 mL的5%亞硝酸鈉溶液 (靜置反應6分鐘)、1 mL的10%硝酸鋁溶液 (靜置反應6分鐘)與10 mL的4.3%氫氧化鈉試液 (靜置反應15分鐘)後,使用分光光度計測定其於波長510 nm的吸光值,並將所得數據依標準曲線換算成蘆丁的當量,而後依照式(IV)計算總黃酮含量,式(IV)如下:
請參照表七,其係實施例1的中草藥發酵物與比較例1的中草藥發酵物的總黃酮含量分析結果。由表七的內容可知,每毫升之實施例1的中草藥發酵物含有0.197 mg的總黃酮,相較於比較例1的中草藥發酵物的總黃酮含量為0.104 mg為佳。
6. 本發明之中草藥發酵物的麩胱甘肽含量分析6. Analysis of glutathione content of the Chinese herbal medicine fermentation product of the present invention
麩胱甘肽 (glutathione)又稱穀胱甘肽或麩胺基硫,其是由麩胺酸、半胱胺酸及甘胺酸等3種胺基酸所組成的小分子蛋白質,為人體之抗氧化酵素榖胱甘肽過氧化酶 (Glutathione Peroxidase, GPx)的重要成分,可幫助人體細胞對抗自由基,降低疾病的產生。因此,本試驗進一步以高效液相層析分析實施例1的中草藥發酵物與比較例2的中草藥水萃物的麩胱甘肽含量,而分析麩胱甘肽含量的高效液相層析的檢測條件列示於表八。
請參照表九,其係實施例1的中草藥發酵物與比較例2的中草藥水萃物的麩胱甘肽含量分析結果。由表九的內容可知,每毫升之實施例1的中草藥發酵物含有10.94 μg的麩胱甘肽,相較於比較例2的中草藥水萃物的麩胱甘肽含量具有30%的提升。
7. 本發明之中草藥發酵物的鞣花酸含量分析7. Analysis of ellagic acid content of the Chinese herbal medicine fermentation product of the present invention
鞣花酸 (Ellagic acid
)是存在於眾多水果和蔬菜中的天然酚類抗氧化劑,尤以黑莓、草莓、石榴、枸杞、覆盆子、白橡實、蔓越莓、山核桃等植物的果實中含量最為豐富,而鞣花酸的抗癌細胞增生與抗氧化活性使其廣泛的被應用於各領域中。因此,本試驗進一步以高效液相層析分析實施例1的中草藥發酵物與比較例2的中草藥水萃物的鞣花酸含量,而分析鞣花酸含量的高效液相層析的檢測條件列示於表十。
請參照表十一,其係實施例1的中草藥發酵物與比較例2的中草藥水萃物的鞣花酸含量分析結果。由表十一的內容可知,每毫升之實施例1的中草藥發酵物含有245.281 μg的鞣花酸,相較於比較例2的中草藥水萃物的鞣花酸含量具有86.19%的提升。
8. 本發明之中草藥發酵物的阿魏酸含量分析8. Analysis of ferulic acid content of Chinese herbal medicine fermentation products of the present invention
阿魏酸 (Ferulic acid)是桂皮酸的衍生物之一,是存在於植物細胞壁中的一種豐富的酚類,其具有清除自由基、抑制酪氨酸酶活性等功效。因此,本試驗進一步以高效液相層析分析實施例1的中草藥發酵物與比較例2的中草藥水萃物的阿魏酸含量,而分析阿魏酸含量的高效液相層析的檢測條件列示於表十二。
請參照表十三,其係實施例1的中草藥發酵物與比較例2的中草藥水萃物的阿魏酸含量分析結果。由表十三的內容可知,每毫升之實施例1的中草藥發酵物含有2.667 μg的阿魏酸,相較於比較例2的中草藥水萃物的阿魏酸含量具有71.62%的提升。
9. 本發明之中草藥發酵物的升麻素苷含量分析9. Analysis of Cimicin Glycoside Content of Chinese Herbal Medicine Fermentation Products of the Invention
研究顯示,防風等藥材中的升麻素苷 (prim-O-glucosylcimifugin)具有解熱、鎮痛、抗炎、消腫、抗血小板凝結等作用,是以本試驗進一步以高效液相層析分析實施例1的中草藥發酵物與比較例2的中草藥水萃物的升麻素苷含量,而分析升麻素苷含量的高效液相層析的檢測條件列示於表十四。
請參照表十五,其係實施例1的中草藥發酵物與比較例2的中草藥水萃物的升麻素苷含量分析結果。由表十五的內容可知,每毫升之實施例1的中草藥發酵物含有6.289 μg的升麻素苷,相較於比較例2的中草藥水萃物的升麻素苷含量具有73.44%的提升。
10. 本發明之中草藥發酵物的毛蕊異黃酮葡萄糖苷含量分析10. Analysis of the callisoflavone glucoside content of the Chinese herbal medicine fermentation product of the present invention
黃耆包含有效成分毛蕊異黃酮葡萄糖苷 (calycosin-7-glucoside),使其具有補氣、脫毒、生肌、利水的功效,是以本試驗進一步以高效液相層析分析實施例1的中草藥發酵物與比較例2的中草藥水萃物的毛蕊異黃酮葡萄糖苷含量,而分析毛蕊異黃酮葡萄糖苷含量的高效液相層析的檢測條件列示於表十六。
請參照表十七,其係實施例1的中草藥發酵物與比較例2的中草藥水萃物的毛蕊異黃酮葡萄糖苷含量分析結果。由表十七的內容可知,每毫升之實施例1的中草藥發酵物含有131.254 μg的鞣花酸,相較於比較例2的中草藥水萃物的毛蕊異黃酮葡萄糖苷含量具有56.92%的提升。
由上述結果可見,本發明之中草藥發酵物的製備方法相較於傳統的萃取方法可更有效地萃取本發明之中草藥原料中所含有的麩胱甘肽、鞣花酸、阿魏酸、升麻素苷與毛蕊異黃酮葡萄糖苷,並可有效保留其活性,進而使本發明之中草藥發酵物的製備方法所製備之中草藥發酵物具有優異的市場應用潛力。It can be seen from the above results that the preparation method of the Chinese herbal medicine fermentation product of the present invention can more effectively extract glutathione, ellagic acid, ferulic acid, and cohosh contained in the Chinese herbal medicine raw materials of the present invention than the traditional extraction method. The fermented Chinese herbal medicine fermented product prepared by the preparation method of the Chinese herbal medicine fermented product of the present invention has excellent market application potential.
三、本發明之中草藥發酵物的抗氧化活性分析3. Analysis of antioxidant activity of Chinese herbal medicine fermentation products of the present invention
本試驗是利用芬頓 (Fenton)反應中過氧化氫及亞鐵離子反應所產生之羥自由基,並透過水楊酸捕獲前述反應所產生的羥自由基而產生紫色產物的特性來測量本發明之中草藥發酵物清除羥自由基的能力。詳細而言,由於前述之紫色產物對波長為510 nm的光線具有較大吸光值,若本發明之中草藥發酵物具有抗氧化功效,在將其加入含有過氧化氫、亞鐵離子及水楊酸的反應溶液後,可降低反應溶液對波長為510 nm的光線的吸光值,並可藉此評估本發明之中草藥發酵物的抗氧化活性。This test utilizes the hydroxyl radicals generated by the reaction of hydrogen peroxide and ferrous ions in the Fenton reaction, and uses salicylic acid to capture the hydroxyl radicals generated by the reaction to produce a purple product to measure the characteristics of the present invention. The ability of Chinese herbal fermentation products to scavenge hydroxyl radicals. In detail, since the aforementioned purple product has a large absorbance value for light with a wavelength of 510 nm, if the Chinese herbal fermentation product of the present invention has antioxidant effects, it is added with hydrogen peroxide, ferrous ions and salicylic acid. After using the reaction solution, the absorbance value of the reaction solution for light with a wavelength of 510 nm can be reduced, and the antioxidant activity of the Chinese herbal medicine fermentation product of the present invention can be evaluated.
在實驗方面分別取體積百分濃度為10%、15%、20%、25%與30%的實施例1的中草藥發酵物和與比較例2的中草藥水萃物與9 mmol/L的FeSO 4、8.8 mmol/L的H 2O 2和9 mmol/L的水楊酸乙醇溶液混合後,於37°C下水浴15分鐘並以分光亮度計測量各樣本於波長為510 nm的吸光值,並依照以下述式(V)計算羥自由基的清除率: X% = [1-(AU/AU 0)] × 100% 式(V); 其中X%表示羥自由基清除率,AU表示實施例1的相對吸光值,AU 0則表示比較例2的相對吸光值。 In terms of experiments, the Chinese herbal medicine fermentation product of Example 1 and the Chinese herbal medicine aqueous extract of Comparative Example 2 and 9 mmol/L FeSO 4 were taken with volume percentage concentrations of 10%, 15%, 20%, 25% and 30% respectively. After mixing 8.8 mmol/L H 2 O 2 and 9 mmol/L salicylic acid ethanol solution, place in a water bath at 37°C for 15 minutes and measure the absorbance value of each sample at a wavelength of 510 nm with a spectrophotometer, and Calculate the hydroxyl radical scavenging rate according to the following formula (V): X% = [1-(AU/AU 0 )] × 100% Formula (V); wherein The relative absorbance value of AU 0 represents the relative absorbance value of Comparative Example 2.
再者,本試驗亦包含對照例1與對照例2,其中對照例1與對照例2的吸光值是分別用以做為實施例1及比較例2的空白 (blank)背景值,以排除呈色反應以外的因素所造成的吸光值變化,從而獲得標準化後的相對吸光值。而實施例1、比較例2、對照例1與對照例2用以進行試驗的試劑配比請參表十八。
請參照第2圖,其係本發明之中草藥發酵物的羥自由基清除能力之分析圖。如第2圖所示,隨著體積百分濃度的增加,實施例1的中草藥發酵物的羥自由基清除率係逐漸提升,而當實施例1的中草藥發酵物的體積百分濃度為30%時,其羥自由基清除率為18.60%,相較於相同濃度之比較例2的中草藥水萃物具有較佳的表現,可見本發明之中草藥發酵物的製備方法所製得之中草藥發酵物相較於傳統的萃取方法所製得之萃取物具有較佳的抗氧化功效,並有潛力應用於改善皮膚狀態的組合物或其他化妝品之中而具有相關市場的應用潛力。Please refer to Figure 2, which is an analysis chart of the hydroxyl radical scavenging ability of the Chinese herbal medicine fermentation product of the present invention. As shown in Figure 2, as the volume percentage concentration increases, the hydroxyl radical scavenging rate of the Chinese herbal medicine fermentation product of Example 1 gradually increases, and when the volume percentage concentration of the Chinese herbal medicine fermentation product of Example 1 is 30% When, its hydroxyl radical scavenging rate is 18.60%, which is better than the Chinese herbal medicine aqueous extract of Comparative Example 2 with the same concentration. It can be seen that the Chinese herbal medicine fermentation product prepared by the preparation method of the Chinese herbal medicine fermentation product of the present invention is relatively good. Compared with the extract obtained by traditional extraction methods, it has better antioxidant effect and has the potential to be used in compositions for improving skin condition or other cosmetics and has application potential in related markets.
四、本發明之中草藥發酵物的保濕能力評估4. Evaluation of the moisturizing ability of the Chinese herbal fermentation product of the present invention
本試驗是利用經皮水分散失量 (trans-epidermal water loss,TEWL)做為皮膚表層含水量的指標,以評估實施例1的中草藥發酵物的保濕能力。另外,本試驗同樣包含前述之比較例2的中草藥水萃物,以進一步評估以本發明之中草藥發酵物的製備方法所製得之中草藥發酵物的保濕效果。This test uses transepidermal water loss (TEWL) as an indicator of the moisture content of the skin surface to evaluate the moisturizing ability of the Chinese herbal fermentation product in Example 1. In addition, this test also includes the Chinese herbal medicine aqueous extract of the aforementioned comparative example 2 to further evaluate the moisturizing effect of the Chinese herbal medicine fermentation product prepared by the preparation method of the Chinese herbal medicine fermentation product of the present invention.
在實驗方面,首先在3位受試者的手臂皮膚設定對照區域及實驗區域 (各為3 cm × 3 cm),並利用市售之經皮水分散失測定儀TM300 (Courage + Khazaka Electronic)檢測對照區域與實驗區域之皮膚的經皮水分散失量。接著,將使用量為2.0±1.0 mg/cm 2之實施例1的中草藥發酵物與比較例2的中草藥水萃物單次塗抹於實驗區域的皮膚上,但不對對照區域的皮膚進行任何處理,並於測試前、使用後第1天、使用後第5天、使用後第7天及使用後第14天測量經皮水分散失量,並將所測得之數據以對照區域的經皮水分散失量數據進行校正而得皮膚含水量的評估結果。 In terms of experiments, first, a control area and an experimental area (each 3 cm × 3 cm) were set on the arm skin of three subjects, and the control area was detected using a commercially available transepidermal water loss meter TM300 (Courage + Khazaka Electronic). Transepidermal water loss from the skin of the regional and experimental areas. Next, the Chinese herbal fermentation product of Example 1 and the Chinese herbal aqueous extract of Comparative Example 2 were applied to the skin of the experimental area in a single dose at a dosage of 2.0 ± 1.0 mg/cm 2 , but no treatment was performed on the skin of the control area. The transepidermal water loss was measured before the test, on the 1st day after use, on the 5th day after use, on the 7th day after use and on the 14th day after use, and the measured data were used as the transepidermal water loss in the control area. The skin moisture content assessment results are obtained by correcting the volume data.
請參照第3圖,其係本發明之中草藥發酵物的改善肌膚含水量的分析結果圖。如第3圖所示,在使用後第1天、使用後第5天、使用後第7天及使用後第14天,受試者使用實施例1的中草藥發酵物後的皮膚水分含量皆明顯高於比較例2的皮膚水分含量,顯示使用實施例1的中草藥發酵物後的經皮水分散失量較少。由此可見,本發明之中草藥發酵物的製備方法所製得之中草藥發酵物具有優異保濕功效並可提高皮膚之保濕能力,並有潛力應用於改善皮膚狀態的組合物或其他化妝品之中而具有相關市場的應用潛力。Please refer to Figure 3, which is an analysis result of the improvement of skin moisture content by the Chinese herbal medicine fermentation product of the present invention. As shown in Figure 3, on the 1st day after use, the 5th day after use, the 7th day after use, and the 14th day after use, the skin moisture content of the subjects after using the Chinese herbal fermentation product of Example 1 was all obvious. The skin moisture content is higher than that of Comparative Example 2, indicating that the amount of transcutaneous water loss after using the Chinese herbal fermentation product of Example 1 is less. It can be seen from this that the Chinese herbal fermentation product prepared by the preparation method of the Chinese herbal fermentation product of the present invention has excellent moisturizing effect and can improve the moisturizing ability of the skin, and has the potential to be used in compositions or other cosmetics to improve skin condition. Application potential in relevant markets.
五、本發明之中草藥發酵物的皮脂調理效果分析5. Analysis of the sebum conditioning effect of the Chinese herbal fermentation product of the present invention
本試驗是測試施用實施例1的中草藥發酵物後的皮膚油脂含量而評估其調理皮膚之皮脂分泌能力的效果。另外,本試驗同樣包含前述之比較例2的中草藥水萃物,以進一步評估以本發明之中草藥發酵物的製備方法所製得之中草藥發酵物的皮脂調理效果。This test is to test the oil content of the skin after applying the Chinese herbal fermentation product of Example 1 to evaluate its effect on regulating the sebum secretion of the skin. In addition, this test also includes the aqueous Chinese herbal medicine extract of Comparative Example 2 mentioned above to further evaluate the sebum conditioning effect of the Chinese herbal medicine fermentation product prepared by the preparation method of the Chinese herbal medicine fermentation product of the present invention.
在實驗方面,首先在3位受試者的額頭皮膚設定對照區域及實驗區域 (各為3 cm × 3 cm),並利用市售之皮膚油脂測試儀Sebumeter SM300 (Courage + Khazaka Electronic)檢測對照區域與實驗區域之皮膚的油脂含量。接著,將使用量為2.0±1.0 mg/cm 2之實施例1的中草藥發酵物與比較例2的中草藥水萃物單次塗抹於實驗區域的皮膚上,但不對對照區域的皮膚進行任何處理,並於測試前、使用後第1天、使用後第5天、使用後第7天及使用後第14天測量皮膚的油脂含量,並將所測得之數據以對照區域的皮膚的油脂含量數據進行校正而得皮膚的油脂含量差值的結果。 In terms of experiments, first, a control area and an experimental area (each 3 cm × 3 cm) were set on the forehead skin of three subjects, and the control area was detected using the commercially available skin oil tester Sebumeter SM300 (Courage + Khazaka Electronic). and the oil content of the skin in the experimental area. Next, the Chinese herbal fermentation product of Example 1 and the Chinese herbal aqueous extract of Comparative Example 2 were applied to the skin of the experimental area in a single dose at a dosage of 2.0 ± 1.0 mg/cm 2 , but no treatment was performed on the skin of the control area. The oil content of the skin was measured before the test, on the 1st day after use, on the 5th day after use, on the 7th day after use and on the 14th day after use, and the measured data were compared with the skin oil content data of the control area. The result of correction is the difference in skin oil content.
請參照第4圖,其係本發明之中草藥發酵物的皮脂調理效果的分析結果圖。如第4圖所示,在使用後第5天、使用後第7天及使用後第14天,受試者使用實施例1的中草藥發酵物後的皮膚油脂含量差值明顯低於比較例2的皮膚油脂含量差值,顯示實施例1的中草藥發酵物在施用於皮膚後具有較佳的皮脂調理效果,並可有效地調理皮膚之皮脂分泌能力,並有潛力應用於改善皮膚狀態的組合物或其他化妝品之中而具有相關市場的應用潛力。Please refer to Figure 4, which is an analysis result of the sebum conditioning effect of the Chinese herbal fermentation product of the present invention. As shown in Figure 4, on the 5th day after use, the 7th day after use, and the 14th day after use, the difference in skin oil content of the subjects after using the Chinese herbal fermentation product of Example 1 was significantly lower than that of Comparative Example 2. The difference in skin oil content shows that the Chinese herbal fermentation product of Example 1 has a better sebum conditioning effect after being applied to the skin, and can effectively regulate the sebum secretion ability of the skin, and has the potential to be used in compositions to improve skin condition. or other cosmetics and have application potential in relevant markets.
六、本發明之中草藥發酵物的抗敏舒緩效果評估6. Evaluation of the anti-allergic and soothing effect of the Chinese herbal fermentation product of the present invention
本試驗是將實施例1的中草藥發酵物塗抹於3位受試者之皮膚上,以觀察其上之紅血絲的消退情形而評估本發明之中草藥發酵物的抗敏舒緩效果。In this test, the Chinese herbal fermentation product of Example 1 was applied to the skin of three subjects to observe the fading of red blood streaks and evaluate the anti-allergic and soothing effect of the Chinese herbal medicine fermentation product of the present invention.
在實驗方面,首先在3位受試者的手臂皮膚設定實驗區域 (各為3 cm × 3 cm),並將使用量為2.0±1.0 mg/cm 2之實施例1的中草藥發酵物塗抹於實驗區域的皮膚上,每日塗抹2次並持續14天,並拍攝測試前與使用後第14天的皮膚影像及根據所得影像計算其紅血絲平均值。 In terms of experiments, first, an experimental area (each 3 cm × 3 cm) was set on the arm skin of three subjects, and the Chinese herbal fermentation product of Example 1 was applied to the experiment in an amount of 2.0 ± 1.0 mg/cm 2 Apply to the skin of the area twice a day for 14 days, and take skin images before the test and on the 14th day after use, and calculate the average redness value based on the images obtained.
請參照第5A圖、第5B圖、第6圖與表十九,其中第5A圖係本發明之中草藥發酵物的抗敏舒緩效果的一試驗影像,第5B圖係本發明之中草藥發酵物的抗敏舒緩效果的另一試驗影像,第6圖係本發明之中草藥發酵物的抗敏舒緩效果的分析結果圖,表十九則記錄受試者1與受試者2在未使用實施例1的中草藥發酵物與使用14天後的紅血絲平均值,以及受試者3在未使用實施例1的中草藥發酵物與使用7天、14天後的紅血絲平均值。
由第5A圖、第5B圖、第6圖與表十九的內容可見,在使用實施例1的中草藥發酵物14天後,受試者2與受試者3的皮膚紅腫現象明顯消退,其紅血絲平均值亦明顯降低,且3位受試者的皮膚紅血絲平均降低8.2%,顯示本發明之中草藥發酵物的製備方法所製得之中草藥發酵物具有相當的抗敏舒緩效果,並有潛力應用於改善皮膚狀態的組合物或其他化妝品之中而具有相關市場的應用潛力。It can be seen from Figure 5A, Figure 5B, Figure 6 and Table 19 that after 14 days of using the Chinese herbal fermentation product of Example 1, the skin redness and swelling of
七、本發明之中草藥發酵物的安全性評估7. Safety evaluation of the fermented Chinese herbal medicine of the present invention
本試驗是利用封閉式貼布試驗 (Closed Patch Test)評估實施例1的中草藥發酵物對皮膚的潛在刺激性,以確認本發明之中草藥發酵物應用於人體的安全性。在實驗方面,首先將0.2 mL至0.25 mL的實施例1的中草藥發酵物滴加於濾紙片上,再將濾紙片置於斑試器中備用。在受試者方面,隨機選擇年齡為18歲至60歲共30位受試者,並以無刺激性的膠帶於每位受試者的背部固定2個斑試器,使斑試器中的濾紙片貼附於受試者的背部皮膚上24小時,在試驗期間內不移除斑試器,也盡量避免斑試器沾染汗液或水。另外,每個試驗組別皆對應設定一控制組,控制組係以0.20 mL至0.25 mL的蒸餾水進行測試。在測試24小時後取下斑試器,並分別於取下斑試器30分鐘後、24小時後及48小時後觀察斑試器下的皮膚 (以下稱為「受試區域」)及受試區域周圍的皮膚是否出現紅斑、浸潤、水腫、丘疹及/或皰疹等過敏或發炎的症狀。This test uses a closed patch test to evaluate the potential irritation of the Chinese herbal fermentation product of Example 1 to the skin to confirm the safety of the Chinese herbal fermentation product of the present invention when applied to the human body. In terms of experiment, first, 0.2 mL to 0.25 mL of the Chinese herbal medicine fermentation product of Example 1 was dropped onto the filter paper piece, and then the filter paper piece was placed in a spot tester for later use. Regarding the subjects, a total of 30 subjects ranging in age from 18 to 60 years old were randomly selected, and 2 patch testers were fixed on the back of each subject with non-irritating tape, so that the The filter paper piece was attached to the subject's back skin for 24 hours. The spot tester was not removed during the test, and the spot tester was also tried to avoid being contaminated with sweat or water. In addition, each test group corresponds to a control group, and the control group is tested with 0.20 mL to 0.25 mL of distilled water. Remove the patch tester 24 hours after the test, and observe the skin under the patch tester (hereinafter referred to as the "test area") and the subject 30 minutes, 24 hours and 48 hours after removal of the patch tester. Whether the skin around the area shows symptoms of allergies or inflammation such as erythema, infiltration, edema, papules and/or herpes.
請參照表二十,其為實施例1的中草藥發酵物的安全性測試結果。安全性測試結果共分五個等級,其中圖號「-」代表受試區域為陰性反應而未有任何過敏或發炎的症狀,圖號「±」代表受試區域出現可疑反應 (如:具有輕微的紅斑),圖號「+」代表受試區域出現弱陽性反應 (亦稱為紅斑反應,即出現紅斑、浸潤、水腫及/或丘疹),圖號「++」代表受試區域出現強陽性反應 (亦稱為皰疹反應,即出現紅斑、浸潤、水腫、丘疹、皰疹,且皰疹反應超出受試區域以外的皮膚),圖號「+++」代表受試區域出現極強陽性反應 (亦稱為融合性皰疹反應,即出現明顯紅斑、嚴重浸潤、水腫、融合性丘疹,且融合性皰疹反應超出受試區域以外的皮膚)。另外,於表二十中同時計算反應程度為陽性反應 (即圖號為+、++與+++)的人數,以更清楚確認本發明之中草藥發酵物的製備方法所製得之中草藥發酵物的人體安全性。
如表二十所示,在取下斑試器30分鐘後、24小時後或48小時後,受試者在受試區域及其周圍的皮膚皆無紅斑、浸潤、水腫、丘疹、皰疹及融合性皰疹等反應,顯示本發明之中草藥發酵物的製備方法所製得之中草藥發酵物具有優異的生物安全性,並具有相關市場的應用潛力。As shown in Table 20, 30 minutes, 24 hours or 48 hours after removing the spot tester, the subject's skin in and around the test area showed no erythema, infiltration, edema, papules, herpes or fusion. Herpes and other reactions show that the Chinese herbal fermentation product prepared by the preparation method of the Chinese herbal medicine fermentation product of the present invention has excellent biological safety and has application potential in relevant markets.
需補充的是,本發明雖以特定的菌株、特定的材料、特定的比例、特定的製程、特定的分析方法或特定儀器做為例示,以說明本發明之中草藥發酵物具有抗氧化、保濕、調理皮膚之皮脂分泌能力等功效,惟本發明所屬技術領域中任何具有通常知識者可知,在不脫離本發明之精神和範圍內,本發明之中草藥發酵物的成分與功效亦可使用其他的菌株、其他的材料、其他的比例、其他的製程、其他分析方法或其他儀器進行分析,且本發明並不以此為限。It should be added that although the present invention uses specific strains, specific materials, specific ratios, specific processes, specific analysis methods or specific instruments as examples to illustrate that the Chinese herbal fermentation product of the present invention has antioxidant, moisturizing, and However, anyone with ordinary knowledge in the technical field to which the present invention belongs will know that the ingredients and functions of the Chinese herbal fermentation product of the present invention can also be used in other strains without departing from the spirit and scope of the present invention. , other materials, other proportions, other processes, other analysis methods or other instruments for analysis, and the present invention is not limited thereto.
綜上所述,本發明的中草藥原料由黃耆、白朮、白蘞、黃精、川芎、杏仁、防風所組成,而上述藥材在經本發明之中草藥發酵物的製備方法以酵母菌進行發酵後可得含有高濃度之活性成分的中草藥發酵物,且前述之有效成分在彼此協同作用下具有優異的抗氧化、保濕、調理皮膚之皮脂分泌能力等功效,並具有絕佳的生物安全性而可應用於改善皮膚狀態的組合物中,具有優異的市場應用潛力。To sum up, the Chinese herbal medicine raw materials of the present invention are composed of Astragalus, Atractylodes, Bletilla, Polygonatum, Ligusticum chuanxiong, almond, and Fangfeng, and the above-mentioned medicinal materials can be fermented by yeast through the preparation method of the Chinese herbal fermentation product of the present invention. A Chinese herbal ferment containing a high concentration of active ingredients is obtained, and the aforementioned active ingredients have excellent antioxidant, moisturizing, and regulating sebum secretion abilities of the skin under the synergistic effect of each other, and have excellent biological safety and can be used It has excellent market application potential in compositions for improving skin condition.
雖然本發明已以實施方式揭露如上,然其並非用以限定本發明,任何熟習此技藝者,在不脫離本發明之精神和範圍內,當可作各種之更動與潤飾,因此本發明之保護範圍當視後附之申請專利範圍所界定者為準。Although the present invention has been disclosed in the above embodiments, it is not intended to limit the present invention. Anyone skilled in the art can make various modifications and modifications without departing from the spirit and scope of the present invention. Therefore, the protection of the present invention is The scope shall be determined by the appended patent application scope.
100:中草藥發酵物的製備方法 110,120,130,140:步驟 100: Preparation method of Chinese herbal medicine fermentation products 110,120,130,140: steps
為讓本發明之上述和其他目的、特徵、優點與實施例能更明顯易懂,所附圖式之說明如下: 第1圖係繪示本發明之中草藥發酵物的製備方法的步驟流程圖; 第2圖係本發明之中草藥發酵物的羥自由基清除能力之分析圖; 第3圖係本發明之中草藥發酵物的改善肌膚含水量的分析結果圖; 第4圖係本發明之中草藥發酵物的皮脂調理效果的分析結果圖; 第5A圖係本發明之中草藥發酵物的抗敏舒緩效果的一試驗影像; 第5B圖係本發明之中草藥發酵物的抗敏舒緩效果的另一試驗影像;以及 第6圖係本發明之中草藥發酵物的抗敏舒緩效果的分析結果圖。 In order to make the above and other objects, features, advantages and embodiments of the present invention more apparent and understandable, the accompanying drawings are described as follows: Figure 1 is a flow chart showing the steps of the preparation method of Chinese herbal medicine fermentation product of the present invention; Figure 2 is an analysis chart of the hydroxyl radical scavenging ability of the Chinese herbal medicine fermentation product of the present invention; Figure 3 is a diagram showing the analysis results of the Chinese herbal medicine fermentation product of the present invention for improving skin moisture content; Figure 4 is a diagram showing the analysis results of the sebum conditioning effect of the Chinese herbal medicine fermentation product of the present invention; Figure 5A is a test image of the anti-allergic and soothing effect of the Chinese herbal fermentation product of the present invention; Figure 5B is another test image of the anti-allergic and soothing effect of the Chinese herbal ferment of the present invention; and Figure 6 is a graph showing the analysis results of the anti-allergic and soothing effect of the Chinese herbal fermentation product of the present invention.
100:中草藥發酵物的製備方法 100: Preparation method of Chinese herbal medicine fermentation products
110,120,130,140:步驟 110,120,130,140: steps
Claims (10)
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
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CN202210350255.5 | 2022-04-02 | ||
CN202210350255.5A CN116920046A (en) | 2022-04-02 | 2022-04-02 | Preparation method of Chinese herbal medicine fermentation product and composition for improving skin condition |
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