TWI715172B - Composition in suppressing body fat accumulation - Google Patents
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本發明係有關於一種組合物；更詳而言之，特別係關於一種具有抑制體脂肪形成之組合物。The present invention relates to a composition; more specifically, it particularly relates to a composition that inhibits body fat formation.
橙皮苷為柑橘類水果中的一種黃酮類化合物，其主要儲存於柑橘類水果之果皮中，且近年來有不少研究指出服用橙皮苷萃取物對於人體具有降血壓、降膽固醇和預防動脈硬化之效果，但卻未見有人採用發酵的方式使柑橘類水果中的橙皮苷含量增加，並使其功效不亞於橙皮苷萃取物。Hesperidin is a flavonoid in citrus fruits, which is mainly stored in the peel of citrus fruits. In recent years, many studies have pointed out that taking hesperidin extract can lower blood pressure, lower cholesterol and prevent arteriosclerosis. It is effective, but no one has used fermentation to increase the content of hesperidin in citrus fruits and make it as effective as hesperidin extract.
有鑑於此，本案申請人遂依其多年從事相關領域之研發經驗，針對前述之缺失進行深入探討，並依前述需求積極尋求解決之道，歷經長時間的努力研究與多次測試，終於完成本發明。In view of this, the applicant in this case conducted in-depth discussions on the aforementioned deficiencies based on his years of research and development experience in related fields, and actively sought solutions to the aforementioned needs. After a long period of hard research and multiple tests, he finally completed the invention.
本發明之主要目的在於提供一種能降低三酸甘油脂之組合物。The main purpose of the present invention is to provide a composition capable of reducing triglycerides.
為達上述之目的，本發明具有抑制體脂肪形成之組合物，其係含有315~620μg/mL之橙皮苷以及160~230μg/mL之聖草次苷，且該組合物之製造方法係先將檸檬裂解成檸檬汁後，再將檸檬汁進行發酵而得。In order to achieve the above purpose, the present invention has a composition for inhibiting body fat formation, which contains 315~620μg/mL hesperidin and 160~230μg/mL eriocidin, and the preparation method of the composition is first After splitting the lemon into lemon juice, the lemon juice is fermented.
而該裂解的過程係將整顆檸檬連皮帶籽裂解成檸檬汁，又發酵時係將胚芽乳酸桿菌(Lactobacillus plantarum)以及季也蒙有孢漢遜酵母菌(Hanseniaspora guilliermondii)植入100%純檸檬汁中。The lysis process is to lyse the whole lemon and the seeds into lemon juice, and during fermentation, Lactobacillus plantarum and Hanseniapora guilliermondii are implanted into 100% pure lemons. In the juice.
為期許本發明之目的、功效、特徵及結構能夠有更為詳盡之瞭解，茲舉較佳實施例並配合圖式說明如後。In order to have a more detailed understanding of the purpose, efficacy, features, and structure of the present invention, preferred embodiments are described below in conjunction with the drawings.
本發明具有抑制體脂肪形成之組合物，其係含有315~620μg/mL之橙皮苷以及160~230μg/mL之聖草次苷，且該組合物之製造方法係先將檸檬裂解成檸檬汁後，再將檸檬汁進行發酵而得。The present invention has a composition for inhibiting the formation of body fat, which contains 315~620μg/mL hesperidin and 160~230μg/mL eriocidin, and the preparation method of the composition is to first crack lemon into lemon juice After that, the lemon juice is fermented.
而該檸檬的裂解係將整顆檸檬連同果皮和種子裂解成檸檬汁，接著再直接將益生菌植入100%純檸檬汁內進行發酵，且發酵的過程中需將溫度控制在26℃~32℃之間並持續攪拌12到24天，而在攪拌的過程中會不間斷的透過攪拌設備將空氣打入檸檬汁內，使檸檬汁、益生菌以及空氣三者能充分的混合。The lysis system of the lemon splits the whole lemon together with the peel and seeds into lemon juice, and then directly implants the probiotics into 100% pure lemon juice for fermentation, and the temperature must be controlled at 26℃~32 during the fermentation process Keep stirring between ℃ for 12 to 24 days, and during the stirring process, air will be pumped into the lemon juice through the stirring equipment, so that the lemon juice, probiotics and air can be fully mixed.
其中，該益生菌係由胚芽乳酸桿菌(Lactobacillus plantarum)以及季蒙也有孢漢遜酵母菌(Hanseniaspora guilliermondii) 以混合比例介於1~3:1~2之間所組成，又該檸檬汁和益生菌的混合比例介於1~2:0.04~0.4之間，該胚芽乳酸桿菌(Lactobacillus plantarum)寄存於德國菌種中心，寄存編號為DSM 32004，該季蒙也有孢漢遜酵母菌(Hanseniaspora guilliermondii)寄存於德國菌種中心，寄存編號為DSM 32005。Among them, the probiotic strain is composed of Lactobacillus plantarum and Hanseniaspora guilliermondii in a mixing ratio of 1~3:1~2, and the lemon juice and probiotics The mixing ratio of the bacteria is between 1~2:0.04~0.4. The Lactobacillus plantarum is deposited in the German Culture Center, and the deposit number is DSM 32004. This season also contains Hanseniaspora guilliermondii. Deposited in the German Culture Center, deposit number is DSM 32005.
本實施例以動物實驗作驗證，藉此來證實服用本發明之組合物確實具有調整三酸甘油脂之功效，有關動物實驗之相關方法以及檢測結果如下所示。In this example, animal experiments are used for verification to confirm that the composition of the present invention has the effect of adjusting triglycerides. The related methods and test results of animal experiments are as follows.
試驗動物：選用來自樂斯科生物科技股份有限公司的十週齡大鼠(Sprague-Dawley, SD)。Experimental animals: Ten-week-old rats (Sprague-Dawley, SD) from Lesco Biotechnology Co., Ltd. were selected.
飼育管理：實驗用之大鼠飼育環境控制在22±3℃，濕度在50±20%，並控制光照期為早上六點到晚上六點，黑暗期則為晚上六點到翌日早上六點，光照期與黑暗期各十二小時，且需每日監測環境溫度及濕度。Feeding management: The experimental rat breeding environment is controlled at 22±3℃, humidity is 50±20%, and the light period is controlled from 6 am to 6 pm, and the dark period is from 6 pm to 6 am the next day. The light period and the dark period are twelve hours each, and the ambient temperature and humidity need to be monitored daily.
飼料配方：正常組飼料採用Research Diets, Inc的AIN-93M，而對照組飼料則採用高熱量飼料，正常組和對照組的大鼠均採自由攝食方式餵食，而高熱量飼料之配方如下表(一)所示。
飲水：將自來水經逆滲透處理機處理後裝入經高溫高壓滅菌之塑料瓶後供大鼠自由飲用。Drinking water: Treat the tap water with a reverse osmosis treatment machine and put it into a high-temperature and high-pressure sterilized plastic bottle for rats to drink freely.
試驗組別設計：詳細的投予量如下表(一)所示，劑量推算係根據2005年美國食品藥物管理局所公告之實驗出其估算方法，以60公斤之成人為基準，使用高等實驗動物進行試驗時，劑量之換算原則為人體每日每公斤體重之建議攝取量的6.2倍為大鼠1倍劑量。
試驗劑量之配製：將本發明以冷凍乾燥之方式製備成粉末，以滅菌水直接配製濃縮液至投予體積，以50mL的本發明為例，其經濃縮處理後之體積為13.5mL，以人體與大鼠之劑量換算如下，低劑量對照組大鼠投予
投予期間、方法：將不同劑量之本發明與滅菌水配置後，直接進行管餵口服投予並持續八週。Administration period and method: After preparing different doses of the present invention and sterilized water, they are directly administered by tube orally for 8 weeks.
臨床症狀觀察：試驗期間每周定期量測大鼠體重並比較試驗開始與結束時之體重，且每日記錄各組大鼠之飼料攝取量並在試驗結束後計算食物利用率。Observation of clinical symptoms: During the experiment, the weight of the rats was measured regularly every week and the weight at the beginning and the end of the experiment was compared. The feed intake of each group of rats was recorded daily and the food utilization rate was calculated after the experiment.
體脂肪收集與量測：試驗結束後將大鼠犧牲並取出內臟脂肪(腹膜腔內之附睪、腎臟周圍及腸繫膜上面的脂肪)進行秤重並換算體脂肪率。Body fat collection and measurement: After the test, the rats were sacrificed and the visceral fat (the testicles in the peritoneal cavity, the fat around the kidney and the mesentery) was taken out and weighed and converted into body fat percentage.
臟器收集與量測：大鼠犧牲後取出其肝臟、腎臟、脾臟和心臟秤重。Organ collection and measurement: After the rats were sacrificed, their liver, kidney, spleen and heart were weighed.
血清學檢測：大鼠犧牲前禁食12小時後，以二氧化碳麻醉並收集腹腔靜脈血液，將血液離心後取上清液(血清)以全自動血清生化分析儀檢測(Automated biochemistry analyzer, Vitros 5.1 FS, Johnson & Johnson. Inc., USA)進行分析，分析的項目包含有：血脂、血糖、肝功能、腎功能、電解質平衡狀態。Serological test: After the rats were fasted for 12 hours before sacrifice, they were anesthetized with carbon dioxide and collected blood from the abdominal vena cava. , Johnson & Johnson. Inc., USA). The analyzed items include: blood lipids, blood sugar, liver function, kidney function, and electrolyte balance.
肝臟脂肪檢測：檢驗大鼠肝臟中之總膽固醇和三酸甘油脂含量，再依照Folch萃取法，取0.2g的肝臟加入20倍質量的氯仿/甲醇(2:1；v/v)溶液，以均質機研磨後取1.5mL上清液以氮氣吹乾並去除有機溶劑，再以triton X-100溶液回溶，溶液中之總膽固醇和三酸甘油脂以全自動血清生化分析儀檢測(Automated biochemistry analyzer, Vitros 5.1 FS, Johnson & Johnson. Inc., USA)檢測。Liver fat test: test the content of total cholesterol and triglycerides in rat liver, and then according to Folch extraction method, take 0.2g liver and add 20 times the mass of chloroform/methanol (2:1; v/v) solution to After grinding with a homogenizer, take 1.5 mL of the supernatant and blow dry with nitrogen to remove the organic solvent, and then re-dissolve it with triton X-100 solution. The total cholesterol and triglycerides in the solution are detected by an automatic serum biochemical analyzer (Automated biochemistry) analyzer, Vitros 5.1 FS, Johnson & Johnson. Inc., USA).
在本次動物試驗中，主要係探討本發明對於不易形成體脂肪之功效，並將試驗分為正常組(給予正常飼料)、負對照組(給予高熱量飼料)、低劑量對照組(給予高熱量飼料以及1倍相對人體劑量之本發明)、中劑量對照組(給予高熱量飼料以及2倍相對人體劑量之本發明)、高劑量對照組(給予高熱量飼料以及6倍相對人體劑量之本發明)，且每組均為12隻試驗動物，經連續給予8週後進行血液和臟器之相關檢測，而檢測結果以平均值正負標準差表示，而檢測數據以單因子變異數分析(One-way ANOVA)進行分析，並以鄧肯多重差距檢定(Duncan's multiple range test)進行事後檢定，其結果如下列所示：In this animal experiment, the main purpose of this experiment is to explore the effect of the present invention on not easily forming body fat, and divide the experiment into normal group (given normal feed), negative control group (given high-calorie feed), low-dose control group (given high Calorie feed and 1 times the relative human dose of the present invention), medium-dose control group (given high-calorie feed and 2 times the relative human dose of the present invention), high-dose control group (given high-calorie feed and 6 times the relative human dose of the present invention) Invention), and each group consists of 12 experimental animals. After 8 weeks of continuous administration, blood and organ related tests are performed. The test results are expressed by the mean plus or minus standard deviation, and the test data is analyzed by single factor variance analysis (One -way ANOVA) for analysis, and a post-test with Duncan's multiple range test. The results are as follows:
首先，就各組大鼠之每週平均體重之記錄如下表(三)所示。
表(三)中之上標英文字母a、b、c、d表示統計之結果，而不同字母表示組間統計有顯著差異（p>0.05）。In Table (3), the superscript letters a, b, c, and d indicate statistical results, while different letters indicate statistically significant differences between groups (p>0.05).
由上表(三)可看出除了正常組於試驗期間之體重皆顯著低於負對照組，表示高熱量飼料可成功誘導大鼠肥胖，中劑量對照組只在第八週的平均體重顯著低於負對照組，反觀高劑量對照組從第二週到第八週知平均體重均顯著低於負對照組，但低劑量對照組則和負對照組無明顯差異。It can be seen from the above table (3) that except for the normal group, the body weight during the test period was significantly lower than that of the negative control group, indicating that the high-calorie diet can successfully induce obesity in rats. The average weight of the middle-dose control group was significantly lower only in the eighth week. In the negative control group, the average weight of the high-dose control group from the second week to the eighth week was significantly lower than that of the negative control group, but there was no significant difference between the low-dose control group and the negative control group.
接著，就各組大鼠之每日飼料攝食量並統計該週之平均攝食量如下表(四)所示。
表(四)中之上標英文字母a、b、c、d表示統計之結果，而不同字母表示組間統計有顯著差異（p>0.05）。In Table (4), the superscript letters a, b, c, and d indicate statistical results, and different letters indicate statistically significant differences between groups (p>0.05).
由上表(四)可看出中劑量對照組之第7週以及高劑量對照組第1週、第4週至第7週的平均攝食量均顯著低於負對照組（p>0.05）。It can be seen from the above table (4) that the average food intake in the 7th week of the middle-dose control group and the first week, the 4th week to the 7th week of the high-dose control group were significantly lower than the negative control group (p>0.05).
接續，就各組大鼠之食物利用率和體脂肪率如下表(五)所示。
表(五)中之上標英文字母a、b、c表示統計之結果，而不同字母表示組間統計有顯著差異（p>0.05）。In Table (5), the superscript letters a, b, and c indicate statistical results, and different letters indicate statistically significant differences between groups (p>0.05).
由上表(五)可知在食物利用率方面正常組顯著低於給予高熱量飼料之各對照組，而在各對照組中之中劑量對照組和高劑量對照組之食物利用率顯著低於其他對照組。From the above table (5), it can be seen that the food utilization rate of the normal group is significantly lower than that of the control groups given high-calorie feed, and the food utilization rate of the middle-dose control group and the high-dose control group in each control group is significantly lower than the others Control group.
就體脂肪方面正常組的平均體脂肪重量明顯低於各對照組，而對各照組中又以中劑量對照組和高劑量對照組之體脂肪量顯著低於負對照組，另外在體脂肪率的部分同樣也是中劑量對照組和高劑量對照組之體脂肪率顯著低於負對照組。In terms of body fat, the average body fat weight of the normal group was significantly lower than that of the control groups. In the control groups, the body fat mass of the medium-dose control group and the high-dose control group was significantly lower than that of the negative control group. The percentage of body fat is also significantly lower in the middle-dose control group and the high-dose control group than the negative control group.
再來，就各組大鼠之血液中之尿酸(uric acid, UA)以及肌酐酸(creatinine, CRE)含量作為評估腎功能之指標，其結果如下表(六)所示。
由上表(六)可看出各組別血液中之尿酸以及肌酐酸均無差異，因此可知服用本發明不會影響腎臟正常之功能。From the above table (6), it can be seen that there is no difference in uric acid and creatinine acid in the blood of each group. Therefore, it can be seen that taking the present invention will not affect the normal function of the kidney.
接續，就各組大鼠血液中天門冬胺酸轉胺酶(AST)和丙胺酸轉胺酶(ALT)酵素活性來觀察試驗動物之肝臟功能，其結果如下表(七)所示。
由上表(七)可看出血液中之天門冬胺酸轉胺酶(AST)和丙胺酸轉胺酶(ALT)酵素活性在高熱量飲食誘導的組別中，不論投予劑量的多寡均和正常組無差異。From the above table (7), it can be seen that the enzyme activities of aspartate transaminase (AST) and alanine transaminase (ALT) in the blood are in the group induced by the high-calorie diet, regardless of the amount of dose administered. There is no difference from the normal group.
接續，就各組大鼠體內電解質平衡，其結果如下表(八)所示。
由上表(八)可看出在高熱量的飼料餵食下，不論是否服用本發明，大鼠血液中之鈉、鉀離子量均和正常組無差異，因此可知服用本發明不會影響大鼠體內之電解質平衡。It can be seen from the above table (8) that under the high-calorie feed, whether the present invention is taken or not, the amount of sodium and potassium ions in the blood of rats are the same as the normal group. Therefore, it can be seen that taking the present invention will not affect the rats. Electrolyte balance in the body.
接續，就各組大鼠血液中之血糖質檢測，其結果如下表(九)所示。
由上表(十)可看出血液中之血糖值在高熱量飲食誘導的組別中，不論投予劑量的多寡均和正常組無差異。It can be seen from the above table (10) that the blood glucose level in the high-calorie diet-induced group has no difference with the normal group regardless of the amount of dose administered.
接續，就各組大鼠血清中之總膽固醇(Total cholesterol)、三酸甘油脂(Total triglyceride)、高密度膽固醇(HDL-C)以及低密度膽固醇(LDL-C) 檢測，其結果如下表(十一)所示。
由上表(十一)可看出低、中、高劑量對照組血清中三酸甘油脂以及總膽固醇濃度均顯著低於負對照組，而在高密度膽固醇和低密度膽固醇濃度方面僅有高劑量對照組顯著低於負對照組。It can be seen from the above table (11) that the serum triglyceride and total cholesterol concentrations in the low, medium, and high-dose control groups are significantly lower than those in the negative control group, while the high-density cholesterol and low-density cholesterol concentrations are only high. The dose control group was significantly lower than the negative control group.
接續，就各組大鼠血液中之非酯化游離脂肪酸(FFA)含量檢測，其結果如下表(十二)所示。
由上表(十二)可看出在高熱量飲食誘導的對照組中非酯化游離脂肪酸的含量比正常組相比有顯著上升，而低、中、高劑量對照組之非酯化游離脂肪酸含量則和負對照組無明顯差異。It can be seen from the above table (12) that the content of non-esterified free fatty acids in the control group induced by the high-calorie diet was significantly higher than that in the normal group, while the non-esterified free fatty acids in the low, medium and high dose control groups The content is not significantly different from the negative control group.
接續，就各組大鼠肝臟中之總膽固醇(Total cholesterol)、三酸甘油脂(Total triglyceride)濃度含量檢測，其結果如下表(十三)所示。
由上表(十三)可看出在高熱量飲食誘導的對照組中，大鼠肝臟中的總膽固醇以及三酸甘油脂均高於正常組，而低、中、高劑量對照組肝臟中的總膽固醇以及三酸甘油脂均顯著低於負對照組。It can be seen from the above table (13) that in the control group induced by a high-calorie diet, the total cholesterol and triglycerides in the liver of rats are higher than those in the normal group, while the liver levels in the low, medium and high dose control groups The total cholesterol and triglycerides were significantly lower than the negative control group.
綜合上述表格可知大鼠在服用了本發明之後，其肝、腎機能以及電解質平衡不受影響，而低、中、高劑量對照組之大鼠肝臟脂肪含量顯著低於無服用本發明之負對照組，由此可知服用本發明是有助減輕體重、減少體脂肪量以及降低體脂肪率。Based on the above table, it can be seen that the liver and kidney function and electrolyte balance of rats are not affected after taking the present invention, while the liver fat content of rats in the low, medium and high dose control group is significantly lower than the negative control without taking the present invention From this, it can be seen that taking the present invention can help reduce body weight, reduce body fat mass and reduce body fat rate.
故，本發明在同類產品中具有極佳之進步性以及實用性，同時查遍國內外關於此類結構之技術資料文獻後，確實未發現有相同或近似之構造存在於本案申請之前，因此本案應已符合『創作性』、『合於產業利用性』以及『進步性』的專利要件，爰依法提出申請之。Therefore, the present invention has excellent advancement and practicability among similar products. At the same time, after searching through domestic and foreign technical documents about this type of structure, it is indeed not found that the same or similar structure exists before the application of this case. Therefore, this case The patent requirements of "creativeness", "applicability for industry" and "progressiveness" should have been met, and an application should be filed according to law.
唯，以上所述者，僅係本發明之較佳實施例而已，舉凡應用本發明說明書及申請專利範圍所為之其它等效結構變化者，理應包含在本發明之申請專利範圍內。However, the above-mentioned are only the preferred embodiments of the present invention. Any other equivalent structural changes made by applying the specification of the present invention and the scope of the patent application should be included in the scope of the patent application of the present invention.
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Citations (2)
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| CN1124133C (en) * | 1997-10-28 | 2003-10-15 | 韩国科学技术研究院 | Hesperidin and hesperetin as inhibitor of acyl coa-cholesterol-o-acyltransferase, inhibitor of macrophage-lipid complex accumulation on the arterial wall and preventive agent |
| CN1946830A (en) * | 2004-12-27 | 2007-04-11 | 百佳株式会社 | Antioxidant material, anti-deterioration agent and food or beverage |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1124133C (en) * | 1997-10-28 | 2003-10-15 | 韩国科学技术研究院 | Hesperidin and hesperetin as inhibitor of acyl coa-cholesterol-o-acyltransferase, inhibitor of macrophage-lipid complex accumulation on the arterial wall and preventive agent |
| CN1946830A (en) * | 2004-12-27 | 2007-04-11 | 百佳株式会社 | Antioxidant material, anti-deterioration agent and food or beverage |
Non-Patent Citations (1)
| Title |
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| Hashemi, Seyed Mohammad Bagher, et al. "Fermented sweet lemon juice (Citrus limetta) using Lactobacillus plantarum LS5: Chemical composition, antioxidant and antibacterial activities." Journal of Functional Foods 38 (2017): 409-414. * |
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