食用昆蟲之熱穩定蛋白質特性分析及其抗體開發之應用

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Title:	食用昆蟲之熱穩定蛋白質特性分析及其抗體開發之應用 Characterization of edible insect thermally stable proteins and its application of the development of edible insect-specific antibody
Authors:	陳翊寧 CHEN, I-NING
Contributors:	食品安全碩士學位學程陳奕廷
Keywords:	食品過敏;食用昆蟲;熱穩定蛋白質;抗體開發 food allergy;edible insect;thermally stable protein
Date:	2023-07-20
Issue Date:	2023-12-07 09:50:16 (UTC+8)
Abstract:	食用昆蟲具有環境友善與營養價值，因此成為解決未來糧食危機的永續性糧食。2021至2023年麵包蟲（Tenebrio molitor）、家蟋蟀（Acheta domesticus）、蝗蟲（Locusta migratoria）、外米擬步行蟲幼蟲（Alphitobius diaperinus）通過歐盟新穎性食品審查。昆蟲成為新穎食品的同時，臨床研究指出食用昆蟲對於甲殼類與塵?過敏患者具有潛在的致敏風險。免疫測定法以具專一性的抗體檢測作為標的物的特定蛋白質，免疫測定法已普遍應用於食品過敏原檢測。目前仍無檢測食品中昆蟲成分的免疫測定法，因此本研究著重於昆蟲中熱穩定性蛋白質的特性分析，以此為基礎開發檢測昆蟲的抗體。本實驗以歐盟與各國公告之食用昆蟲作為研究樣本，利用不同食品加工方法分析食用昆蟲蛋白質的熱穩定性，並搭配質譜分析對熱穩定性蛋白質進行身份鑑定。經質譜分析，37 kDa蛋白質鑑定為原肌球蛋白（tropomyosin）。因此本研究以tropomyosin為標的物，開發檢測昆蟲的多株抗體（pAb-I）。所開發的pAb-I具有昆蟲專一性，可辨識所有測試的食用昆蟲樣本，對甲殼動物、軟體動物、魚類及禽畜動物樣本無交叉反應性。其抗原蛋白分布在37~150 kDa間，37 kDa蛋白質為各個昆蟲樣本中的主要抗原蛋白，此37 kDa蛋白質可耐受常見的加工方法（水煮、蒸煮、烘烤、油炸）與長時間加熱處理。以pAb-I檢測添加不同比例昆蟲粉（分別添加家蟋蟀、麵包蟲、蝗蟲的粉末）的食品時，pAb-I可檢測含10 %昆蟲粉的麵條與含10 ％蟋蟀粉的加工肉丸，pAb-I對添加家蟋蟀粉末的樣本檢測能力最佳。此外，pAb-I能檢測乾燥、罐頭、煙燻以及烘焙加工食品中的昆蟲成分。根據研究結果，pAb-I具有昆蟲專一性且能檢測食品中昆蟲成分於未來可用於建立檢測昆蟲的免疫檢測法。 Edible insect is an environmentally friendly and nutritious food. Therefore, it has become an important sustainable food for solving future global food crises. Mealworm, house cricket, migratory locust, and lesser worm have been authorized as novel food by European Union (EU) since 2023. The food industries of edible insects are undergoing rapid growth and development. Clinical studies have indicated that insects can cause allergic reactions in individuals with shellfish and dust mite allergies. It showed potential risks when patients with crustacean allergy consume insects. Currently, edible insects are not part of food allergens in EU food regulations. Due to the potential risks of insects, the EU stipulates that edible insect products must be labeled with insect species and scientific names, besides a warning statement for individuals with shellfish allergies to avoid consumption. At present, there is no immunoassay available for detecting insect components in food. The first step in developing an immunoassay is to determine a suitable target protein, followed by the detection of this target protein using specific antibodies. The edible insects in this study included EU-approved insect species and common edible insect species in other countries. This study characterized thermally stable proteins in edible insects by different food processing and analyzed by SDS-PAGE and LC/MS-MS for protein identification. The results showed that a 37 kDa protein was the major thermally stable protein among insect species tested. The 37 kDa protein of house cricket (Acheta domesticus) and mealworm (Tenebrio molitor) kept its integrity during different food processing, including boiling, steaming, roasting, and frying. The 37 kDa protein was identified as tropomyosin which was the major allergen in insects. These results revealed that tropomyosin is a suitable marker protin for the presence of edible inects in food products. The developed polyclonal antibody (pAb-I) detected heated samples of all 7 insect species without crossreaction with crustaceans, mollusks, fishes, poultries, livestocks and amphibian. The pAb-I can detect edible insect ingraident in food products with different food processing including smoking, dried, canned, and baking. The pAb-I showed better capability for detecting house cricket than mealworm and migratory locust. Based on the results, the edible insect-specific pAb-I has been successfully developed and can be used in the development of an immunoassay for detecting insect components in food products.
Description:	碩士指導教授:陳奕廷委員:陳玉華委員:侯又禎委員:陳奕廷
Data Type:	thesis
Appears in Collections:	[Master Program in Food Safety] Dissertations/Theses

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