尘螨过敏原蛋白及其硝化产物的鉴别

Identification of house dust mite allergens and their nitrated products

过敏原蛋白是引发尘螨过敏性疾病的主要因素。空气污染物O3和NOX能够引起过敏原蛋白的硝化,从而增强其致敏性。然而,目前还缺乏尘螨过敏原蛋白硝化产物的准确定量方法。本文建立了一种能够同时测定环境中两种主要尘螨过敏原蛋白(Der f 1和Der p 1)及其硝化产物(Nitrated Der f 1和Nitrated Der p 1)的检测方法。该方法将样品中的过敏原蛋白酶解消化成小分子的多肽,通过超高效液相色谱-串联质谱法(UPLC-MS/MS)测定小分子多肽,从而对过敏原蛋白及其硝化产物进行定性和定量分析。该方法采用同位素内标法定量,方法检测限(MDL)为0.33µg·g^(−1)(Der p 1)—0.57µg·g^(−1)(Nitrated Der p 1),相对回收率在43.0%—80.0%之间,方法选择性好,准确度和分析通量高。利用该方法对办公室灰尘中的尘螨过敏原蛋白及其硝化产物进行分析,结果表明所测灰尘样本中Der f 1含量为0.84—1.36µg·g^(−1),Nitrated Der f 1含量为3.71—35.6µg·g^(−1),Der p 1含量为10.3—30.0µg·g^(−1),Nitrated Der p 1含量为2.16—8.54µg·g^(−1)。Der f 1硝化率为80%—96%,Der p 1硝化率为14%—43%。风险评估结果显示所测灰尘样本的风险指数(RI)值为9—38,均处于高风险水平。

Allergen is the main contributor of allergic diseases caused by dust mite.Airborne pollutants O3 and NOx can lead to nitration of allergens,resulting in increase of the sensitization of these allergens.However,there is no accurate method to quantify the nitrated products of dust mite allergens up to date.A method was developed for the simultaneously determination of house dust mite allergens(Der f 1 and Der p 1)and their nitrated products(Nitrated Der f 1 and Nitrated Der p 1)in the environment.In this method,the allergen in the sample was enzymatically digested into small molecule polypeptides,and the small molecule polypeptides were determined by ultra-high performance liquid chromatography coupled with tandem mass spectrometry(UPLC-MS/MS),so as to conduct qualitative and quantitative analysis of allergens and their nitrated products.Isotope internal standards were employed for quantification.The method detection limits(MDL)ranged from 0.33µg·g^(−1)of Der p 1 to 0.57µg·g^(−1)of Nitrated Der p 1.The relative recoveries were 43.0%—80.0%for the analytes.The method showed good selectivity,high accuracy and analytical throughput.The house dust mite allergens and their nitrated products in office dusts were analyzed by this method.The results showed that the content of Der f 1 in the dust samples was 0.84—1.36µg·g^(−1),the content of Nitrated Der f 1 was 3.71—35.6µg·g^(−1),the content of Der p 1 was10.3—30.0µg·g^(−1),and the content of Nitrated Der p 1 was 2.16—8.54µg·g^(−1).The nitrification rate was 80%—96%for Der f 1 and 14%—43%for Der p 1,respectively.The risk assessment showed that the risk index(RI)value of the measured dust samples was 9—38,all of which were at a high risk level.