云南楚雄3种环柄菇中鹅膏肽类毒素的检测与分析

Detection and analysis of amatoxins in 3 species of Lepiota from Chuxiong,Yunnan Province

目的对云南楚雄地区3种环柄菇中的6种鹅膏肽类毒素含量进行分析。方法采集的蘑菇45℃烤干,采用形态学和分子生物学方法进行物种鉴定。将蘑菇分为3个不同生长时期(生长期、成熟期、衰老期)和不同组织部位(菌盖、菌柄、整体),用超纯水超声提取毒素,采用超高效液相色谱-三重四极杆串联质谱法、电喷雾(ESI-)模式下多反应监测(MRM)方式检测,外标法定量。结果细环柄菇(Lepiota clypeolaria)和冠状环柄菇(Lepiota cristata)未检出鹅膏肽类毒素,肉褐鳞环柄菇(Lepiota brunneoincarnata)中检测出α-鹅膏毒肽(α-AMA)、β-鹅膏毒肽(β-AMA),其中α-AMA含量高于β-AMA。α-AMA总含量成熟期(1.67 mg/g)>生长期(1.56 mg/g)>衰老期(0.40 mg/g),β-AMA总含量成熟期(0.71 mg/g)>生长期(0.60 mg/g)>衰老期(0.14 mg/g)。两种毒素在生长期和成熟期的分布为:菌盖>菌柄,而衰老期为菌柄>菌盖。结论楚雄地区的肉褐鳞环柄菇L.brunneoincarnata主要含有鹅膏毒肽,包括α-AMA、β-AMA、Amanin、Amaninamide,不含鬼笔毒肽及毒伞肽,并且在不同生长期、不同组织部位中的α-AMA、β-AMA含量具有差异,能够为该类蘑菇中毒的临床诊断及治疗提供参考。

Objective To analyze the content of 6 kinds of amatoxins in 3 species of Lepiota collected from Chuxiong of Yunnan Province.Methods The collected mushrooms were baked and dried at 45℃,and species identification was carried out by morphological and molecular biology methods.The mushrooms were divided into three different growth periods(growth period,mature stage,aged stage)and different tissue parts(pileus,stipe,whole),and the toxins were extracted by ultrapure water ultrasound.The determination was performed by ultra-performance liquid chromatographytriple quadrupole tandem mass spectrometry with electrospray(ESI-)in multiple-reaction monitoring(MRM)mode,and the content was quantitatively analyzed by external standard method.Results No amatoxins were detected in Lepiota clypeolaria and Lepiota cristata,whileα-amanitin andβ-amanitin were detected in Lepiota brunneoincarnata,and the content ofα-amanitin was higher thanβ-amanitin.The total content ofα-amanitin was highest in the mature stage(1.67 mg/g),followed by the grow stage(1.56 mg/g)and least in the aged stage(0.40 mg/g).And the total content ofβ-amanitin was also highest in the mature stage(0.71 mg/g),followed by the growth stage(0.60 mg/g)and lowest in the aged stage(0.14 mg/g).The cap part was richer inα-amanitin andβ-amanitin than the stipe part in the growth and mature stages,while the content ofα-amanitin andβ-amanitin in the aged stage was higher in the stipe than in the pileus.Conclusion In Chuxiong Prefecture,Yunnan Province,the flesh-brown ring-stalk mushroom L.brunneoincarnata mainly contains Amatoxins,includingα-AMA,β-AMA,Amanin,and Amaninamide,excluding phalloidin and virotoxin.The content ofα-AMA andβ-AMA varies in different growth periods and tissues,providing reference for the clinical diagnosis and treatment of poisoning by this type of mushroom.