Gambaro Veniero, Roda Gabriella, Visconti Giacomo Luca, Arnoldi Sebastiano, Casagni Eleonora, Dell'Acqua Lucia, Farè Fiorenza, Paladino Eleonora, Rusconi Chiara, Arioli Stefania, Mora Diego
Dipartimento di Scienze Farmaceutiche, Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milano, Italy.
Dipartimento di Scienze Farmaceutiche, Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milano, Italy.
J Pharm Biomed Anal. 2016 Jun 5;125:427-32. doi: 10.1016/j.jpba.2016.03.043. Epub 2016 Mar 23.
The taxonomic identification of the biological material contained in the hallucinogenic mushrooms culture media, was carried out using a DNA-based approach, thus highlighting the usefulness of this approach in the forensic identification of illegal samples also when they are present as basidiospores mixed in culture media and spore-bearing fruiting body are not present. This approach is very useful as it allows the unequivocal identification of potentially illicit material before the cultivation and it enables to stop the material to the Customs and to destroy it due to its dangerousness without cultivating the "grow-kits" and without instructing a criminal case. In fact, even if psilocin and psilocybin and the whole mushrooms are illegal in many countries, there is no specific indication in the law about the so called "grow-kits", containing the spores. To confirm the data obtained by the taxonomic identification, a simple, reliable, efficient LC-UV method, using tryptamine as internal standard, suitable for the forensic quali-quantitative determination of psilocin and psilocybin in hallucinogenic mushroom was optimized, validated and applied to the mushrooms grown after the cultivation of the grow-kits seized by the judicial authority, with the authorization of the Ministry of Health. A cation exchange column was used in a gradient elution mode (Phase A: 50mMK2HPO4; 100mM NaCl pH=3 Phase B: methanol). The developed method was linear over the calibration range with a R(2)>0.9992 for both the analytes. The detection and quantification limits were respectively 0.01 and 0.1μg/mL for psilocybin and 0.05μg/mL and 0.1μg/mL for psilocin and the intra- and inter-day precision was satisfactory (coefficients of variation <2.0% for both the analytes). The content of psilocybin in the mushrooms grown from the seized "grow-kits" ranged from 1.02 to 7.60mg/g of dry vegetable material, while the content of psilocin from 0.415 to 8.36mg/g.
使用基于DNA的方法对致幻蘑菇培养基中所含生物材料进行分类鉴定，从而突出了该方法在法医鉴定非法样本中的实用性，即便非法样本是以担孢子形式混于培养基中且不存在带孢子子实体的情况下。这种方法非常有用，因为它能在培养前明确鉴定出潜在的非法材料，并能让海关拦截该材料，因其危险性而将其销毁，无需培养“种植套件”，也无需立案。事实上，尽管在许多国家，裸盖菇素、赛洛新和整个蘑菇都是非法的，但法律中没有关于含有孢子的所谓“种植套件”的具体规定。为了确认分类鉴定获得的数据，优化、验证了一种简单、可靠、高效的LC - UV方法，该方法使用色胺作为内标，适用于法医对致幻蘑菇中裸盖菇素和赛洛新进行定性定量测定，并将其应用于经卫生部授权、由司法当局查获的“种植套件”培养出的蘑菇。使用阳离子交换柱进行梯度洗脱模式（A相：50mM磷酸氢二钾；100mM氯化钠，pH = 3；B相：甲醇）。所开发的方法在校准范围内呈线性，两种分析物的R(2)>0.9992。裸盖菇素的检测限和定量限分别为0.01μg/mL和0.1μg/mL，赛洛新的检测限和定量限分别为0.05μg/mL和0.1μg/mL，日内和日间精密度令人满意（两种分析物的变异系数<2.0%）。从查获的“种植套件”中生长出的蘑菇中，裸盖菇素的含量范围为1.02至7.60mg/g干植物材料，而赛洛新的含量范围为0.415至8.36mg/g。
