| Clinical chemical toxicants can be divided into five classes according to their different toxicities:ultrahigh toxic,high toxic,toxic,low-toxic,and micro-toxic.Among all vast chemical toxicants derived from various kinds and resources,more than5000 of them are with ultrahigh or high toxicity.In China,clinical intoxication accicidents resulted from the misuses or intentional poisoning of rodents,pesticides,sedative hypnotics,and plant toxic components occurred frequently in each year,posing a great threat to public safety and human health.Chemical toxicants can invade the body through various ways includng the respiratory tract,digestive tract and skin,causing acute toxicity,physiological dysfunction,multiple organ failure and even death to human.The primary fundemental of clinical chemical poisoning treatment is to identify which kind,how much dose,which way of intoxication,therefore,it is very important to develop the rapid and accurate clinical detection techniques towards chemical toxicants.Dried Blood Spot(DBS)is an advanced tool for blood collection and preservation.The preparation of DBS only consists of pricking finger or toe,dropping the blood on the filter paper card and drying at room temperature for more than 2 hours.The benefits of DBS include,simple sample collection,minimal invasion and convenient transportation and storage,which is currently highly valued in the field of clinical rapid test.DBS facilitates further combination with other analytical technique to implement rapid detection and continuous monitoring,such as liquid chromatography-tandem mass spectrometry,or even atmospheric mass spectrometry,etc.For the toxicants with low concentration,more structural similiarity with analogues,and much different properties,liquid chromatography-tandem mass spectrometry can realize a highly sensitive and selective detection,providing reliable qualitative and quantitative results for the clinical diagnosis and treatment for chemical toxicants.For the rapid detection of clinical chemical toxicants in DBS,how could we accomplish universal and efficient sample extraction,has become a key factor in the method development in DBS coupled with this liquid chromatography-tandem mass spectrometry.In addition,ambient ionization mass spectrometry is a promising tool for rapid and direct analysis of complex samples under the ambient atmosphere,enabling the collection of high throughput mass spectrometric information,so as to meet the clinical rapid detection and diagnosis need towards multiple samples.Low Temperature Plasma(LTP)ionization is one of ambient ionization mass spectrometric technique,which has unique features of simple construction,free of electrospraying solvent and low plasma temperature,which are beneficial for the non-destructive analysis of the body.Is the LTP mass spectrometry can be conviently coupled with DBS in clinical rapid tests on chemical toxicants?The difficulty lies in how could we enhance the ionization efficiency,and selectively detect trace analytes in complicated matrices.In this thesis,focusing on clinic DBS samples,we mainly investigated the kind of filter papers,the wetting step,and the kind of extraction solvents,and established a universal pretreatment method consisting of water soaking and methanol extraction.Using mycophenolic acid as an internal standard,we developed a liquid chromatographic-triple quadrupole mass spectrometric(LC-QqQ-MS/MS)method on qualitative and quantitative determination of twelve anticoagulant rodenticides in DBS.The results showed that,water-moisturizing contributes to higher extraction recoveries by helping the target compounds liberate from the cellulose chains.During the analysis of anticoagulant rodenticides in DBSs,different extraction efficiency was proived by DMPK-A,-B,-C and 903 cards.While when we used the filter card of 903 type as a supporting matrix,the extraction efficiency of the twelve rodenticides ranged from 66%to 115%with intraday relative standard derivations(RSDs)less than 15%.Good method validation results were found in all twelve anticoagulant rodenticides.The linearity for pindone was from 20 to 500 ng/mL with a correlation coefficient(R~2)of0.9987,and from 5 to 500 ng/mL with the R~2 varied from 0.9988 to 0.9996 for the other eleven analytes.The recoveries of all twelve analytes were ranged from 61%to 105%,while the matrix effects were ranged from 71%to 193%with intraday RSDs less than15%.The established method is accurate,sensitive,rapid,and convenient,it has been successfully applied in the detection of three clinic blood samples poisoned by rodenticides.It provides a new method for clinical diagnosis and forensic toxicant analysis on anticoagulant rodenticide intoxication accidents.Based on the above experiences,we developed a rapid pretreatment method for clinical toxicant colchicine in DBS prepared on 903 filter paper card,using a water-moisturizing step and a extraction step based on 0.1%formic acid-methanol.Compared with the pretreatment procedures towards plasma,the DBS pretreatment is simple,short time-consuming and with higher extraction efficiency.Remifentanil was used as an internal standard for the qualitative and quantitative detection of colchicine by LC-QqQ-MS/MS.The method validation showed that,colchicine has a linear range from 2 to 100 ng/mL(R~2=0.9991).The recovery of colchicine was ranged from 100 to115%,while the matrix effects were ranged from 108%to 153%with intraday RSDs less than 15%.All the stabilities met the method validation requirements,with the RSDs less than 15%.We have successfully applied this method to the detection of clinical poisoning samples,including plasma,urine,bone marrow,and dialysate effluent.The results showed that the results from DBS and plasma were in good agreement,and the concentration of colchicine in each biological sample was consistent with that reported in the literature.It provides a reliable new diagnosis method for clinical colchicine poisoning,and guiding the corresponding clinical treatment.To achive a direct,real-time detection towards clinical DBS,we developed a low temperature plasma-quadrupole-time of flight mass spectrometry(LTP-QTOF-MS)on rapid determination of psychoactive fentanyl drugs.We have performed preliminary investigation on the improvement of ionization efficiency and the modification methods on paper substrates.Using caffeine as a model compound,LTP parameters were inestigated and the most optimized conditions include,using copper wire as the external electrode,the electrode spacing was 3.5 cm,the voltage was 2~2.5 kV,and the He gas flow rate was 0.25 L/min,and the distance from the probe to the mass spectrometer inlet was 2 cm.For five kinds of fentanyl analogs,the tip of the LTP probe was reconstructed through the tip cannula to focus the plasma beam,which can increase the signal of the fentanyl compound by 2 to 3.5 times.In addition,a better mass spectral response can be obtained from the DBS paper substrate wetted with a 0.1%formic acid-methanol solution.Graphite-coating assisted laser irradiation helps to increase the conductivity and thermal conductivity of the paper substrate,the signals of analytes were then increased by 1.4~2.2 times.The dry gas flow rate was the main factor affecting the MS singal response while the analytes enter the analyzer of QTOF MS/MS after their desorption.The fentanyls had the highest response in the conditions of,the capillary signal voltage was 800 V,the dry gas flow rate was 8 L/min,and the sprayer pressure was 20 psig.Currently sensitivity of five fentanyl compounds in the LTP-QTOF MS was 350 ng/mL,while the sensitivity in LTP-QTOF MS/MS was slightly improved,it is still 1~2 orders of magnitude lower than that of LC-QTOF MS/MS.We will further optimize multi-factors such as high-powered laser-assisted ionization,distance from the tail plasma to the ion source,and end-focusing the LTP,etc,as well as the introduction of doping spray solvents to increase the ionization efficiency. |
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