| Objective: Paraquat(PQ)poisoning has become one of the most common pesticide poisoning.The death rate of PQ oral poisoning is up to 70%.There are still no specific antidotes for PQ poisoning at home and abroad.Currently,clinically comprehensive treatment is adopted,among which poison clear-up in vivo is a key therapeutic method,including gastric lavage,intentional diarrhea,diuresis and blood perfusion.Since the oral poison can quickly go into the intestinal tract,gastric lavage alone has limitations in clearing up the poison.As a kind of widely used adsorbent,activated charcoal is also used as clinical therapy for acute oral poisoning,but it hasn’t been generally accepted to routinely use the activated charcoal gavage to adsorb the poison.Especially for PQ clear-up,it still lacks strong experimental data support.To explore an effective method to clear up the poison in the gastrointestinal tract quickly and constantly,we decided to conduct an animal experiment research by using activated charcoal gastric administration for adsorption therapy,which aims to explore the activated charcoal’s adsorption effect on PQ in the gastrointestinal tract in order to provide basic reference data for clinical therapy of PQ oral poisoning patients.Methods:(1)To Draw standard curve of PQ concentration in blood plasma.Surface-enhanced Raman spectroscopy(SERS)method was adopted.Instrument: The i-Raman portable spectrometer.Spectrum conditions: 785 nm in wavelength,image resolution 4.5 cm-1.60 m W in laser power.Au NPs with a grain size of 50 nm.The sampling time of single samples is 10 s and the pre-processing time of samples is within1 minutes.We formulated PQ working solution of 1 ng/ml,5 ng/ml,10 ng/ml,50 ng/ml,100 ng/ml,200 ng/ml with the PQ standard solution.Then we measured the Roman peak value and valley value to of each PQ working solution by SERS to calculate the difference value,that is Raman intensity.Then according to the concentration and the corresponding Raman intensity,we drew the PQ standard curve,with PQ concentration as the horizontal coordinate and PQ Raman intensity as the vertical coordinate.The linearity was good when PQ concentration was between 1ug/L and 200ug/L.Quantitative lower limit is 1ug/L,and quantitative higher limit is 200ug/L.Then we analyzed it with weighted least square method(WLS).The linear regression equation is the standard curve equation.(2)To establish hepatic portal vein blood sampling path.To directly observe PQ’s adsorption effects of activated charcoal in the gastrointestinal tract,the method of compare and analysis of the poison concentration in the hepatic portal veins and peripheral veins is adopted.Before operation,the beagles can’t eat for 8 hours and can’t drink for 2 hours.The experimental animals were given propofol injection(1.0 ml/kg)for anesthesia,and then by orotracheal intubation,we connected the anespirator set with a mixture of 1.5%-3.0%(v/v)isoflurane and oxygen.Then the operation was made by placing one end of PICC into the hepatic portal vein of a beagle and leaving the other end out of its body to establish hepatic portal vein blood sampling path.The anesthesia effects were good in the operation group.Anesthesia induction and the recovery were all quick,and the vital signs were stable during the operation.When finishing the operation,the beagles can stand about 10 minutes after dose administration.The average operating time was about 1 hour and the average amount of bleeding was about 15 ml.After operation,normal drinking was recovered and 4 hours later normal feeding was recovered.5-7 d later,the operation incision healed.PICC went beneath the skin to the back neck and then it was fastened,which can prevent beagles from biting it and can also keep it clean.After operation,normal saline was used to wash the tube and 125u/ml heparin sodium solution was used to keep it smooth.After 3 d for operation,the vital signs of beagles recovered as before,then we can conduct the relative research.(3)To better observe the effects of poison clear-up,the model of PQ exposure to beagles’ digestive tracts is adopted in the animal experiment.According to the degree of toxicity and the survival rate of clinical PQ poisoning patients,moderate or severe patients who take 20-40mg/kg dose of PQ is the focus of clinical therapy.We calculated the administrated dose according to the body surface area ratio of human to beagles.Through preliminary experiments which proved that the behaviors and PQ concentration in plasma of beagles after exposure to PQ were similar to clinical patients of moderate poisoning,we determined that the administrated dose of PQ for beagles is30 mg/kg.Then the model of PQ exposure to beagles through the gastric tube is established.AACT and EAPCCT recommend that the dosage of activated charcoal foran adult be 25-100 g(about 0.5-2.0 g/kg)and the time be within 1 hour after oral poisoning.Based on this,considering the results of pre-experiment and the time when patients are sent to hospitals,we determined that the dosage for beagles is 1.0 g/kg mixed with 100 ml the normal saline after exposure to PQ for 30 minutes.(4)Grouping and disposal of dogs.Twenty healthy male beagles were randomly divided into experimental group and control group,with 6 beagles in each group.20%PQ solution(a dose of 30 mg/kg)was prescribed through stomach for beagles in both groups.After exposure to PQ for 30 minutes,the beagles in experimental group were given activated charcoal suspension(1.0 g/kg of type I activated charcoal powder mixed with 100 ml of normal saline)by gavage,while the control group was only given equal volume of normal saline.After exposure to PQ for 10 minutes,30 minutes,and 1,2,4,8,12,24,and 48 hours,blood was collected from hepatic portal veins and peripheral veins to detect the PQ concentration change in the plasma.The blood from hepatic portal vein and peripheral vein was separately injected into the anticoagulation tubes with heparin sodium for centrifugation(8000 r/min for 10 minutes).The centrifuged plasma was used to detect PQ concentration.The toxicokinetics software DAS2.1.1 was applied to analyze PQ concentration and compare the change in toxicokinetics parameters between the both groups.The change in vital signs including heart rate(HR),respiratory rate(RR)and pulse oxygen saturation(Sp O2)was dynamically monitored 10 minutes before exposure,4 hours and each day from the 1st to the 7th day after exposure.Results:(1)Effects of activated charcoal gastric administration on PQ concentration in the plasma of hepatic portal veins and peripheral veins.Through statistics analysis and toxicokinetics analysis of PQ concentration data,the results were as followings.After exposure to PQ,the poison concentration in the plasma of hepatic portal veins and peripheral veins in the control group rose quickly and reached peak 4 hours later.Then it fell quickly and 8 hours later it fell slowly.But in the experimental group,the increase rate to the peak was significantly slow.Besides,PQ peak fell more obviously than that in the control group and it was about 50% of the control group [ug/L: 123.50±11.67 vs.255.18±12.29 in blood form hepatic portal veins,122.35±11.72 vs.250.86±11.15 in blood form peripheral veins,all P<0.01].After 8 hours it fell much more quickly than that of the control group.After exposure to PQ for 48 hours,PQ concentration in the plasma was still lower than that of the control group [ug/L: blood form hepatic portal veins 0.53±0.18 vs.15.98±5.58,blood form peripheral veins 0.31±0.01 vs.15.03±4.82,all P<0.05].With the toxicokinetics analysis,the area under the curve(AUC)and Cmax of PQ in the plasma of hepatic portal veins and peripheral veins in the experimental group were significantly lower than those of the control group [AUC(mg·L-1·h-1): blood form hepatic portal veins 1.6±0.2 vs.3.3±0.4,blood form peripheral veins 1.5±0.2 vs.3.2±0.3;Cmax(ug/L):blood form hepatic portal veins 125.07±9.49 vs.255.18±12.29,blood form peripheral veins 123.38±9.52 vs.250.86±11.15,all P<0.05].PQ plasma half-life(t1/2)in the experimental group was significantly shorter than that of the control group [hours: blood form hepatic portal veins 3.8±1.2 vs.15.4±3.7,blood form peripheral veins 3.5±1.0 vs.15.5±2.7,all P<0.05].Time to the peak(Tmax)of PQ in the experimental group was slower than that of the control group [hours:blood form hepatic portal veins 5.3±1.9 vs.4.0±0.0,blood form peripheral veins 4.7±1.5 vs.4.0±0.0,all P<0.05].PQ mean retention time(MRT)was significantly shorter than that of the control group [hours:blood form hepatic portal veins 8.0±1.5 vs.13.4±1.2,blood form peripheral veins 7.6±1.3 vs.13.3±1.2,all P<0.05].(2)Effects of activated charcoal gastric administration on vital signs of poisoning beagles.The change in vital signs including heart rate(HR),respiratory rate(RR)and pulse oxygen saturation(Sp O2)was dynamically monitored 10 minutes before exposure,4 hours and each day from the 1st to the 7th day after exposure.With the repeated measures ANOVA analysis of data about vital signs in two groups,the results were as followings.After exposure to PQ,HR and RR in both the experimental group and the control group tended to speed up and reached to the peak in about 4d and then it began to slow down gradually;Sp O2 tended to slow down gradually and reached to the valley in about 4d and then it began to recover,but the change range of vital signs in the experimental group was smaller than that of the control group,and the parameters were significantly better than those of control group [5-day HR(bpm): 130.50±4.57 vs.143.83±2.67,5-day RR(times/min): 30.50±0.50 vs.33.83±0.69,5-day Sp O2:0.912±0.007 vs.0.880±0.008,all P<0.01].Conclusions: 1.Early use of activated charcoal in the digestive tract can slow down the rising rate of PQ concentration in the plasma,cut down the peak concentration and has less influence on vital signs,which indicates that activated charcoal has a good adsorption of PQ in the gastrointestinal tract.Activated charcoal gavage in the early stage has vital application value for rescuing patients of PQ poisoning.2.It needs further research about when we don’t choose activated charcoal for therapy after oral administration of PQ. |
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