Galli A, Mori F, Gori I, Lucherini M
Department of Preclinical and Clinical Pharmacology, University of Florence, Firenze, Italy.
Biochem Pharmacol. 1992 Jun 9;43(11):2427-33. doi: 10.1016/0006-2952(92)90323-b.
The protective action of 1,2,3,4-tetrahydro-9-aminoacridine (THA) against the long-lasting inactivation of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) brought about by diisopropylfluorophosphate (DFP) and physostigmine, as well as by neostigmine in the case of AChE only, was evaluated by a dilution technique using Electrophorus electricus AChE and horse serum BuChE as target enzymes. In parallel experiments, the ability of physostigmine itself to protect these enzymes from DFP was evaluated and compared with that of THA. THA pretreatment was seen to prevent in a dose-dependent manner the inhibition of both AChE and BuChE. However, it was appreciably more potent towards AChE than towards BuChE. THA mean EC50 values for protecting AChE against 10, 40 and 100 microM DFP were 0.04, 0.16 and 0.45 microM, respectively; against 1 microM physostigmine the value was 1.8 microM and against 1.2 microM neostigmine it was 3.0 microM. The THA mean EC50 value for protecting BuChE against 3 microM physostigmine was 0.55 microM and the values for protecting against 3, 10 and 40 microM DFP were 1.5, 3 and greater than 10 microM, respectively. The protective action of THA was time independent: recovery of the maximal enzymic activity was immediate upon dilution. Unlike THA, the protective action of physostigmine developed progressively after dilution and was maximal within 3-4 (AChE) or 6-8 hr (BuChE). Under our experimental conditions, 0.3 microM physostigmine protected approximately 70% of AChE from 40 microM DFP and 5 microM physostigmine protected 9 and 47% of BuChE from 40 and 3 microM DFP, respectively. The results of this work suggest that THA exerts its protective action by shielding the active site of AChE and BuChE from the attack of the inactivating agents on account of its higher enzymic affinity, whereas the protective action of physostigmine against DFP takes advantage also of the carbamylation of the enzyme. These results are in line with the hypothesis that protection of AChE is the primary mechanism responsible for the antidotal action of THA against organophosphorus poisoning.
采用稀释技术，以电鳗乙酰胆碱酯酶（AChE）和马血清丁酰胆碱酯酶（BuChE）作为靶酶，评估了1,2,3,4 - 四氢 - 9 - 氨基吖啶（THA）对由二异丙基氟磷酸酯（DFP）和毒扁豆碱引起的乙酰胆碱酯酶（AChE）和丁酰胆碱酯酶（BuChE）的持久失活的保护作用，对于AChE而言，还评估了其对新斯的明引起的失活的保护作用。在平行实验中，评估了毒扁豆碱自身保护这些酶免受DFP影响的能力，并与THA的保护能力进行了比较。观察到THA预处理能以剂量依赖的方式防止AChE和BuChE的抑制。然而，它对AChE的作用明显比对BuChE更强。THA保护AChE免受10、40和100微摩尔DFP抑制的平均半数有效浓度（EC50）值分别为0.04、0.16和0.45微摩尔；针对1微摩尔毒扁豆碱，该值为1.8微摩尔，针对1.2微摩尔新斯的明，该值为3.0微摩尔。THA保护BuChE免受3微摩尔毒扁豆碱抑制的平均EC50值为0.55微摩尔，保护其免受3、10和40微摩尔DFP抑制的值分别为1.5、3和大于10微摩尔。THA的保护作用与时间无关：稀释后酶的最大活性立即恢复。与THA不同，毒扁豆碱的保护作用在稀释后逐渐显现，并在3 - 4小时（AChE）或6 - 8小时（BuChE）内达到最大。在我们的实验条件下，0.3微摩尔毒扁豆碱可保护约70%的AChE免受40微摩尔DFP的影响，5微摩尔毒扁豆碱分别可保护9%和47%的BuChE免受40和3微摩尔DFP的影响。这项工作的结果表明，THA因其更高的酶亲和力，通过保护AChE和BuChE的活性位点免受失活剂的攻击而发挥其保护作用，而毒扁豆碱对DFP的保护作用还利用了酶的氨基甲酰化。这些结果与以下假设一致，即保护AChE是THA对有机磷中毒解毒作用的主要机制。
