Richard J. Kitz

Activity-structure relationships in the reactivation of diethylphosphoryl acetylcholinesterase by phenyl-1-methyl pyridinium ketoximes

Abstract The effect of structure on the activity of phenyl-1-methyl pyridinium ketoximes and bis quaternary derivatives in reactivating diethylphosphoryl acetylcholinesterase was investigated. The inhibited enzyme was reactivated in the presence of acetylcholine, with a pH stat to maintain the pH and record the amount of ester hydrolyzed vs. time during the reactivation. A graphical technique was devised to obtain the pseudo first-order rate constant from this record. The configurations of the 4-derivatives were assigned on the basis of the Beckmann rearrangement. The anti -2-derivative is 20 times more active than the syn , whereas the anti -4-derivative is 6 times less active than the syn . The remarkable increase in activity that occurs in the bis-4-quatemary compounds occurs with both configurations. Compared to the aldoximes, introducing a phenyl group has a modest negative effect on the activity in the 4-derivatives, but the effect is more pronounced in the 2-derivatives.

THE CHEMISTRY OF ANTICHOLINESTERASE ACTIVITY

The molecular level chemistry of the acetylcholine‐acetylcholinesterase system has been examined in some detail. The mechanisms of normal substrate hydrolysis and anticholinesterase activity were stressed.

The reaction of acetylcholinesterase with O-dimethylcarbamyl esters of quaternary quinolinium compounds

Abstract The dimethylcarbamate esters of some mono- and diquaternary quinolinols were prepared and their anticholinesterase properties investigated by using acetyl-cholinesterase from electric eel. The I 50 values in the steady state of equal rates of carbamylation and decarbamylation were measured and the values of the second-order rate constants for carbamylation were evaluated. Compounds containing the dimethyl-carbamyl function substituted in the 5 and 7 positions were of the same order of activity as neostigmine to which they are structurally related. When substituted in the 3 position, the dimethylcarbamyl derivative was about as active as pyridostigmine which is structurally similar. It was concluded that the structure of the leaving group is important in determining the activity of these compounds and the p K a value of the leaving group appears to be relatively unimportant.

A study in vitro of new short-acting, non-depolarizing neuromuscular blocking agents.

Abstract Presently available short-acting, depolarizing myoneural blocking agents have significant side effects such as increased intraocular tension, cardiac arrhythmias desensitization of the postjunctional membrane, skeletal muscle injury and muscle pains. Non-depolarizing blocking drugs do not produce profound blocks of short duration. A program was evolved to synthesize compounds producing a non-depolarizing block at low concentrations with a brief action span requiring no antagonists. Several series of mono- and diquaternary esters of alkyl aminoethanols were prepared and studied in vitro . The rates of spontaneous hydrolysis and the hydrolysis catalyzed by acetylcholinesterase and plasmacholinesterase were measured. In addition, the anticholinesterase effects of these agents were evaluated. The myoneural blocking properties were studied with intracellular recordings in frog sciatic nerve-sartorius muscle preparations. Activity-structure relationships are reported. Several compounds warrant further study including investigations in vivo .

The reaction of acetylcholinesterase (AChe) with some quaternary hydroxy aminophenols

Abstract A series of mono- and diquaternary hydroxyl derivatives of quinolinol, isoquinolinol, and stilbazole were synthesized. The dimethylcarbamyl and diethylphosphoryl analogs of these hydroxyl compounds are potent anticholinesterase agents. The anticholinesterase properties of the hydroxyl analogs were evaluated by measuring the values of their competitive and noncompetitive binding constants for the reaction with electric eel acetylcholinesterase. The p K a values were also measured. When compared to the nonhydroxylated reference compounds, the addition of the OH group was found to increase binding of the undissociated species to free AChe (competitive component). These derivatives are generally more poorly bound to the acetyl enzyme (noncompetitive component), and the diquaternary compounds are less potent than their monoquaternary analogs. The values of the binding constants for the reaction of the undissociated forms of the inhibitor with the enzyme were found to correlate qualitatively with the second-order rate constants for carbamylation of the enzyme by the dimethylcarbamyl analogs, but not with the constants for phosphorylation by the diethylphosphoryl compounds. The latter compounds have been shown to correlate well with the p K a value of the quaternary hydroxyl derivatives.

HUMAN TISSUE CHOLINESTERASES: RATES OF RECOVERY AFTER INHIBITION BY NEOSTIGMINE; MICHAELIS-MENTEN CONSTANTS.

Abstract The kinetics of inhibition of acetylcholinesterase by neostigmine is briefly reviewed. The acid-transferring mechanism of inhibition which is similar to that of diisopropylfluorophosphate and tetraethylpyrophosphate has recently been shown to be also appropriate for neostigmine. Both solutions and suspensions of cholinesterases were obtained from human brain, skeletal muscle, and red cells. Using in vitro techniques, it was demonstrated that the half-time for the recovery of catalytic ability after inhibition by neostigmine was similar in both the suspensions and the solutions for the three tissues and averaged 34 min. The Michaelis-Menten constants for the tissue acetylcholinesterases were also shown to have similar values, about 2 × 10 −4 M. These constants had not previously been reported for human brain or skeletal muscle enzymes. Both the recovery times and the Michaelis-Menten constants are of the same order as reported by Wilson for eel acetylcholinesterase. These studies indicate the kinetic similarity of human and eel acetylcholinesterase with respect to carbamate inhibition. The relationship of these investigations to clinical observations of patients treated with neostigmine is discussed.

Teaching effectiveness: evaluation of computer-assisted instruction for cardiopulmonary resuscitation.

Computer instruction for cardiopulmonary resuscitation (CPR), using a case-simulation program, was evaluated for its teaching effectiveness. First-month anesthesia residents, divided into two groups, A and B, underwent a cognitive evaluation on their 1st day of arrival and again on their 5th day. Both groups were provided with clear educational objectives and reading materials, a tutor was as signed to each student to answer specific questions. Students in group A, also given computer-assisted instruction (CAI), showed a significantly higher learning than those in group B, who served as controls. That CAI was a highly motivational form of learning was established by later voluntary participation in CAI of all group B students.

The pharmacology of new short-acting nondepolarizing ester neuromuscular blocking agents: clinical implications.

OR over 15 years, a need has been recogF nized in anesthesia for a short-acting nondepolarizing neuromuscular blocking agent.l-4 Such an agent might well eliminate many or all of the clinical problems currently experienced with succinylcholine.3~4 To this end, we have designed, synthesized, and pharmacologically evaluated more than 100 diester diquaternary ammonium compounds,- of which some in the current series may merit clinical trial, in view of the results obtained in experimental animals.

The reaction of acetylcholinesterase with methanesulfonyl esters of quaternary quinolinium compounds

The methanesulfonyl esters of quaternary and diquaternary quinolinium compounds were synthesized and studied as inhibitors of purified eel acetylcholinesterase. Some derivatives produced only reversible inhibition of the enzyme; the values of their binding constants were measured. For those compounds producing irreversible (acid-transferring) inhibition, the values of the binding constants, first-order and second-order rate constants were measured. It was shown that the values of the second-order rate constants cannot be correlated with the pKa value of the leaving group (unlike the previously studied diethylphosphoryl quinolinium compounds). The values of the binding constants for the reaction of the quaternary hydroxyl analogs with the enzyme were compared with the values measured for the respective methanesulfonyl compounds. The data indicate that, depending on position, the methanesulfonyl group has little effect on or is detrimental to non-covalent binding between enzyme and inhibittor. The potency of these compounds is generally less than their respective dimethylcarbamyl and diethylphosphoryl analogues. However, several decamethylene bridged diquaternary compounds are the most potent of the methanesulfonate anticholinesterase agents.

Decamethonium and Serum Potassium in Man

Decamethonium and succinylcholine were used to study the effects of depolarizing muscle relaxants on serum potassium in 60 patients, free of neuromuscular disease, during major orthopedic surgery. Significant increases in serum K+were found after administration of decamethonium or succinylcholine in the usual clinical doses. The abnormal elevations of serum K+found in patients with burns, massive trauma, or muscle denervation are thus acentuations of the process that occurs in normal man following use of these depolarizing drugs. The administration of any depolarizing agent to these abnormal patient groups would, therefore, appear contraindicated.