Cholinesterase. Its significance in anaesthetic practice.

Plasma cholinesterase is an enzyme which has importance to the anaesthetist primarily for its role in the metabolism of suxamethonium, although other anaesthetic related drugs that this enzyme metabolises are also increasingly important. In this article we review current thoughts on the function, profile and chemistry of plasma cholinesterase. Causes of variations in the activity of the enzyme are described and the basis of genetic variations is explained.

Recognition of the Ek1Ek1 homozygote for plasma cholinesterase.

A family is reported in which the propositus was found to be sensitive to suxamethonium. Both parents were heterozygote each having genotype Ea1Ek1. The sibling of the propositus had the most common phenotype as defined by dibucaine, fluoride and RO2 numbers. Genetic analysis, however, indicated that this sibling must be an Ek1Ek1 homozygote.

Inheritance of low plasma cholinesterase activity in two families with a history of suxamethonium sensitivity.

An hypothesis advanced in 1970 concerning family inheritance of low enzymic activities of plasma cholinesterase has been substantiated. The recognition of three E1kE1s genotypes is reported in this family after investigation of a new generation. A second family with similar genetic characteristics is described.

Segregation of the E1j gene for plasma cholinesterase in family studies.

Four families are reported in which the E1j gene is segregating. Two E1jE1k and one E1jE1f genotypes have been recognised by genetic analysis. The biochemical characteristics of several E1j genotypes are presented.

Family studies of the E1kE1s genotype for plasma cholinesterase.

Six individuals in three families with a history of suxamethonium sensitivity have been found to have genotype E1kE1s. The biochemical data for the recognition of this genotype have been analysed and the mean values compared with similar parameters for the usual phenotype. Individuals with genotype E1kE1s will be sensitive to suxamethonium.

Recognition of two new phenotypes segregating the E1k allele for plasma cholinesterase.

The first identification of the cholinesterase variants E1kE1k and E1kE1s is reported from a family study. The evidence is based on the biochemical parameters of enzymic activity, and dibucaine, fluoride and RO2 numbers. Two individuals appear to be homozygotes E1kE1k and two are heterozygotes E1kE1s with family evidence supportive of these genotypes. The heterozygotes E1kE1s will be sensitive to suxamethonium.

Analysis of suxamethonium sensitivity following termination of pregnancy.

The cholinesterase genotypes in the majority (25/35) of patients with suxamethonium sensitivity following termination of pregnancy are heterozygotes with an E1a gene. Twelve of these patients have the genotype E1uE1a. The reported duration of apnoea is minimal in the heterozygotes lacking the E1a gene (about 5-10 min) and maximal in the homozygotes E1aE1a (about 35 min). With few exceptions, the heterozygotes having an E1a gene are apnoeic for 10-15 min. The apparent low frequency of suxamethonium in these patients is discussed.

Phenotyping of individuals sensitive to suxamethonium. The Cholinesterase Research Unit at the Royal Postgraduate Medical School.

Four hundred and thirty blood samples from suxamethonium-sensitive individuals have been phenotyped by the Cholinesterase Research Unit following its transfer from Exeter to the Hammersmith Hospital. The distribution of genotypes has been shown to be similar to that found in Exeter. Screening for the Elk and Elj genes has not yielded any major differences in the gene frequencies of sensitive individuals, even during pregnancy. The uneven sex distribution of the patients, as well as other unusual points that have arisen, are discussed. A new gene for the biosynthesis of cholinesterase has probably been identified.

E1h, a new allele at cholinesterase locus 1.

Unusual inhibition characteristics in two unrelated suxamethonium-sensitive individuals were indicative of a new allele, E1h, segregating with the E1a gene. Family studies substantiate this hypothesis and three new genotypes are recognised.

Plasma cholinesterase variants. Family studies of the E1k gene.

The use of RO2-0683 as differential inhibitor has shown that 13% of 181 heterozygotes, believed to be E1uE1a, are in fact E1aE1k. The results indicate that slightly more than 1% of the British population could be homozygote E1kE1k. Analysis of 47 families segregating the E1aE1k phenotype in more than 2 blood relatives is restricted by the inability to differentiate the genotypes E1uE1k and E1kE1k from E1uE1u and E1uE1f.

Comparison of a commercially available assay system with two reference methods for the determination of plasma cholinesterase variants.

For assaying plasma cholinesterase (EC 3.1.1.8) activity and phenotyping by means of dibucaine inhibition, we have compared a commercially available kit, in which butyrylthiocholine is used as substrate, with two reference methods, one using benzoylcholine and the other propionylthiocholine. With 50 different samples of three of the most common genetic variants, we could clearly differentiate the variants with benzoylcholine and dibucaine, whereas there was some overlap of the E1uE1u and E1uE1a phenotypes with the other two substrates at 30 degrees C. The phenotypes were better differentiated at 25 degrees C, and in our hands the use of butyrylthiocholine was preferable to propionylthiocholine for phenotyping with dibucaine. The affinity of the usual and atypical homozygotes for fluoride with butyrylthiocholine gave an inverted response to the affinity of these variants for the anion with benzoylcholine. We suggest that this may be explained by the role of the chromogen or its products in the assay procedure with the thiocholine substrate.

Suxamethonium apnoea associated with pregnancy and liver dysfunction in a treated cretin.

A patient, with a history of cretinism, who developed suxamethonium apnoea as a consequence of a reduction in plasma cholinesterase activity secondary to the concurrence of pregnancy and liver dysfunction associated with pre-eclampsia is reported. The patient had a normal phenotype for plasma cholinesterase.

Studies on the inhibition by propranolol of some human erythrocyte membrane enzymes and plasma cholinesterase.

Human erythrocyte acetylcholinesterase and the plasma cholinesterase variants are not only inhibited by propranolol but have been found to show stereospecificity for its isomers. The erythrocyte enzyme has a greater affinity for the L-isomer than either the racemate or the D-isomer. In contrast the plasma cholinesterases have greater specificity for the D-isomer than the other isomer or racemate. The usual enzyme shows greater stereospecificity than the atypical enzyme and these findings present additional evidence that these enzyme variants differ in structure at the catalytic active site. Neither Na+ + K+ -ATPase nor Mg2+-ATPase show stereo-specificity for the isomers of propranolol although both enzymes are inhibited by the drug. The action of the drug on the four enzymes in blood samples obtained from patients having Huntington's disease was found to be identical to those observed on the enzymes in blood samples from healthy controls.

Inhibition of the plasma cholinesterase variants by propranolol.

The inhibitory effect of (+/-) propranolol 1.69 x 10(-4)-1.69 x 10(-7) mol litre-1 on normal and atypical plasma cholinesterase variants was investigated. The atypical enzyme is less sensitive to inhibition by (+/-) propranolol or either of its enantiomorphs than the usual enzyme. Propranolol 8.45 x 10(-6) mol litre-1 was used as differential inhibitor of 643 plasma samples from individuals of known genotype. Although the measurement of propranolol inhibition alone is not always unambiguous for assigning a definite genotype to a given individual, the correlation of propranolol inhibition with fluoride inhibition gives clear differentiation of the E1uE1u and E1UE1f phenotypes as well as other phenotypes.

Malignant hyperthermia and the fluoride-resistant gene.

One hundred and six individuals from 33 families with a history of malignant hyperthermia have been investigated for plasma cholinesterase variants. An increased frequency of the fluoride-resistant gene has been found. Although an adequate explanation for our results is elusive, some hypotheses are discussed.

Differential inhibition of plasma cholinesterase variants using the dibutyrate analogue of pancuronium bromide.

A steroid, the dibutyrate analogue of pancuronium bromide (9.8 X 10(-8)M), has been used as differential inhibitor in the study of the plasma cholinesterase variants. Pancuronium dibutyrate numbers have been measured for 190 individuals, and the mean values for six of the known genotypes, E1uE1u, E1uE1f, E1uE1a, E1fE1a, E1aE1a, and E1fE1f, have been calculated. Evidence is presented that a combination of the pancuronium dibutyrate number and the fluoride number give better resolution of the six genotypes than the combination of the pancuronium dibutyrate and the dibucaine number. This new differential inhibitor has real potential for revealing the probable existence of new genotypes.

A comparison of some methods of phenotyping the plasma cholinesterase variants using benzoylcholine as substrate.

Five differential inhibitors of plasma cholinesterase have been compared using benzoylcholine as substrate. None of the inhibitors (dibucaine, NaF, NaBr, NaCl, or pancuronium dibutyryloxy bromide) could be used singly to resolve all the variants. Better resolution was obtained when two inhibitors were used in conjunction. Clear differentiation of all six genotypes was obtained only with the combined use of pancuronium dibutyryloxy bromide and sodium fluoride. The limitations of some of the parameters are discussed.

Inhibition of the plasma cholinesterase variants by pancuronium bromide and some of its analogues.

Pancuronium bromide, a 3,17-diacetoxy-5 alpha-androstane, and three of its analogues, the 17-desoxy, the 3,17-dibutyryloxy and the 16-N-monoquaternary ammonium derivatives have been used as inhibitors of the usual and atypical plasma cholinesterase variants. In all cases the usual enzyme is more sensitive to inhibition by the substituted steroids than the dibucaine resistant enzyme. The relative affinities of the bis-quaternary ammonium compounds for either enzyme is in the order dibutyryloxy derivative > 17-desoxy derivative greater than or equal to pancuronium bromide. The monoquaternary compound has the least affinity of all the inhibitors for the usual enzyme but the greater affinity for the atypical enzyme. These observations show that the bis-quaternary compounds are very powerful differentiators of the variants. The monoquaternary derivative shows less differential inhibition, but provides additional evidence that the usual and dibucaine resistant variants differ in structure at or near their esteratic active site.