Serum Pseudocholinesterase Activity Research Articles

Relationship between pseudocholinesterase and lipids in hyperlipoproteinaemia

The biological r d e of serum pseudocholinesterase (PChE) is enigmatic: the protein is normally detected and defined in terms of its catalytic hydrolysis of choline esters but no necessity for such enzymic activity in serum has ever been demonstrated. However, the now well-established association between obesity and PChE in genetic and dietary animal models (Kutty et a[., 1981, 1983), and also in clinical situations (Cucuianu et al., 1968; Chu et al., 1978), suggests that PChE may play a significant part in the aetiology of coronary heart disease (CHD), a common end-stage consequence of obesity and hyperlipoproteinaemia. Serum was collected after an overnight fast from 66 patients with confirmed C H D (80% males; age 35-60 years) and 83 matched controls. Sixteen of the C H D patients were re-investigated after treatment for 2-3 months with diet, or medication, or both, to reduce serum lipids. Cholesterol and triglycerides were measured by standard methods. Lowdensity lipoprotein (LDL) was measured by a turbidimetric method (Walton & Scott, 1964) that included both LDL and very-low-density lipoprotein (VLDL). PChE was measured with butyrylthiocholine iodide substrate (Dietz et al., 1973). Serum PChE activities in the control (mean = 3.15 pmol of thiocholine/min per ml of serum; S.D. = 0.40) did not differ significantly with age or sex in adult subjects. However, 34 C H D patients (50%) had PChE activities exceeding the control 95th percentile, and all C H D patients’ PChE activities exceeded the control mean (Fig. 1). As expected, 50% of the patients were hypercholesterolaemic and 80% were hypertriglyceridaemic by accepted clinical criteria. Correlation analysis of the combined observations from patients and controls demonstrated that PChE is positively correlated with LDL (r = 0.51) and its associated lipids, cholesterol ( r = 0.48) and triglycerides (r = 0.52). Of even greater potential significance is that 18% of the C H D patients with significantly elevated PChE had normal lipid levels. During treatment, the patients’ PChE activities dropped from 4.7 f 0.34 to 3.9 k 0.26pmol of thiocholinelmin per ml ( I = 10.1) in concert with declines in cholesterol, triglyceride and total LDL levels. Animal models (Kutty et al., 1981, 1983) suggest that excess dietary intake is a major determinant of the PChE levels. Genetically obese (oblob), diabetic (dbldb) and normal mice with gold thioglucose-induced obesity exhibit a marked increase (50-150%) in liver and serum PChE activity when diet is unrestricted. Levels of PChE are almost normal when these hyperphagic animals are maintained on restricted diets. The parallel between these experimental models and hypertrophic obesity in humans is clear. However, the problem remains of non-obese, normolipidaemic C H D patients with significantly elevated serum PChE: what is the aetiology of hyperpseudocholinesterasaemia in these individuals and is non-dietary elevated serum PChE an independent risk factor in susceptibility to atheroscleortic heart disease? How could elevated PChE levels be implicated in atherosclerosis? It is possible that the true function of PChE

Pseudocholinesterase Activity and Atracurium V. Suxamethonium Block

Serum pseudocholinesterase activity in pregnant women was assayed spectrophotometrically using either propionylthiocholine or suxamethonium as substrate. All patients had a normal phenotype as indicated by normal dibucaine and fluoride numbers. However, the mean cholinesterase activity was lower than in the general population. There was a negative correlation between cholinesterase activity and the duration of neuromuscular blockade following suxamethonium, but no correlation was observed between the cholinesterase activity and the duration of block following atracurium.

Levels of serum lipids, apolipoproteins A-I and B and pseudocholinesterase activity and their discriminative values in patients with coronary by-pass operation

Serum lipids and apoproteins A-I and B were measured in 115 male patients and serum pseudocholinesterase activity (PChE) was determined in 83 patients with 3 vessel coronary artery disease (CAD). The control subjects were matched according to sex, smoking, relative weight and age and were free from heart disease. The CAD patients had significantly higher serum VLDL cholesterol and triglyceride levels and lower HDL cholesterol and apo A-1 levels and lower HDL to total cholesterol ratio than the controls. The concentrations of serum total cholesterol and LDL cholesterol were only slightly (6.4% and 8.8%, on an average) higher in CAD patients than in controls. The apo B levels of CAD patients were also slightly lower in patients than in controls. The CAD patients had slightly higher PChE activities than controls. The ratios of apo A-I to PChE and HDL cholesterol to PChE were significantly (about 30%, P < 0.001) lower in patients than in controls. In discriminant analysis between the groups HDL cholesterol and apo A-I showed the best (74% success in reclassifying the patients to correct groups), and total cholesterol, triglycerides, LDL cholesterol and apo B remarkably weak discriminating power among the single variables of serum lipids and lipoproteins. In discriminating analysis the apo A-I/PChE and HDL cholesterol/PChE ratios showed relatively high (77.1 and 71.1% success from the patients to correct groups) and serum PChE activity weak discriminating power. These results indicate that low levels of HDL cholesterol and apo A-I and the low ratio of HDL cholesterol to total cholesterol are the most potent metabolic risk factors for 3 vessel coronary artery disease in a population with relatively high serum total cholesterol level. The determinations of apo A-I/PChE and HDL cholesterol/PChE ratios may be an additional, valuable tool in discriminating the risk for CAD.

Rate assay for determination of serum pseudo-cholinesterase activity.

A simple and reproducible method for the determination of serum pseudo-cholinesterase activity was developed by making use of a stable substrate, p-hydroxybenzoylcholine, with p-hydroxybenzoate hydroxylase as a linked enzyme. The method is based on spectrophotometric measurement of the decrease of NADPH. p-Hydroxybenzoate released from p-hydroxybenzoylcholine is hydroxylated by the action of p-hydroxybenzoate hydroxylase in the presence of NADPH and O2 to produce 3,4-dihydroxybenzoate and NADP+. This method is superior to the conventional methods in that this substrate is extremely stable up to pH 9.0, which is close to the optimum pH for the assay (pH 8.0). Serum interference was resolved by the use of p-hydroxybenzoate hydroxylase as a linked enzyme. The Km value of pseudocholinesterase for p-hydroxybenzoylcholine is 1 X 10(-5) M. The results of our method and Garry's method (Clin. Chem. 11, 91-96, 1965) correlated well (r = 0.962). The within-run and between-run C.V. values were 2.1 and 2.7, respectively.

Increased plasma antithrombin III level in hyperlipidemic subjects

When compared to values obtained in healthy normolipidemic normal weight control subjects, the plasma antithrombin III level determined by immunological, clotting and thrombin-agarose diffusion techniques was found to be obviously decreased in patients with decompensated cirrhosis of the liver and slightly but significantly increased in hyperlipidemic and especially in hypertriglyceridemic subjects. Plasma antithrombin III was positively correlated with serum cholesterol level, the logarithm of serum triglyceride concentration and serum pseudocholinesterase activity. A weaker correlation between plasma fibrinogen and antithrombin III was noted in the investigated clinical material. It is suggested that the accelerated fatty acid and lipoprotein turnover occurring in many subjects with type IIb and type IV hyperlipoproteinemia might be accompanied by an enhanced hepatic protein synthesis involving various liver secretion enzymes and clotting factors as well as antithrombin III.

O-toluoylcholine as substrate for measurement of serum pseudo-cholinesterase activity

New synthetic substrates for serum pseudo-cholinesterase activity were compared with the common substrates for the routine assays, with regard to reactivity, specificity and stability; o- and m-toluoylcholine as well as o- and m-toluoyldimethylaminoethanol esters had selective specificities for pseudocholinesterase. The last three substrates, however, were unstable in solution at 4°C. On the other hand, o-toluoylcholine could be stored in solution for several days with no appreciable degradation, and it was extremely stable with regard to pH and temperature. No or little hydrolysis of o-toluoylcholine was observed by various enzymes other than pseudo-cholinesterase. The enzymatic method using o-toluoylcholine as substrate was reproducible, and the results correlated well with those obtained using butylthiocholine as substrate and 5,5'-dithiobis-(2-nitrobenzoic acid) as color reagent. In conclusion, o-toluoylcholine is a favorable substrate for the determination of serum pseudo-cholinesterase activity.

Similar behaviour of lecithin:cholesterol acyltrans- ferase and pseudocholinesterase in liver disease and hyperlipoproteinemia

Using exogenous substrate for its assay, lecithin: cholesterol acyltransferase (LCAT) was found to be decreased in liver disease and higher than normal in endogenous hypertriglyceridemia. LCAT activity was positively correlated with serum cholesterol and triglyceride. However in the six patients with excessive hypertriglyceridemia (type V), LCAT activity was lower than in type IV hyperlipoproteinemia. LCAT activity was not changed significantly in type II-a hyperlipoproteinemia. A striking parallel was noted between plasma LCAT and serum pseudocholinesterase activity. It is suggested that both these liver secretion enzymes might be induced by an accelerated turnover of serum lipids and lipoproteins. Pathogenical implications of these findings are briefly discussed.

New enzymatic assay of cholinesterase activity

A new enzymatic method for the determination of serum pseudo-cholinesterase activity is described. Choline, which is liberated from benzoylcholine as substrate by cholinesterase, is oxidized by choline oxidase to betaine with the simultaneous production of hydrogen peroxide, which oxidatively couples with 4-aminoantipyrine and phenol in the presence of peroxidase to yield a chromogen with maximal absorbance at 500 nm. The calibration curve is linear up to 1500 units per liter of serum. The method is reproducible, and the results correlate well with those obtained by the method using butyrylthiocholine as substrate and 5,5'-dithiobis-(2-nitrobenzoic acid) as color reagent.

PSEUDOCHOLINESTERASE ACTIVITY IN LIVER TRANSPLANTATION

Serum-pseudocholinesterase activity has been studied in five patients with orthotopic liver transplantation, and decreased activity has been demonstrated in the presence of acute rejection and cholangitis. Successful treatment of both complications was followed by restoration of normal enzyme activity. Monitoring of the serum-pseudocbolinesterase may be useful as an early warning in the follow-up of patients after liver transplantation.

Serum pseudocholinesterase activity in schizophrenia

Serum pseudocholinesterase activity was determined in clinically diagnosed untreated schizophrenic patients and compared with normals. It was concluded that pseudocholinesterase activity was increased significantly when acetylcholine, benzoylcholine and butyrylcholine were used as substrates. The ratios of hydrolyses rates of different substrates showed significant decrease in acetyl/butyrl, but benzoyl/butyrl and benzoyl/acetyl were not significantly changed. No correlation could be observed between the pseudocholinesterase activity and the liver function tests. It is also suggested that schizophrenia is not the result of the increase in cholinesterase activity, but rather it is due to a factor which also increases the pseudocholinesterase activity. Liver function tests were found to be normal except for the thymol turbidity test, which may be because of the alteration in the lipid globulin complex or in the morphologic damage of the liver.

Etude Critique d'Une Methode De Dosage Automatisee Des Pseudocholinesterases Selon La Reaction d'Hestrin

1. Attempts were made to develop an automated method which could perform quantitative measurements of total serum pseudocholinesterase (E.C. 3.1.1.8.) activity and dibucaïne number by use of the Hestrin reaction. 2. Unusual patterns of inhibition were observed with dibucaïne; some inhibited sera gave absorbance values which were higher than the absorbance of the substrate itself. This is due to the fact that certain sera contain “ester-like” substances which dialyze and thereafter react with hydroxylamine to produce a complex absorbing at the same wavelength as the acylhydroxamate ferric complex. 3. Application of the Hestrin reaction for the determination of total serum pseudocholinesterase activity and dibucaïne number with an automated system equipped with a dialyzer therefore requires corrections for serum blank values. Consequently, the procedure does not appear to be practical for large scale determinations of these parameters.

Etude critique d'une methode de dosage automatisee des pseudocholinesterases selon la reaction d'hestrin

1. Attempts were made to develop an automated method which could perform quantitative measurements of total serum pseudocholinesterase (E.C. 3.1.1.8.) activity and dibucaïne number by use of the Hestrin reaction. 2. Unusual patterns of inhibition were observed with dibucaïne; some inhibited sera gave absorbance values which were higher than the absorbance of the substrate itself. This is due to the fact that certain sera contain “ester-like” substances which dialyze and thereafter react with hydroxylamine to produce a complex absorbing at the same wavelength as the acylhydroxamate ferric complex. 3. Application of the Hestrin reaction for the determination of total serum pseudocholinesterase activity and dibucaïne number with an automated system equipped with a dialyzer therefore requires corrections for serum blank values. Consequently, the procedure does not appear to be practical for large scale determinations of these parameters.

Etude critique d'une methode de dosage automatisee des pseudocholinesterases selon la reaction d'hestrin*

Attempts were made to develop an automated method which could perform quantitative measurements of total serum pseudocholinesterase (E.C. 3.1.1.8.) activity and dibucaïne number by use of the Hestrin reaction. Unusual patterns of inhibition were observed with dibucaïne; some inhibited sera gave absorbance values which were higher than the absorbance of the substrate itself. This is due to the fact that certain sera contain “ester-like” substances which dialyze and thereafter react with hydroxylamine to produce a complex absorbing at the same wavelength as the acylhydroxamate ferric complex. Application of the Hestrin reaction for the determination of total serum pseudocholinesterase activity and dibucaïne number with an automated system equipped with a dialyzer therefore requires corrections for serum blank values. Consequently, the procedure does not appear to be practical for large scale determinations of these parameters.

Serum proteins in human trichinosis

Of 200 patients suffering from trichinosis, the course of the disease was mild in one hundred, moderate in sixty-six and severe in thirty-four. The following laboratory tests were performed serially: serum total protein and its electrophoretic pattern, thymol turbidity, serum pseudocholinesterase activity and serologic tests (i.e., complement fixation and precipitation). Proteinuria was estimated by the tannin method. The greatest decrease in serum total protein, especially albumin, was found in the fourth and fifth week of the disease and was related to its severity. Proteinuria was also more prominent in severe cases of trichinosis. A relative and absolute increase in gamma globulins paralleled the increased titer of serologic tests and abnormal thymol turbidity test. In thirty-five persons, follow-up examinations were made within one to twenty months after discharge and almost complete normalization of the previously abnormal tests was found.

Pseudocholinesterase activity in thyroid disease.

The present evidence indicates that serum pseudocholinesterase activity is increased to about 20% above the normal level in thyrotoxicosis and decreased by about 30% in myxoedema. Treatment restores the esterase level to the average mean activity of random normal as the patient becomes euthyroid. There is no evidence in the present investigation that any of the pseudocholinesterase variants modify these conclusions.

Two Further Serum Pseudocholinesterase Phenotypes as Causes of Suxamethonium Apnoea

Suxamethonium, a muscle relaxant whose properties were first described by Bo vet and his colleagues in 1949, has proved useful in anaesthesia because the duration of its action is short. Thus after the administration of 50-100 mg. the muscle paralysis and associated apnoea last only about two minutes. This brief action is due to the rapid destruction of the drug by serum pseudo cholinesterase (Bovet-Nitti, 1949). Rare individuals exist who possess a low level of serum pseudocholin esterase, and who, after the usual dose of suxa methonium, develop a prolonged apnoea which may last several hours (Bourne, Collier, and Somers, 1952 ; Evans, Gray, Lehmann, and Silk, 1952). This low serum pseudocholinesterase activity and the associated suxa methonium sensitivity have been shown to be inherited (Lehmann and Ryan, 1956). Kalow and his colleagues have shown that these individuals are homozygotes for a rare gene, and possess not a low level of the normal enzyme but an abnormal serum pseudocholinesterase. This abnormal or atypical enzyme differs from the usual type in two ways: it is less active in hydrolysing a large number of substrates, including suxamethonium (Davies, Marton, and Kalow, 1960), and more resistant to a majority of cholinesterase inhibitors (Kalow and Davies, 1958). Kalow and Genest (1957) have proposed the use of one such inhibitor, the local anaesthetic dibucaine (cinchocaine, nupercaine ), as a screening test to identify these homozygotes. In this test a 10~* M concentration of dibucaine inhibits the normal enzyme by about 80% and that found in suxamethonium sensitive individuals by only 20%. The percentage inhibition is called the dibucaine number, or DN. H?t?rozygotes, possessing a mixture of both enzymes, have dibucaine numbers of between 50 and 69. Various population surveys have shown that the suxamethonium sensitive homozygote occurs with a frequency of about 1 in 2,800, while 3.8% of the population are h?t?rozygotes (Kalow and Gunn, 1958 ; Kattamis, Zannos-Mariolea, Franco, Liddell, Lehmann, and Davies, 1962). Another genetic variant has been described in which the homozygote, detected because of a prolonged apnoea following suxamethonium administration, showed a complete absence of serum pseudocholin esterase activity (Liddell, Lehmann, and Silk, 1962). The gene responsible has been called the silent pseudocholinesterase gene. Harris and Whittaker (1961, 1962) have described a third variant of the enzyme. They discovered that sodium fluoride, like dibucaine, inhibits the atypical enzyme less strongly than normal pseudocholinesterase. Thus a 5 x 10~5 M concentration of sodium fluoride inhibits the normal enzyme by about 60%, the atypical enzyme by about 25%, and sera from h?t?rozygotes by abou 45%. Harris and Whittaker called this inhibition the fluoride number, or FN. When sera from large numbers of persons were examined a few individuals were found the dibucaine numbers of whose sera were sl ghtly lower and the fluoride numbers markedly lower than expected. Harris and Whittaker suggested that this was due to the presence of another variant of pseudocholinesterase with a slightly lower dibucaine number and markedly lower fluoride number than normal. Using both the dibucaine and fluoride te hniques, five types of individual can be recognized : 1. DN 80, FN 63 ; normal enzyme. 2. DN about 74, FN about 52 ; h?t?rozygotes possessing both he normal enzyme and the fluoride-resistant enzyme described by Harris and Whittaker. 3. DN about 64, FN about 48 ; h?t?rozygotes possessing oth the normal and atypical enzymes. 4. DN about 54, FN about 34 ; h?t?rozygotes possessing both the atypical and the fluoride-resistant enzymes. 5. DN about 20, FN about 23 ; homozygotes for the atypical enzyme.

A simple colorimetric method for estimating serum pseudocholinesterase

A simple colorimetric method for estimating serum pseudocholinesterase is presented. It depends on the hydrolysis of phenyl benzoate at pH 8.6 by the serum with the subsequent determination of the liberated phenol by the use of Folin and Ciocalteu's reagent. The evidence that the results are a measure of serum pseudocholinesterase activity is (1) complete inhibition of enzymic hydrolysis by prostigmin and (2) a comparison between the serum esterase activity and the pseudocholinesterase as determined by the Warburg technique.