Indian Journal of Clinical Biochemistry 1991, 6, 73-75

E R Y T H R O C Y T E A L D E H Y D E D E H Y D R O G E N A S E A C T I V I T Y I N

A L C O H O L I C S A N D N O N - A L C O H O L I C S A N D I T S V A L U E AS A

B I O C H E M I C A L M A R K E R I N A L C O H O L I S M

GOVIND SHARMA, R. L. TRIPATHI, K. S. S. SAIBABA and L. M. SRIVASTAVA* Department of Biochemistry All India Institute of Medical Sciences NEW DELHI - 110 029 INDIA.

Received on October 25, 1990

ABSTRACT

Erythrocyte aldehyde dehydrogenase (ALDH) activity in a normal reference population from North India was determined from results of 22 healthy subjects (12 male and 10 female). We compared these data with results of 19 alcoholic patients (13 male and 6 female). The mean erythrocyte ALDH activity of male and female alcohol abusers was 16.2% and 20.9% lower than the mean values for healthy subjects, respectively (P <0.001 and <0.01). It is proposed that e ~ ALDH activity may be a useful clinical chemical marker of alcohol abuse and alcoholism.

INTRODUCTION

Acetaldehyde, the first metabolite in ethanol metabolism, plays an important role in alcohol-related toxicity in humans (1, 2). Acetaldehyde is produced in vivo from ethanol by alcohol dehydrogenase (ADH). Most of the acetaldehyde is rapidly metabolized to acetate by a NAD-dependent aldehyde dehydrogenase (ALDH). Blood acetaldehyde concentrations have been thought to increase in alcoholics as a consequence of either an enhanced rate of ethanol metabolism (3, 4)'and/or a reduced acetaldehyde metabolism clue to a decreased hepatic ALDH activity (5, 7). Recent studies show that alcoholics have a decreased ALDH activity in liver (5, 6) as well as in blood (8, 9).

This study was undertaken to investigate the value of erythrocyte ALDH activity levels in alcoholics and non-alcoholic controls from a random sample from North India for the purpose of diagnosis and prognosis of alcohol abuse.

*Correspondence Address : Dr: L. M. Srivastava Additional Professor Department of Biochemistry All India Institute of Medical Sciences NEW DELHI- 110 029.

74 SHARMA ET AL 1991

MATERIALS AND METHODS

Thirteen male chronic alcohol abusers ranging in age from 25 to 56 years (mean 33.2 yrs.) and six female alcohol abusers ranging in age from 22 to 43 years (mean 31.5 yrs.), without clinical evidence of liver disease, were studied. Twelve normal healthy non- alcoholic males ranging in age from 23 to 52 years (mean 32.5 yrs.) and ten normal healthy non-alcoholic females ranging in age from 24 to 42 years (mean 30.2 yrs.) were included in the study to obtain normal control values of erythrocyte ALDH activity.

EDTA blood was collected from these subjects by venous puncture. The plasma and buffy coat were separated from the erythrocytes by centrifugation at 2000 g for 10 min and the red cells were washed three times with one volume of 0.25 M sucrose. Thereafter, one minilitre of the washed cells were mixed with one volume of 100 mM sodium phosphate buffer, pH 6.0, containing lmM EDTA and 1 mM dithioerythtitol. The cell suspension was frozen at -20~ and thawed to break the cell membrane. For the preparation of stroma- free hemolysate, a 500/~l aliquot of the cells were mixed with 1 volume of chloroform. After centrifugation of the mixture at 27,000 g for 15 min, the top hemolysate layer was carefully removed with a pipette and used further for hemoglobin determination as well as for the preparation of hemoglobin-free lysate. This 500 #1 hemolysate was applied to a small column (1 x5 cm.) of CM Sephadex G - 5 0 equilibrated with 30 mM sodium phosphate buffer, pH 6.0, containing 1 mM EDTA and lmM dithioerythritol. Subsequently, 2 ml of the same buffer was dropwise passed through the column. The whole procedure was carried out at 4~ The first 500 t~l of the eluate which contains no ALDH activity was discarded. The hemoglobin-free eluate containing ALDH activity was collected and determination of enzyme activity was carried out by the method of Agarwal et al (10). The data were evaluated statistically by applying the student's 't ' test.

RESULTS AND DISCUSSION

As shown in Table 1, the mean ALDH enzyme activity in male and female alcohol abusers w~s found to be 0.1404-0.004 units/g Hb and 0.121 -r units/g Hb, respectively. Whereas in non-alcoholic male and female controls the enzyme activity was found to be 0.167 4-0.007 units/g Hb and 0.1534-0.004 units/g Hb, respectively. One unit of enzyme activity is defined as the formation of 1 t~mol NADH H +/min under the assay conditions and expressed per g hemoglobin. On comparison, mean erythrocyte ALDH activity of the alcoholic male and female abusers was found to be lowered by 16.17% and 20.92% than the mean value for healthy male and female controls respectively. The reduction in ALDH activity was found to be statistically significant (P values: <0.001 and <0.01).

The observed lower erythrocyte ALDH activity in female alcoholics and non-alcoholic controls confirms earlier studies (8,11).

This study also confirms previous reports that erythrocyte ALDH activity is significandy reduced in alcohol abusers, chronic alcoholics and subjects with other alcohol- related disorders (8,9,11,12). Whether a reduced ALDH activity represents a primary genetic defect in ~coholism or it is the consequence of alcohol-related tissue damage remains to be clearly understood. Assay of red cell ALDH activity may however, offer a potential

Vol. 6 Aldehyde dehydrogenase as alcoholism marker 75

TABLE 1: Erythrocyte Aldehyde Dehydrogenase (ALDH) Activity in Alcoholic and Non-Alcoholic Subjects

Subjects ALDH Activity (units/g Hb)

Male Female

Non-alcoholic controls

Alcoholics

0.167 0.153 4- 4-

0.007 0.004 (12) 00)

0.140 0.121 4-

0.004** 0.010" (13) (6)

[16.17] [20.92]

Note: 1. Values are mean-t-S.D, 2. Figures in small parenthesis indicate sample size. 3. Figures in capital parenthesis indicate per cent change. 4. *P < 0.01.

**P < 0.(301.

marker to monitor alcohol abuse during treatment and management of the disease alcohofism.

ACKNOWLEDGEMENT

The authors thank Prof. D. P. Agarwal, Institute of Human Genetics, University of Hamburg, Germany for his interest and support.

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