Ethanol Metabolism

ETHANOL METABOLISM Learning Objectives:

1. Explain the nutritional worth of alcohol usage

2. Write the pathway used in the conversion of ethanol to acetate and describe the properties of the enzymes involved.

3. Describe what is known about the mechanism underlying the flush response and

that associated with disulfuram usage. 5. Explain the metabolic basis underlying methanol toxicity. 6. Delineate the major physiological factors leading to the precipitation of alcoholic

ketosis. 7. Describe the metabolic chain of events leading to alcoholic ketosis. 8. Describe the major diagnostic markers for alcoholic ketosis and explain how these

differ from those for diabetic ketoacidosis. Key Words: Alcohol dehydrogenase Aldehyde dehydrogenase MEOS Cytochrome P-450 Anabuse Disulfuram Acetaldehyde Cirrhosis Antifreeze Ethylene glycol Oxalate Glycolate Methanol Wood alcohol Formaldehyde Formate Formyl-THF Systemic acidosis

Competitive inhibitor 4-Methylpyrazole Hemodialysis Alcoholic ketosis Hypovolemia Insulin Glucagon Adrenalin Cortisol Lactic acidosis Ketone bodies Acetoacetate B-Hydroxybutryate NADH/NAD ratio Lipolysis Malonyl CoA Carnitine acyltransferase B-Oxidation

Clinical Vignette

99:163 - Medical Biochemistry Rubenstein

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A 44 yr old woman drank approximately 720 ml of ethylene glycol in the form of antifreeze. She had previously attempted suicide by injecting the same substance into her buttocks. When admitted to the hospital, the patient was unresponsive and incontinent and was receiving ventilation. Her temperature was 37.1o, her pulse was 110/min., and her blood pressure 130/70. Her pupils were fixed and dilated, and she nad no corneal, gag, or deep-tendon reflexes. The serum sodium was 140 mM, the potassium 6.6 mM, the chloride 110 mM, and the bicarbonate 1mM (anion gap 29 mM). The serum creatinine was 2.8 mgldl and the lactate concentration was 10.1 mM. The osmolar gap was 84 mOsm/kg. Urinalysis revealed calcium oxalate crystals. The serum concentration of ethylene glycol was 2600 mg/liter. At admission, pH was 6.46, pCO2 was 18 mm Hg, pO2 was 327 mm Hg. She was being ventilated at the rate of 18 liters of O2/min. Two hours after admission, hemodialysis was begun and continued intermittently for 48 hrs. To reduce the conversion of ethylene glycol to its acid metabolites, ethanol was added to the dialysate and then given by gavage in a dose of 600 mg/kg of body weight followed by intragastric infusion at a rate of 200 mg/kg per hour for 37 hr. Renal failure did not develop, and the patient recovered completely. Blakeley et al. (1993) New England Journal of Medicine 328, 515-516.


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Overall Considerations

A. Ethanol is metabolized to acetate which is converted to acetyl-CoA. Therefore, ingested ethanol can only be converted to fatty acids or oxidized to CO2. It cannot be used for gluconeogenesis.

B. The caloric content of ethanol is 7 calories/gm, almost as high as that for fat. C. Hence, ethanol is a great source of energy via oxidative metabolism. The body

uses it efficiently when it amounts to less than about 15% of one’s total caloric intake.

D. Excessive use of alcohol often leads to malnutrition because one’s caloric needs

are satisfied by the alcohol, but nutrients necessary for proper health (sugars, amino acids, vitamins) are not ingested because the person’s hunger is satiated by the alcohol.

E. Evidently, alcohol in moderate amounts in some forms may actually be beneficial

(French paradox - red wine consumption). You can ask more about this in pharmacology.

II. Pathway


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A. Major site of alcohol metabolism is the liver

B. ADH (alcohol dehydrogenase)

1. Cytosolic

2. Requires Zn+2 for activity (interacts with the OH group) 3. Uses NAD

C. Aldehyde dehydrogenase

1. Mitochondrial 2. Requires NAD 3. Inhibited by Anabuse (disulfuram) - Causes build-up of acetaldehyde which

makes one very sick: vomiting, headache, etc. 4. Half of the Japanese have an isozyme of this enzyme with lower activity than

that found in Caucasians. Causes a “flush” response when alcohol is imbibed.

D. Microsomal Ethanol Oxidizing System (MEOS)

1. A cytochrome P-450 dependent enzyme. 2. Uses molecular oxygen and NADPH to oxidize the alcohol to acetaldehyde 3. Very high Km for ETOH (20 mM or 0.1% alcohol), so contribution to total

alcohol oxidation is probably pretty minimal.

E. Acetate produced from EtOH metabolism is released into the blood and is converted to acetyl CoA elsewhere.

III. Chronic alcohol abuse:

A. Changes in membrane fluidity of brain and heart B. Damage to the heart muscle, especially capacity of mitochondria to function

properly leading to heart failure. C. Cirrhosis of the liver and decreased liver function D. Thiamine deficiency


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METHANOL (WOOD ALCOHOL) POISONING

I. Sources of problem

A. Sterno, alcohol distilled from wood products (moonshine) B. Chronic alcoholics are often unable to purchase ethanol

C. A similar problem is caused by ingestion of antifreeze (ethylene glycol)

II. Biochemistry

A. Pathway of methanol and ethylene glycol metabolism

B. Consequences:

1. The formaldehyde generated is not the harmful agent in this situation. 2. Formaldehyde that is made from the methanol by alcohol dehydrogenase is

rapidly converted to formic acid. 3. Optic nerve is extremely sensitive to formic acid. Exposure causes

irreversible damage leading in worst case to blindness. 4. Formic acid continues to build up in bloodstream causing severe systemic

acidosis.

a. The only way formic acid can be eliminated is by urination or conversion to formyl-tetrahydrofolic acid.


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b. Formyl-THF synthase is not present in large amounts and cannot keep up with the production of formic acid that occurs following ingestion of a large amount of methanol.

III.Treatment

A. Prevent conversion of methanol to formaldehyde by alcohol dehydrogenase.

1. Administer large doses of EtOH i.v. which can then act as a competitive inhibitor of the ADH.

2. Administer nonmetabolizable agents like 4-methylpyrazole which also inhibit

the action of the enzyme. 3. Hemodialysis to cleanse toxins from the blood.

B. NaHCO3 was also routinely used in the past to treat the acidosis acutely.

However, there is currently controversy as to whether this is a good idea.

1.Intracellular pH, not blood pH is the most important factor. As long as blood is pumping through the lungs, protons will be pulled out of the cell and expelled from the body.

2. HCO3 equilibrates with CO2. A bolus of HCO3 will produce a sharp increase in serum CO2. This is freely diffusable across the blood brain barrier, and when it accumulates in the brain will further suppress breathing response preventing respiratory compensation for the acidosis.


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ALCOHOLIC KETOSIS

Wren et al., Am. J. Med. 91, 119 (1991) I. Precipitating conditions

A. Chronic starvation (Malnutrition)

1. Absence of liver glycogen 2. Induction of glucagon expression 3. Supression of insulin secretion 4. Gluconeogenesis

B. Binge drinking (100-200 g. EtOH/day))

C. Vomiting - induced hypovolemia

1. Cortisol release 2. Adrenalin release

II Biochemical observations

A. Large increase in the NADH/NAD ratio in the cytosol due to conversion of large amount of EtOH to acetate.

B. Lactic acidosis (often but not always)

C. Impaired ability to synthesize glucose from lactate

Lactate -------> Pyruvate ----------->OAA -----------> PEP

Malate D. Elevated ketone bodies (β-OH-Butyr. > AcAc) in the range of 6-10 mM.


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E. Acid base imbalance and electrolyte abnormalities

1. Vomiting can cause a metabolic alkalosis 2. Blood pH can be normal because acidity of ketone bodies will be

compensated by the alkalosis due to the vomiting. III. Alteration of normal fatty acid metabolism in the liver

A. Increased glucagon, decreased insulin, and increased catecholamines cause massive liberation of fatty acids from extrahepatic tissues into blood (3-5 mM) via the hormone sensitive lipase (cAMP-dependent)

B. High glucagon and low insulin inhibit synthesis of malonyl CoA.

1. Fatty acid synthesis is shut down 2. Carnitine acyltransferase (cytosol) is fully activated allowing fatty acids to

enter mitochondrion.

C. High concentration of fatty acids in liver

1. Feedback inhibit synthesis of fatty acids from acetyl-CoA being produced 2. Saturate the carnitine system in the liver. 3. Fatty acyl CoA builds up in the mitochondrion and saturates the β-oxidation

enzymes. 4. NADH and FADH2 saturate electron carriers of the ETS, and deplete the

mitochondrion of NAD and FAD. 5. Krebs cycle slows since

a. capacity for β-oxidation exceeds that for Krebs Cycle operation

b. since concentrations of electron carriers needed for oxidative decarboxylation reactions are decreased

c. Oxaloacetate needed for citrate synthase rxn. is being converted back to

malate due to high NADH/NAD ratio.

D. NADH from cytosol saturates the malate/aspartate shuttle further skewing the NADH/NAD ratio in the mitochondrion.


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IV. Ketosis

A. High concentration of NADH slows down ETS and Krebs Cycle. B. Acetyl CoA generated as a result of β-oxidation cannot be reconverted to fatty

acids nor efficiently degraded in the mitochondrion. C. Excess acetyl CoA thus converted to acetoacetate. D. High concentration of NADH allows further reduction of acetoacetate to β-

hydroxybutyrate. E. Differences between alcoholic ketosis and diabetic ketoacidosis.

1. In the diabetic situation, there is usually a systemic acidosis whereas in alcoholic ketosis, serum pH can be at or near normal value.

2. In the alcoholic situation, ratio of β-hydroxybutyrate/acetoacetate >> than in

case of diabetic. 3. Dipstick assay used to detect ketone bodies in urine does not detect β-

hydroxybutyrate. One needs to use an enzyme-based assay. Therefore, in the alcoholic patient, combination of normal pH coupled with use of a dipstick assay only can lead to a bad diagnosis and severe mismanagement of the patient.


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V. Treatment

A. Administer saline-glucose to break the metabolic spiral with addition of other ions as needed to correct electrolyte imbalance.

1. Blood becomes normoglycemic

2. Glucagon and adrenalin production is turned off; insulin production is turned

on. 3. Lipolysis no longer occurs in extrahepatic tissues and concentration of free

fatty acids declines causing cessation of β-oxidation and saturation of mitochondria with excess reducing agents and acetyl CoA.

4. In presence of insulin excess acetyl CoA disappears in hepatocyte.

a. Acetyl CoA ----------> Malonyl CoA---------->Fatty acids b. Malonyl CoA inhibits carnitine acyl transferase keeping free fatty acids

out of mitochondria thus suppressing β-oxidation. c. Formation of ketone bodies is suppressed due to removal of excess

acetyl CoA.

B. Insulin administration

1. Advocated by some to help break the catabolic spiral. 2. Can be dangerous, because once the body begins to equilibrate, the result

can be too much insulin leading to acute hypoglycemia and insulin shock.

C. Bicarbonate administration is not called for because of reasons stated above.

Clinical vignette A 50-year-old man who has been a chronic alcohol abuser for the past 10 years has developed a markedly diminished appetite for food. On a Saturday, according to his landlady, he became unusually irritable and confused after drinking two fifths of Scotch whiskey and eating very little. His landlady convinced him to call his doctor and accompanied him to the hospital. Upon examination, he had a heart-beat of 104 beats per minute, his blood pressure was 150/60, and he was in early congestive heart failure. His neck veins were abnormally dilated, and he had noticeable edema of the ankles and lower legs. His skin had a dry red appearance. He was poorly oriented to time, place, and person. The landlady told the doctor that the gentleman had seemed to become increasingly weak and lethargic over the past few months. A chest X-ray revealed a markedly enlarged heart.


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N. Diseases associated with Krebs Cycle: 1. Beriberi

a. The term means weakness or fatigue b, Three classifications of the disease

i. Dry beriberi: ♦ Patient unable to stand from a squatting position ♦ Signs and symptoms localized to the neuromuscular system

characterized by progressive polyneuritis and increasing involvement of different muscle groups.

ii. Wet beriberi: ♦ Neuromuscular complaints are associated with edema ♦ Patients often have marked loss of appetite, weakness, and

malaise. ♦ Some element of cardiac failure often present. This is a special

type of failure called high output failure that is brought about in part by peripheral vasodilation and the attempts by the heart to efficiently pump blood through this increased vascular space. Characterized clinically by a decreased circulation time in the presence of a failing circulatory system.

iii. Cardiac beriberi ♦ Above symptoms accompanied by cardiac decompensation and

often severe cardiac enlargement. iv. Wernickes syndrome: confusion, ataxia, and opthalmoplegia is often

seen with this disease. c. Biochemistry basis: i. Thiamine deficiency ii. Reasons: ♦ Thiamine deficient diets (especially in Oriental countries where a

staple of the diet is washed rice).


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♦ Chronic heavy alcohol use (chief cause of the disease in Western countries) which blocks uptake of thiamine from the intestine.

iii. Consequences ♦ Decreased thiamine impairs oxidation of pyruvate, operation of the

Krebs Cycle (a-ketoglutarate dehydrogenase) and hence ATP production in tissues that are highly dependent on oxidative metabolism: brain and nervous system.

♦ Decreased thiamine can impair the efficient operation of the non-

oxidative branch of the pentose phosphate shunt causing a pentose imbalance in sensitive tissues (e.g. CNS).

d. Treatment i. For dietary insufficiency, oral thiamine supplementation or inclusion of

thiamine rich foods. ii. For alcohol – based disease, a high dose of injected thiamine will bring

a marked resolution of symptoms, especially cardiac symptoms in a fairly short period of time. Abstention from any alcohol, and better nutrition are important long-term treatments.

iii. Note that alcohol can also directly affect the functioning of the heart

and chronic abuse can lead to a condition called alcoholic cardiomyopathy which is a non-high output dilated cardiomyopathy that is resistant to treatment by thiamine. If not too severe, total abstention can resolve the problem over a long period of time. If damage is too great, however, it is irreversible.


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SUMMARY - ALCOHOL METABOLISM (Rubenstein)

1. Calorically, alcohol has a value of 7 calories/gm and is the equivalent of fat. It can be used for energy production and excess is stored as fat, but it cannot be used to make glucose.

2. Zn-dependent alcohol dehydrogenase (liver) converts alcohol to the corresponding aldehyde

in an NAD-requiring reaction. For ethanol, the product is acetaldehyde. Aldehyde dehydrogenase the oxidizes the aldehyde to the acid.

3. A build-up of acetaldehyde at low levels causes a flush response. High levels can make

one violently ill. Anabuse and the antibiotic metronidazole inhibit aldehyde dehydrogenase and bring on violent vomiting when alcohol is imbibed.

4. Ingestion of methanol leads to elevated formic acid in the blood leading to a severe systemic

acidosis due to the slow metabolism of formate by the body. Formate first poisons the optic nerve and then leads to much more serious consequences. Antifreeze (ethylene glycol) leads to oxalate crystals in the blood with systemic acidosis which may cause renal failure.

5. Binge drinking coupled with malnutrition causes alcoholic ketosis. NADH/NAD levels

increase. Lack of insulin results in massive lipolysis with rampant β-oxidation producing acetyl CoA faster than it can be metabolized. The result is a ketosis characterized by elevate β-hydroxybutyrate/acetoacetate levels since the excess NADH will reduce more of the acetoacetate made in the liver mitochondria. The patient may not be acidotic since the vomiting that accompanies binge drinking may cause a compensating systemic alkalosis. The excess NADH will also interfere with gluconeogenesis

6. To treat the condition give i.v. glucose, fluids, and electrolytes. The glucose will elicit insulin

production. This will turn off glucagon production causing a cessation of lipolysis. It will also inhibit carnitine acyl transferase I preventing fatty acids from entering the mitochondrion thereby causing a cessation of b-oxidation. This will lead to deposition of fatty acids as TAGs and a gradual lowering of serum ketone bodies. Insulin treatment is also possible, but one then runs the risk of driving the patient into hypoglycemia due to excess insulin as recovery proceeds.

7. For systemic metabolic acidosis, administration of bicarbonate is not recommended usually

since one can easily overshoot leading to hypernatremia as the body’s metabolism is returned to normal. As long as perfusion and breathing are maintained, the patient should improve since the intracellular pH should be more near normal due to the continuous removal of acid from the body.


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Ethanol Metabolism