Review

The Physiological Sources of, Clinical

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Significance of, and Laboratory-Testing Methods
for Determining Enzyme Levels
Muneeza A. Esani, MHA, MT(ASCP)

ABSTRACT discuss the physiological sources, clinical significance, and laboratory-

testing methodology for lactate dehydrogenase (LD), aspartate
Enzymes are organic molecules that accelerate biochemical reactions aminotransferase (AST), alanine aminotransferase (ALT), alkaline
but emerge from the reaction unchanged. Abnormal levels of plasma phosphatase (ALP), and gamma-glutamyltransferase (GGT).
enzymes are highly suggestive of damaged cells and provide clues
to parts of the body that may be involved in disease processes. Keywords: enzymes, lactate dehydrogenase, aspartate
Enzyme levels are measured in the clinical laboratory to identify the aminotransferase, alanine aminotransferase, alkaline phosphatase,
site of damage and to quantify the amount of damage. This paper will gamma-glutamyltransferase

Enzymes are organic molecules that accelerate biochemical The aminotransferases are a group of enzymes that
reactions but emerge from the reaction unchanged. For catalyze the conversion of amino acids to 2-oxo-acids by
instance, lactate dehydrogenase (LD) is a tetrameric enzyme transfer of amino groups.3 The highest levels of aspartate
with two distinct subunits, namely, M and H. It is found in aminotransferase (AST) are present in liver, cardiac
cardiac and skeletal muscles, liver, kidneys, erythrocytes, tissue, and skeletal muscle, with smaller amounts present
leukocytes, lungs, lymph nodes, spleen, and brain; it in the kidneys, pancreas, and erythrocytes. Alanine
catalyzes the redox conversion of lactate to pyruvate. LD aminotransferase (ALT), however, is predominantly present
has 6 isoenzymes that have been identified to date in the in the liver.2,4
literature. LD1 consists of HHHH subunits and LD2 consists
of HHHM subunits; both are predominantly found in the Alkaline phosphatase (ALP) catalyzes the hydrolysis of
heart and the erythrocytes. LD3 consists of HHMM subunits various phosphomonoesters at an alkaline pH. The highest
and is found in the lungs. LD4 and LD5 consist of HMMM concentrations of ALP are found in bone, liver, spleen,
and MMMM subunits, respectively; both are found in the intestine, placenta, and kidneys. ALP contains a number of
liver.1 LD catalyzes the reversible conversion of pyruvate to isoenzymes, with bone, liver, and placenta types being the
lactate in a redox reaction.2 most extensively studied.2 Isoenzyme analysis is of limited
clinical use due to ineffective methods of separation and
the availability of more efficient tests for clinical diagnosis.

Gamma-glutamyltransferase (GGT) catalyzes the transfer

DOI: 10.1309/LMBR83WM3GNJEDLS
of the γ-glutamyl residue from γ-glutamyl peptides to
Abbreviations amino acids2; this epithelial enzyme is responsible for the
LD, lactate dehydrogenase; AST, aspartate aminotransferase; ALT, catabolism of extracellular glutathione.5 GGT is present in
alanine aminotransferase; ALP, alkaline phosphatase; GGT, gamma- kidney, brain, prostate, pancreas, and liver tissue.2
glutamyltransferase; MI, myocardial infarction; NAD, nicotinamide
adenine dinucleotide; reduced nicotinamide adenine dinucleotide,
NADH

Department of Clinical Laboratory Science, University of TX, Medical

Branch, Galveston, TX
Clinical Significance
*To whom correspondence should be addressed. LD levels are increased in several disorders due to its
E-mail: [email protected] presence in a variety of tissues; the most dramatic

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 Review

increase is observed in prehepatic disorders, hemolytic

disease, testicular and germ cell tumors, and acute
myocardial infarction (MI). LD levels increase 12 hours
Testing Methods and

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after an MI and peak in 2 days, returning back to normal Interferences
levels in 7 to 14 days. Normally, plasma LD2 levels
predominate in blood, followed by LD1 levels; however, The measurement of enzymes is standardized, simple,
damaged cardiac tissue in MI causes LD1 levels to and inexpensive; it requires no special preparation for the
increase, which triggers a reversal in which LD1 levels patient.4 Due to the small quantity of enzymes normally
become greater than LD2 levels. Isoenzyme analysis has available in blood, the rate of reaction is measured
now become outdated due to the availability of more photometrically, related to the activity of the enzyme. This
sensitive and specific cardiac markers such as troponin enzyme activity is proportional to the concentration of the
for the diagnosis of MIs. Currently, LD is mostly used by enzyme.2 Serum or heparinized plasma is the preferred
laboratorians as a nonspecific screening or monitoring type of sample for enzyme analysis.
tool to determine the extent of tissue damage. The
Reference range for LD is 125 to 220 U/L. 2 Laboratorians can measure LD activity by gauging
the absorbance of coenzyme nicotinamide adenine
ALT and AST are clinically significant aminotransferases.3 dinucleotide (NAD) as it is changed to reduced
Both are markers of liver disease; however, ALT is more nicotinamide adenine dinucleotide (NADH) while lactate is
liver specific than AST.4 AST levels are also significantly oxidized to pyruvate in the following reaction:
increased in skeletomuscular disorders and pulmonary
embolism. In acute inflammatory conditions of the liver,
ALT levels are higher than AST levels because ALT
has a half-life of 16 to 24 hours. 2 Moreover; increased This reaction is derived from glycolysis, which is the
ALT levels have been associated with deaths from liver biochemical pathway for metabolism of glucose in all living
disease5 and all-cause mortality6 in some studies. The cells. Samples must be free of hemolysis because LD
reference range for ALT is 7 to 45 U/L and that of AST is levels are 100- to 150-fold higher in erythrocytes than in
5 to 35 U/L. 2 serum. Also, samples must be stored at room temperature
and analyzed within 48 hours of sample collection.2
ALP levels are highly increased in hepatobiliary and
bone disorders. Levels are normally increased during AST and ALT activity are measured by measuring the
pregnancy and in growing children due to the placental change in absorbance as aspartate and alanine transfer
and bone fractions, respectively. Intestinal ALP levels their amino group to the appropriate respective α-keto
increase in disorders of the digestive tract and liver. ALP acids. Pyridoxal-5-phosphate is the coenzyme for these
levels are decreased in inherited hyperphosphatasia. 2 reactions.
The reference range for ALP varies with age and sex,
with higher levels for younger compared with older
individuals and for men compared to women, probably
due to bone-fraction measurements.

GGT levels are mainly evaluated in hepatobiliary and

liver disorders. Levels are also increased in patients with
chronic alcoholism who take enzyme-inducing drugs Change in absorbance is directly proportional to the
such as warfarin and phenytoin, as well as in conditions concentration of ALT and AST.2
such as pancreatitis and diabetes mellitus. 2,7 Moreover,
high GGT levels are also associated with high rates of ALT and AST reactions are extremely important for
all-cause mortality.6 The reference range for GGT is 6 to synthesis and degradation of amino acids; oxaloacetate
55 U/L for men and 5 to 38 U/L for women. Values are and pyruvate are oxidized by the tricarboxylic cycle (Krebs
lower in females due to higher levels of estrogen and cycle) to provide intermediate products that generate
progesterone. 2 energy for living cells. Samples must be free of hemolysis
for AST; however, ALT levels are unaffected by any
interference from erythrocytes. For best results, samples

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Review

must be stored at refrigerated temperatures after collection tions of the liver. LD levels are most predominant in pre-
and be analyzed within 3 to 4 days.2 hepatic and other hemolytic disorders due to its presence
in erythrocytes. ALT and AST are predominant in hepatic

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ALP activity is measured by a method devised by disorders, with ALT being more liver specific than ALT.
Bowers and McComb8 that involves calculation of ALP and GGT are increased in posthepatic disorders with
activity based on molar absorptivity of p-nitrophenol. GGT more so with chronic alcoholism. Increased activity of
Para-nitrophenylphosphate, a colorless compound, is these enzymes provides clues to the source of the clinical
hydrolyzed to a yellow-colored p-nitrophenol, and increase problem and can be extremely valuable in diagnosis of the
in absorbance can be measured in the following reaction: patient’s condition. LM

The increase in absorbance of p-nitrophenol is directly References

proportional to ALP activity. Samples collected for ALP
1. Quistorff B, Grunnet N. The isoenzyme pattern of LDH does not play
analysis must be free of hemolysis and must be analyzed a physiological role; except perhaps during fast transitions in energy
soon after collection. Ingestion by the patient of a high-fat metabolism. Aging. 2011;3(5):457-460.
meal before specimen collection may also cause falsely 2. Johnson-Davis KL. Enzymes. In: Bishop ML, Fody EP, Schoeff LE,
increased values due to the intestinal fraction.2 However, eds. Clinical Chemistry: Principles, Techniques, and Correlations. 7th
ed. Philadelphia: Lippincott Williams & Wilkins. 2013;262-289.
fasting specimens are not recommended because the
3. Renze B, Mauro P. Enzyme and rate analyses. In: Tietz Textbook
interference is negligible. of Clinical Chemistry and Molecular Diagnostics. 5th ed. St. Louis:
Elsevier Saunders; 2012.
GGT activity is measured when γ-glytamyl-p-nitroanilide 4. Kunutsor SK, Apekey TA, Walley J. Liver aminotranferases and risk
of incident type 2 diabetes: a systematic review and meta-analysis.
transfers its γ-glytamyl residue to glycylglycine. This results
Am J Epidemiol. 2013;178(2):159-171.
in the formation of a chromagen, p-Nitroaniline, as shown
5. Ruhl CE, Everhart JE. Elevated serum alanine aminotransferase and
in the following reaction: γ-glutamyltransferase and mortality in the United States population.
Gastroenterology. 2009;136:477-485.
6. Kim HC, Nam CM, Jee SH, Han KH, Oh DK, Suh I. Normal serum
aminotransferase concentration and risk of mortality from liver
diseases: prospective cohort study. BMJ. 2004;328:983.
The absorbance of p-Nitroaniline is directly proportional to 7. Tynjälä J, Kangastupa P, Laatikainen T, Aalto M, Niemelä O. Effect
the activity of GGT, which is stable in serum samples for 1 of age and gender on the relationship between alcohol consumption
week at refrigerated temperatures. and serum GGT: time to recalibrate goals for normal ranges. Alcohol
Alcohol. 2012;47(5):558-562.
8. Bowers GN, McComb, RB. Measurement of total alkaline
Overall, LD, ALT, AST, ALP, and GGT are enzymes that are phosphatase activity in human serum. Clin Chem. 1975;21:1988-
clinically significant in various disorders including condi- 1995.

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