BIOCHEMICAL TESTS AND

THEIR SIGNIFICANCE
BY DR. TEJASHVI SETH.
 INTRODUCTION

• Biochemical tests are chemical pathology investigations which deals

with investigations of the metabolic abnormalities of the body in disease
states.

• Investigations are carried out by assays of various normal and abnormal

compounds found in body fluids viz. blood, urine, CSF, saliva etc.
 NEED FOR LABORATORY DIAGNOSTIC TESTS
• Aid in discovering occult disease.
• Preventing irreparable damage.
• Early diagnosis after onset of signs and symptoms.
• Differential diagnosis of various possible disease.
• Determining the stage of the disease.
• Estimating the activity of disease.
• Detecting the recurrence of the disease.
• Monitoring the effect of therapy.
ERRORS IN LABORATORY INVESTIGATIONS

Pre-analytic errors

Analytic errors

Post-analytic errors
 TYPES OF LABORATORY TESTS

• Discretionary or on-off tests : useful to support the diagnosis

• Biochemical profile : information on disease status of a patient
• Dynamic function tests : to measure body’s response to external
stimuli
• Screening tests : identify inborn errors of metabolism or to check
entry of toxic agents
• Metabolic work-up tests : to identify endrocrinology disorders
(Sonis, Fazio, Fang 1995)

Laboratory tests
Laboratory tests Laboratory tests
rarely ordered
frequently used by occasionally used
but significant to
dentists by dentists
dentists
LABORATORY TEST FREQUENTLY USED BY DENTISTS
 Complete blood count  Bleeding studies
• Hemoglobin • Prothrombin time
• Hematocrit • Partial thromboplastin time
• White blood count • Bleeding time
• Differential white blood cell count • Platelet count

 Fasting blood sugar  Hepatitis screening

 LABORATORY TESTS OCCASIONALLY USED BY DENTISTS
 Tests for disturbance in bone
• Calcium
• Phosphorous
• Alkaline phosphate
 Erythrocyte sedimentation rate
 Urinalysis
 Syphilis screening
 LABORATORY TESTS RARELY ORDERED BY BUT
SIGNIFICANT TO DENTISTS
 Enzymes
 Bilirubin
• Creatinine phosphokinase
 Reticulocyte count
• Serum glutamic-oxaloacetic
 Creatinine
transaminase (SGOT)
 Blood urea nitrogen
• Serum glutamic-pyruvic
transaminase (SGPT)  Acid phosphatase

• Lactic dehydrogenase
 SPECIMENS
The biological fluids employed in the clinical biochemistry laboratory
include
• Whole blood
• Plasma
• Serum
• Urine
• Cerebrospinal fluid
 COLLECTION OF SPECIMENS

 Blood
• Venous blood drawn from any prominent vein.
• Capillary blood obtained from finger or thumb.
• Arterial blood usually drawn under local anesthesia
 Urine

• Single specimen or for 24 hours

• Single specimens of urine normally collected in the morning are useful for
qualitative tests.

• 24 hour urine collection are employed for quantitative estimation of urinary

constituents.
 Cerebrospinal fluid

• collected by puncturing the interspace between 3rd and the 5th

lumbar lumbar vertebrae under aseptic conditions and local
anesthesia.
 BLOOD GLUCOSE ESTIMATION
 Fasting blood sugar
• used as a screening test for hyperglycemia.
• Normal values – 70-90 mg/100ml

 2 hours post prandial blood glucose :

• a more sensitive measurement of the hyperglycemia associated with
diabetes mellitus.
• Normal value is <140 mg/100ml
• Method for glucose determination : O-toluidine method

• Principle of estimation :
• Glucose + O- toluidine acid medium green coloured
complex.
• Measured in a colorimeter 630 nm.
 Oral Glucose tolerance test
• Used for definitive diagnosis of diabetes mellitus.

• It is performed on series of samples of blood and urine used to confirm the

diagnosis of diabetes mellitus in patient who do not exhibit consistently
elevated fasting blood sugar levels.
 GLYCOSYLATED HEMOGLOBIN ASSAY (GLYCOHEMOGLOBIN
TEST)

•Allows the determination of blood glucose status over 30 to 90 days prior to

collection of the blood sample.

•2 different glycosylated hemoglobin assays:

hemoglobin a 1 (Hba 1 ) test : normal value < 8%
hemoglobin a 1c (Hba 1c ) test : normal value < 6.0 % to 6.5 %

•It is used to monitor glycemic control in patients with previously diagnosed DM.
 GLUCOSE ESTIMATION IN URINE

• Glucose appear in urine when blood sugar exceeds 160-180 mg / dL.

• Glucosuria with hyperglycemia

• Glucosuria without hyperglycemia
Glucosuria with hyperglycemia caused by
• DM,
• Hyperthyroidism,
• General anesthesia,
• Intracranial lesions such as stroke.

Glucosuria without hyperglycemia seen in patient with

• renal diseases
• in 10-15% normal pregnancies,
• under stress,
• following injestion of high CBH meal.
Method: Benedict’s test.

Test principle:
• Benedict’s reagent contains potassium thiocyanate and potassium
ferrocyanide, in addition to sodium citrate, sodium carbonate and
copper sulphate.
• Glucose reduces cupric ions in the solution to cuprous ions, which
react with potassium thiocyanate to form coloured precipitates.
 INFLUENCE OF DIABETES ON PERIODONTIUM

• Enlarged gingiva
• Sessile or pedunculated gingival polyps
• Polypoid gingival proliferation
• Abcess formation
• Periodontitis
• Loosened teeth
(Hirschfeld I, 1934)
 BLOOD UREA NITROGEN
• Most widely used screening test for the evaluation of kidney function

• Aids in differential diagnosis of prerenal, renal and post-renal

hyperuraemia

• Decreased values in severe liver disease, protein malnutrition and

pregnancy
Elevated levels of urea are observed in:

• Pre-renal conditions: Diabetes mellitus, Dehydration,

Cardiac failure, Haematemesis, Severe
burns, High fever etc.

• Renal conditions: Disease of kidneys.

• Post-renal conditions: Enlargements of prostrate, stones in

urinary tract, tumour of bladder
• Test method used : Diacetyl monoxime method .
• Normal values: 5-12 mg/dl.
• Urea + diacetyl-monoxine yellow coloured
complex of
dioxime derivatives
• Measured at 520 nm
 INFLUENCE OF URAEMIA ON PERIODONTIUM

• Xerostomia
• Gingivitis
• Excessive plaque formation
• Uraemic stomatitis
• Periodontal diseases
• Maxillary and mandibular radiographic alterations
• Poor oral hygiene
(Cheng LP, et al. 2006)
 SERUM CALCIUM

• Normal range : 4.6-5.3 mg/dL

• Critical range : < 4.1 or > 5.9 mg/Dl
Increased Decreased

Paget’s disease Rickets

Metastatic bone tumor Vitamin D deficiency
Hyperparathyroidism Renal failure
Multiple myeloma Malabsorption
Osteomalacia
• Method: O-Cresolphthalein complexone method.

• Principle:
• Calcium + cresolphthalein complexon (CPC) complex
reagent containing dimethyl sulfoxide
and 8-hydroxyquinolone

• Measured at a wavelength 660 nm.

 SERUM CLACIUM AND PERIODONTAL DISEASE

• Lower dietary calcium intake and total serum calcium levels have
statistically significant association with more severe level of
periodontitis.
• The effects of calcium on periodontal disease likely related to alveolar
bone change which eventually results in greater clinical attachment loss
(Nishida M, et al. 2000)
 SERUM PHOSPHORUS

 Decreased values seen in

• preliminary hyperparathyroidism,
• rickets,
• fanconi’s syndrome (a disease associated with the defective absorption of phosphorus).

 Increased values seen in

• hypervitaminosis- D,
• hypoparathyroidism and
• in renal failure
• Normal value : Adults = 2.5-4.5 mg/dl
Children = 4.0-7.0 mg/dl
• Test method : precipitation of serum protein by trichloroacetic
acid
• protein free filtrate + molybdic acid phosphomolybdate
(containing inorganic reagent + phosphorous)
1-amino 2- naphthol-4
-sulphonic acid

reduced molybdenum blue

• Blue colour intensity is measured at 689 nm.
 SERUM PHOSPHOROUS AND PERIODONTAL
DISEASE

• Narges Nagash, et al. in 2017 studied the relationship between

periodontal disease and serum factors in patients undergoing
hemodialysis and concluded that

• Serum levels of phosphorous has a significant association with

clinical attachment level.
• With increase in severity of periodontitis the serum levels of
phosphorous also increases in these patients.

• Also these patients have higher dental calculus because of the

increase serum phosphorous and calcium
 ALKALINE PHOSPHATASE

 Clinical significance :

• Alkaline Phosphatase estimation are of interest in diagnosis of 2 groups

of conditions
- Hepatobiliary diseases.
- Bone diseases accommodated with increased osteoblastic activity.
• Increased levels :
• In infancy and childhood, Pregnancy and in healing fracture.

• Moderate increase is seen in hepatic conditions and elevation tends to be

more marked in post-hepatic conditions.

• Rickets - two to four times increase in the normal rate which falls back to
normal values after treatment with vitamin D.
• The highest level - in paget’s disease.

• Moderate rise – osteomalacia

• Very high level - bone cancer

• Slight to moderate elevations - hyperparathyroidism

• Decreased levels :

• In Hypophosphatasia( a rare inherited disorder).

• In acute and chronic leukemias

Method : Para nitrophenyl phosphate method (PNP).

Principle :
• Para nitrophenyl phosphate P-nitrophenoxide ions
(under alkaline conditons)
• exhibit yellow color.
• Intensity of yellow color is directly proportional to enzyme present in the
specimen and can be measured at 405 nm (violet filter).

Normal Range : 20-90 I.U. in adults

93-221 I.U. in children
 ALKALINE PHOSPHATASE AND PERIODONTIUM

•Alkaline phosphatase is an enzyme found in many cells of the periodontium,

including
-osteoblasts,
-fibroblasts,
-neutrophils.

•The concentration of enzyme alkaline phosphatase in GCF is significantly higher

in diseased sites than the healthy sites.
• Alkaline phosphatase activity increases somewhat in the inflamed
gingiva, since it is present in the inflammatory and endothelial cells,
and these are present in larger numbers in chronic gingivitis
 SERUM ACID PHOSPHATASE
• Clinical significance:
• In detecting and monitoring carcinoma of prostrates a disease currently
the third most frequent cause of death in males.
• In case of metastasis of prostatic carcinoma, total ACP activity may reach
40-50 times the upper limit of reference intervals. However in localized
cases of prostatic carcinoma only slightly raised levels of ACP are found.
• Moderate elevation of ACP is also seen in cases of
- Paget’s disease,
- hyperparathyroidism (with skeletal involvement)
- breast cancer (in women).
• Test method: P-nitrophenyl phosphate method.

• Principle:
• The enzyme acts on paranitrophenyl phosphate in citrate buffer at pH 4.9.
• The liberated nitrophenol after the incubation gives the measure of acid
phosphatase.

• Normal range: Total acid phosphatase: 0.9-12 I.U.

 SERUM ACID PHOSPHATASE AND
PERIODONTAL DISEASE
• Indicator of cellular damage in soft tissue of periodontium and inflamed
gingival tissue.
• Increased activity of acid phosphatase is consequence of destructive
process of alveolar bone and associated with the advanced stages of
development of the periodontal disease.
• Increased amount of serum acid phosphatase is released in gingival
crevicular fluid and saliva with increasing severity of the disease
( D S Pushparani, 2015)
 AMINOTRANSFERASES

• Clinically two most important transaminases are:

• Serum glutamic-oxalacetic transaminase (SGOT) or Aspartate

transferase (AST).

• Serum glutamic-pyruvic transaminase (SGPT) or Alanine transferase

(ALT).
• SGOT is rich in heart muscle.

• SGPT is rich in liver – also present in heart,

-kidney
-skeletal muscle.

• Serum AST and ALT levels are increased in viral hepatitis and other
forms of liver disease associated with hepatic necrosis.
ALT and AST also increase in case of
• alcoholic hepatitis,
• extrahepatic cholestasis and
• after administration of various drugs such as ampicillin, salicylates and
opiates.
 Normal value: SGOT = 8-40 IU/ L
SGPT = 5-35 IU/L
SGPT/ SGOT = <1
or ALT/AST = <1
• Method used: Reitman and Frankel’s method.

• Principle:
Keto acid formed by catalysation
(pyruvic acid in SGPT and oxaloacetic acid in case of SGOT
+
2, 4-dinitrophenyl hydrazine reagent (DNPH)

dinitrophenyl hydrazine (brown colour)

• Read intensity at 540 nm.

Determination helps in follwing:
• Differentiation between acute myocardial infarction and coronary
insufficiency.

• Diagnosis of acute MI when the ECG changes are not definitive and
difficult to interpret because of infarction.

• Diagnosis of extension of original infarction or of recurrent acute

infarction.
• If AST is elevated in both serum and spinal fluid, massive
parenchymal brain destruction is suggested.

• SGPT or ALT also increases in children with acute lymphoblastic

leukemia.
 AMINOTRANSFERASE AND PERIODONTAL
DISEASE
• Indicator of tissue necrosis

• AST levels increased during ligature induced experimental periodontitis

( chambers, et al. 1984)

• In experimental gingivitis in humans GCF samples harvested during the

development and resolution of the condition were significantly associated
with the gingival inflammation (Persson et al,1990)
 SERUM ALBUMIN

 Increased levels in dehydration.

 Decreased levels in
• renal diseases,
• nephritic renal insufficiency,
• severe malnutrition,
• pregnancy,
• burns
• diarrhea.

Normal range: 3.3 - 4.8 gm/dl

• Test method : Bromocresol green dye method
• Principle :
• Bromocresol green + albumin intense blue green coloured
complex
• Intenstiy measured at 628 nm
 SERUM ALBUMIN AND PERIODONTAL DISEASE

• Serum albumin has a significant inverse correlation with mean probing

depth.
(Narges Naghsh, et al. 2017)

• An investigation demonstrated that patients with severe periodontal

diseases were more likely to suffer from severe chronic kidney disease
(CKD) than less severe CKD. In addition, the association of serum
albumin content with periodontal status was demonstrated in progressive
stages of CKD (Ausavarungnirun et al. 2016).
 SERUM CREATININE
• Increased levels in renal failure.

• Concentration above 1.5 to 2.0 mg/dl is virtually diagnostic of renal

disease.

• Elevated values also seen in conditions like

- congestive heart failure,
- shock
-mechanical obstruction of the urinary tract.
• Normal values: 0.5 – 1.5 mg/dL

• Test method : Alkaline picrate method

• Principle :

creatinine + alkaline picrate creatine picrate

orange red coloured
complex
• colour intensity measured 530nm
 SERUM CREATININE AND PERIODONTAL
DISEASE

• Shimazaki Y, et al in 2013 studied the relationship between normal

serum creatinine concentration and periodontal disease in Japanese
middle-aged males

• concluded that there is significant inverse association between serum

creatinine concentration and periodontal diseases.
 DETERMINATION OF SERUM CHOLESTROL:

Elevated levels of serum cholesterol are seen with

• artherosclerosis,
• nephrosis,
• diabetes mellitus,
• obstructive jaundice
• myxedema.

 Decreased levels are observed in

• hyperthyroidism,
• malabsorption & anemia.
• Normal values: 150-225 mg/dL

• Test method : Acetic anhydride method

• Principle :

cholesterol + acetic andhyride (glacial acetic acid) green coloured

conc sulphuric acid complex

• colour intensity measured 560nm

 SERUM CHOLESTEROL AND PERIODONTAL DISEASES

• D S Kalsi, et al. in 2015 studied the association of lipid profile test values,
type-2 diabetes mellitus, and periodontitis and concluded that
• hyperlipidemia may be one of the factors associated with periodontitis
and that periodontitis may itself lead to abnormal serum lipid levels.
• Therefore, in addition to effects on diabetes, periodontitis may contribute
to elevated serum lipid levels and therefore potentially to systemic disease
arising from chronic hyperlipidemia.
 SERUM BILIRUBIN
• Clinical significance:
Increased levels in case of
• liver insufficiency,
• billiary obstruction or increased hemolysis,
• abnormal retention of bilirubin usually results in jaundice.

• Normal range:
Total bilirubin up to 1.0 mg/dl
Direct bilirubin up to 0.5 mg/dl
Indirect bilirubin up to 0.5 mg/dl.
• Method: Malloy and Evelyn.

• Principle: based on Van den bergh reaction.

• Bilirubin+ diazo reagent purple colour azobilirubin

• The optical densities of total test & direct test are measured against
respective blanks at 540 nm(green filter)
 CONCLUSION

• Laboratory studies are an extension of the physical examination in which

tissue, blood, urine or other specimens are obtained from the patient and
subjected to microscopic, biochemical, microbiological or immunological
examination.

• With increasing knowledge concerning to variety of diseases affecting the

oral cavity, greater use can be made of information obtained from such
laboratory tests in identifying the nature of disease.
 REFERENCES

• Text Book Of Medical Laboratory Techniques : GODKAR P. PRAFUL

• Burkit’s Oral Medicine: MALCOLM A. LYNCH
• Carranza’s Clinical Periodontology 11th edition: NEWMAN, TAKEI,
CARRANZA
• A Text Book Of Oral Pathology: WILLIAM G. SHAFERS
• Text Book of Biochemistry : U Sataynarayan
• Basic Applied Dental Biochemistry : William/Elliot
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• Yoshihiro Shimazaki, Mitoshi Kushiyama, Masatoshi Murakami, Yoshihisa
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