Unit 5- Ocular Biochemistry | 2nd Semester Bachelor of Optometry

Blood Sugar – Significance of Its Estimation

Blood Sugar Ranges

Introduction

Blood sugar, or blood glucose, refers to the concentration of glucose present in the blood at any given time. Glucose is the body’s primary source of energy, particularly for the brain and red blood cells, which depend almost entirely on it for normal functioning. The measurement of blood sugar is a critical aspect of clinical biochemistry because it provides valuable insight into metabolic health, endocrine function, and the presence of diseases such as diabetes mellitus. Understanding the significance of estimating blood sugar levels is essential for diagnosis, treatment planning, and monitoring of various metabolic disorders.

Normal Blood Sugar Levels

• Fasting blood glucose: 70–100 mg/dL (3.9–5.6 mmol/L)
• Postprandial (2 hours after a meal): < 140 mg/dL (< 7.8 mmol/L)
• Random blood glucose: usually < 200 mg/dL in healthy individuals

These values may slightly vary depending on laboratory methods and clinical guidelines, but they form the baseline for interpreting blood glucose results.

Importance of Glucose in the Body

Glucose plays a central role in metabolism. It is derived from dietary carbohydrates, stored as glycogen in the liver and muscles, and produced endogenously via gluconeogenesis. It is oxidized during glycolysis and the citric acid cycle to produce ATP, the universal energy currency of cells. The regulation of blood glucose involves a balance between glucose intake, utilization, and storage, orchestrated by hormones such as insulin, glucagon, cortisol, epinephrine, and growth hormone.

Why Estimate Blood Sugar?

Estimating blood sugar is important in several clinical contexts:

• Diagnosis of diabetes mellitus – Persistent elevation of fasting or postprandial blood glucose is diagnostic of diabetes.
• Screening in high-risk individuals – People with obesity, a family history of diabetes, or other risk factors benefit from early detection through blood glucose testing.
• Monitoring therapy – Diabetic patients require regular glucose monitoring to ensure effective control and prevention of complications.
• Detection of hypoglycemia – Low blood glucose can be life-threatening and requires rapid diagnosis and treatment.
• Critical care assessment – Stress hyperglycemia is common in critically ill patients, and glucose monitoring helps guide management.

Methods of Blood Sugar Estimation

Clinical biochemistry laboratories use several methods to measure blood glucose levels, each with its own advantages:

• Glucose oxidase method – Specific and widely used, based on the oxidation of glucose to gluconic acid and hydrogen peroxide.
• Hexokinase method – Considered the reference method due to its accuracy and specificity.
• Oral glucose tolerance test (OGTT) – Measures the body’s ability to handle a glucose load, useful in diagnosing impaired glucose tolerance.
• Point-of-care glucometers – Convenient for bedside or home monitoring, though less accurate than laboratory methods.

Significance in Diabetes Mellitus

Diabetes mellitus is characterized by chronic hyperglycemia resulting from defects in insulin secretion, insulin action, or both. Persistent high blood sugar damages blood vessels and nerves, leading to complications such as retinopathy, nephropathy, neuropathy, and cardiovascular disease. Blood glucose estimation is therefore central to:

• Diagnosing the disease (fasting glucose ≥ 126 mg/dL or random glucose ≥ 200 mg/dL with symptoms)
• Monitoring glycemic control in combination with glycated hemoglobin (HbA1c)
• Adjusting treatment regimens to prevent complications

Significance in Hypoglycemia

Hypoglycemia, defined as blood glucose below 70 mg/dL, may result from excessive insulin administration, certain medications, prolonged fasting, critical illnesses, or endocrine disorders. Symptoms include sweating, palpitations, confusion, seizures, and loss of consciousness. Accurate measurement of low blood glucose is vital for prompt intervention to prevent brain damage or death.

Blood Sugar Fluctuations in Other Conditions

Blood sugar estimation is also significant in:

• Gestational diabetes – Screening pregnant women prevents complications for both mother and child.
• Acute illnesses – Severe infections, myocardial infarction, and stroke can cause transient hyperglycemia due to stress hormone release.
• Endocrine disorders – Conditions like Cushing’s syndrome and pheochromocytoma affect glucose regulation.
• Liver disease – Impaired glycogen metabolism affects blood sugar stability.

Clinical Interpretation

While interpreting blood glucose values, clinicians must consider factors such as recent food intake, medications, and concurrent illnesses. For example, steroid therapy can elevate blood sugar, while insulin and oral hypoglycemic drugs lower it. Laboratory error, improper sample handling, and delayed processing can also affect results due to ongoing glycolysis in the sample.

Complications of Abnormal Blood Sugar

Persistent hyperglycemia leads to glycation of proteins, oxidative stress, and endothelial dysfunction, which are central to the development of diabetic complications. On the other hand, recurrent hypoglycemia can cause cognitive decline and increase the risk of accidents and cardiovascular events.

Role in Preventive Medicine

Blood glucose estimation is not only diagnostic but also preventive. Early identification of impaired fasting glucose (100–125 mg/dL) or impaired glucose tolerance enables lifestyle interventions such as diet modification, increased physical activity, and weight loss, which can delay or prevent the onset of type 2 diabetes.

Modern Trends in Blood Sugar Monitoring

Technological advancements have introduced continuous glucose monitoring (CGM) systems that measure interstitial glucose every few minutes and provide trends, alerts, and insights for better control. Such innovations are improving outcomes for both type 1 and type 2 diabetes patients.

Conclusion

Blood sugar estimation is a cornerstone of clinical biochemistry, with wide-ranging significance in diagnosing, monitoring, and preventing metabolic diseases. Accurate measurement helps guide therapy, prevent complications, and improve quality of life. With the global rise in diabetes prevalence, the importance of regular blood glucose monitoring in both clinical and community settings cannot be overstated.

Urea – Significance of its Estimation

Blood Urea Nitrogen Test

Urea is the principal nitrogen-containing compound in the blood and is the main end-product of protein metabolism in humans. It is produced in the liver through the urea cycle (also known as the ornithine cycle) and excreted mainly by the kidneys in urine. Measuring the concentration of urea in the blood, commonly referred to as the Blood Urea Nitrogen (BUN) test, is a fundamental clinical biochemistry practice used to assess kidney function, hydration status, protein metabolism, and liver health.

Biochemistry of Urea Formation

Urea is synthesized in the liver via the urea cycle, which detoxifies ammonia, a by-product of amino acid catabolism. This process involves several steps and enzymes, and requires energy in the form of ATP. Ammonia (NH₃) is toxic to cells and must be converted into a less toxic, water-soluble compound for safe excretion. Urea, being non-toxic and highly soluble, serves this purpose effectively. The cycle takes place partly in the mitochondrial matrix and partly in the cytosol of hepatocytes.

Normal Reference Range

• Blood Urea Nitrogen (BUN): 7–20 mg/dL (2.5–7.1 mmol/L)
• Blood urea (direct measurement): 20–40 mg/dL

Values may vary slightly depending on the laboratory method and the population studied. Factors such as diet, hydration, and age can influence normal ranges.

Clinical Significance of Urea Estimation

The estimation of urea in the blood has several important clinical uses:

1. Assessment of Kidney Function: Urea is primarily excreted by the kidneys, so elevated blood urea levels (uremia) often indicate impaired kidney function.
2. Evaluation of Protein Metabolism: High protein intake or increased protein catabolism increases urea production, while low protein intake or liver disease reduces urea production.
3. Hydration Status: Dehydration can lead to elevated blood urea levels due to reduced kidney perfusion, while overhydration can lower urea levels.
4. Monitoring of Chronic Kidney Disease (CKD): Urea estimation is used alongside creatinine measurements to monitor disease progression.

Causes of Elevated Urea (Uremia)

Increased blood urea levels may result from:

• Prerenal causes: Dehydration, shock, congestive heart failure, severe burns, gastrointestinal bleeding, high-protein diet.
• Renal causes: Acute or chronic renal failure, glomerulonephritis, polycystic kidney disease.
• Postrenal causes: Urinary tract obstruction (stones, tumors, prostatic hypertrophy).

In prerenal conditions, blood flow to the kidneys is reduced, limiting urea excretion. In renal causes, there is direct kidney damage affecting filtration. In postrenal causes, obstruction in the urinary tract prevents normal excretion.

Causes of Decreased Urea

Low blood urea levels may be due to:

• Severe liver disease (impaired urea cycle function)
• Low protein intake or malnutrition
• Overhydration or syndrome of inappropriate antidiuretic hormone secretion (SIADH)
• Pregnancy (due to increased plasma volume and protein synthesis)

Methods of Urea Estimation

Several laboratory methods are used to measure blood urea:

1. Diacetyl Monoxime Method: Urea reacts with diacetyl monoxime in the presence of acid and heat to form a colored complex, which is measured spectrophotometrically.
2. Enzymatic Urease Method: Urease catalyzes the hydrolysis of urea to ammonia and carbon dioxide; ammonia is then quantified using colorimetric methods.
3. Automated Analyzers: Modern clinical chemistry analyzers employ enzymatic methods with high precision and rapid turnaround time.

BUN/Creatinine Ratio

Often, urea estimation is interpreted alongside creatinine to determine the cause of abnormal kidney function. A high BUN/Creatinine ratio suggests prerenal causes (e.g., dehydration), while a normal or low ratio may indicate intrinsic renal disease.

Urea and Ocular Health

While urea estimation is not directly a diagnostic test for eye diseases, kidney dysfunction and systemic metabolic imbalance can have ocular implications, including:

• Hypertensive retinopathy due to chronic kidney disease
• Diabetic retinopathy when linked with diabetic nephropathy
• Uremic optic neuropathy in severe cases of uremia

These associations underline the importance of systemic evaluation in patients presenting with certain ocular symptoms.

Sample Collection and Handling

Blood samples for urea estimation are usually collected from a vein in the arm. Serum or plasma can be used for testing. Fasting is not strictly necessary, but consistency in pre-test conditions improves accuracy. Samples should be analyzed promptly or refrigerated to prevent degradation.

Factors Affecting Urea Levels

• Dietary protein intake: High protein increases urea production; low protein decreases it.
• Hydration status: Dehydration increases urea concentration; overhydration decreases it.
• Medications: Corticosteroids, antibiotics, and certain chemotherapy drugs may alter urea levels.
• Exercise: Intense exercise may transiently elevate urea due to muscle breakdown.

Interpretation in Clinical Context

Urea estimation should never be interpreted in isolation. A complete assessment includes:

• Serum creatinine levels
• Glomerular filtration rate (GFR)
• Urinalysis results
• Patient's hydration status and diet

The integration of all these factors enables accurate diagnosis and management of underlying conditions.

Conclusion

The estimation of urea in clinical biochemistry provides essential information about kidney function, protein metabolism, and systemic health. Its measurement is quick, cost-effective, and easily repeatable, making it a cornerstone in both emergency and routine laboratory testing. While not specific to a single disease, changes in urea levels serve as important indicators that guide further investigation and management. For optometrists and ophthalmologists, awareness of systemic parameters like urea is valuable, as systemic diseases affecting kidneys can also influence ocular health, emphasizing the need for holistic patient care.

Creatinine – Significance of Its Estimation

Introduction

Creatinine is a nitrogenous waste product that is formed as a result of normal muscle metabolism. It is produced from creatine, a compound involved in the storage and supply of energy in muscles. Creatinine is released into the bloodstream at a fairly constant rate, depending largely on muscle mass, and is eliminated from the body through the kidneys. Because of its stable production and almost complete removal by the kidneys, serum creatinine measurement is one of the most widely used indicators for evaluating kidney function.

Biochemistry of Creatinine

Creatinine is formed from creatine phosphate during muscle energy metabolism. Creatine itself is synthesized mainly in the liver and kidneys from amino acids—arginine, glycine, and methionine. Once produced, creatine is transported to muscle tissue, where it is phosphorylated to phosphocreatine by creatine kinase. Phosphocreatine serves as an energy reserve that can rapidly donate phosphate groups to ADP to generate ATP during muscle contraction.

Over time, phosphocreatine spontaneously and irreversibly breaks down into creatinine, which then diffuses into the bloodstream. Since this process is independent of diet, exercise, or most physiological variations, the daily creatinine production is relatively constant for a given individual.

Normal Creatinine Levels

• Men: 0.6 – 1.2 mg/dL (53 – 106 µmol/L)
• Women: 0.5 – 1.1 mg/dL (44 – 97 µmol/L)
• Children: 0.3 – 0.7 mg/dL (27 – 62 µmol/L)
• Newborns: 0.2 – 1.0 mg/dL (18 – 88 µmol/L)

The slightly higher values in men are due to greater average muscle mass compared to women.

Clinical Significance of Creatinine Estimation

Estimating creatinine levels plays an essential role in assessing renal function, particularly the glomerular filtration rate (GFR). Because creatinine is almost entirely excreted by glomerular filtration with minimal tubular reabsorption, its blood concentration is inversely related to GFR. When kidney function declines, GFR decreases, leading to increased blood creatinine levels.

1. Assessment of Kidney Function

Measuring serum creatinine helps detect acute kidney injury (AKI) and chronic kidney disease (CKD). In early stages of kidney impairment, creatinine levels may remain within normal limits, but progressive damage will eventually cause noticeable elevation.

2. Estimation of Glomerular Filtration Rate (GFR)

Formulas like the Cockcroft-Gault equation or the MDRD (Modification of Diet in Renal Disease) study equation use serum creatinine, age, sex, and body weight to estimate GFR. This estimation helps in:

• Staging CKD
• Adjusting drug dosages in patients with renal impairment
• Monitoring the progression of renal disease

3. Detection of Muscle Disorders

Creatinine levels can also reflect muscle mass and activity. Extremely low creatinine levels may be seen in conditions causing muscle wasting, while mild elevations can occur in muscular dystrophies or after intense exercise.

4. Evaluation in Critical Illness

In hospitalized or ICU patients, rising creatinine levels can indicate acute kidney injury due to sepsis, shock, dehydration, or exposure to nephrotoxic drugs.

Causes of Elevated Creatinine

• Acute kidney injury (due to dehydration, infection, toxins, ischemia)
• Chronic kidney disease (diabetes, hypertension, glomerulonephritis)
• Obstruction of urinary tract (stones, tumors, enlarged prostate)
• Increased muscle breakdown (rhabdomyolysis, trauma, burns)
• High protein intake or creatine supplementation (mild temporary rise)

Causes of Low Creatinine

• Reduced muscle mass (aging, malnutrition, prolonged illness)
• Pregnancy (due to increased plasma volume)
• Severe liver disease (reduced creatine synthesis)

Methods of Estimation

1. Jaffe’s Reaction

The most common method for measuring creatinine is the Jaffe reaction, in which creatinine reacts with picric acid in alkaline conditions to form an orange-red complex. Although simple, it is susceptible to interference by substances such as glucose, ketones, and proteins.

2. Enzymatic Methods

These methods involve the enzymatic conversion of creatinine to creatine, then to sarcosine and urea, with subsequent colorimetric or spectrophotometric detection. Enzymatic assays are more specific and less prone to interference compared to Jaffe’s method.

3. Point-of-Care Testing

Portable devices are now available for rapid creatinine estimation in emergency and outpatient settings.

Creatinine Clearance Test

This test measures the volume of blood plasma cleared of creatinine per unit time, providing a more accurate measure of GFR. It requires both a blood sample and a 24-hour urine collection. Creatinine clearance is calculated using:

Creatinine Clearance (mL/min) =
(Urine creatinine × Urine volume) / (Plasma creatinine × Time in minutes)

Role in Drug Dosage Adjustment

Many medications, particularly antibiotics and chemotherapeutic agents, are excreted by the kidneys. In patients with reduced renal function, drug clearance is impaired, leading to toxicity unless the dose is adjusted. Serum creatinine and estimated GFR are vital for determining safe drug dosing.

Limitations of Serum Creatinine

• Serum creatinine can remain normal until a significant portion of kidney function is lost (up to 50%).
• Variations in muscle mass, diet, and hydration can influence results.
• In elderly patients, low muscle mass can mask kidney dysfunction.

Therefore, serum creatinine is best interpreted alongside other tests such as estimated GFR, blood urea nitrogen (BUN), and urinalysis.

Recent Advances

Novel biomarkers like cystatin C are being studied as more sensitive indicators of early kidney damage, but serum creatinine remains the most widely used and cost-effective test globally.

Conclusion

Creatinine estimation is an indispensable part of clinical biochemistry and nephrology. It provides a simple yet powerful tool for assessing renal function, guiding treatment decisions, monitoring disease progression, and adjusting medication dosages. While not perfect, its widespread availability, low cost, and reliability make it a cornerstone in the evaluation of kidney health.

Bilirubin – Significance of Its Estimation

Introduction

Bilirubin is a yellow pigment formed during the normal breakdown of red blood cells. It is a key component of bile and is excreted from the body through the liver, eventually reaching the intestine. Bilirubin metabolism and its measurement are of great clinical significance because abnormal levels can indicate a range of disorders, particularly those involving the liver, gallbladder, and red blood cells. The estimation of bilirubin levels is a routine yet vital laboratory investigation in clinical biochemistry, as it aids in the diagnosis and monitoring of various hepatic and hematological diseases.

Chemistry and Types of Bilirubin

Bilirubin is a tetrapyrrolic bile pigment derived from the catabolism of heme, the iron-containing component of hemoglobin. It exists in two primary forms:

• Unconjugated (Indirect) Bilirubin: This form is lipid-soluble and not water-soluble. It circulates in the bloodstream bound to albumin and is produced by the breakdown of heme. Unconjugated bilirubin is transported to the liver for further processing.
• Conjugated (Direct) Bilirubin: In the liver, unconjugated bilirubin undergoes conjugation with glucuronic acid, catalyzed by the enzyme UDP-glucuronosyltransferase, forming bilirubin diglucuronide. This form is water-soluble and can be excreted in bile.

The sum of both forms is referred to as Total Bilirubin.

Bilirubin Metabolism

Schematic representation of Bilirubin Metabolism 

The process of bilirubin metabolism involves several steps:

1. Heme Degradation: Senescent red blood cells are phagocytosed by macrophages in the spleen, liver, and bone marrow. Hemoglobin is split into heme and globin; globin is degraded into amino acids, while heme is converted to biliverdin by heme oxygenase.
2. Conversion to Unconjugated Bilirubin: Biliverdin is rapidly reduced to unconjugated bilirubin by biliverdin reductase.
3. Transport to the Liver: Unconjugated bilirubin binds to albumin in plasma and is carried to the liver.
4. Conjugation: In hepatocytes, unconjugated bilirubin is conjugated with glucuronic acid to form water-soluble bilirubin diglucuronide.
5. Excretion into Bile: Conjugated bilirubin is secreted into bile and transported to the intestine.
6. Intestinal Metabolism: In the intestine, bacterial enzymes convert conjugated bilirubin into urobilinogen. Some urobilinogen is reabsorbed (enterohepatic circulation), while the rest is converted to stercobilin and excreted in feces, giving stool its characteristic brown color.

Physiological Role of Bilirubin

Although primarily a waste product, bilirubin also exhibits certain beneficial physiological properties, such as antioxidant activity. It can neutralize reactive oxygen species and protect cells from oxidative damage. However, excessive accumulation in the body is toxic, particularly to the brain in newborns (kernicterus).

Normal Bilirubin Levels

• Total bilirubin: 0.3 – 1.2 mg/dL
• Direct (conjugated) bilirubin: 0.1 – 0.3 mg/dL
• Indirect (unconjugated) bilirubin: Calculated as total minus direct bilirubin

Significance of Bilirubin Estimation

Measuring bilirubin levels in the blood is crucial in the diagnosis, differential diagnosis, and monitoring of a variety of diseases:

1. Jaundice Detection: Jaundice occurs when bilirubin levels exceed normal values, causing yellowing of the skin, sclera, and mucous membranes. It can be classified into:
   • Pre-hepatic (Hemolytic) Jaundice: Due to excessive breakdown of red blood cells, leading to elevated unconjugated bilirubin.
   • Hepatic Jaundice: Due to liver cell damage (e.g., hepatitis, cirrhosis) affecting both conjugation and excretion, resulting in elevated levels of both unconjugated and conjugated bilirubin.
   • Post-hepatic (Obstructive) Jaundice: Due to blockage in the bile ducts (e.g., gallstones, tumors) causing elevated conjugated bilirubin.
2. Assessment of Liver Function: Bilirubin estimation is part of the liver function test (LFT) panel, which helps evaluate the liver’s capacity to conjugate and excrete bilirubin.
3. Hemolytic Anemia Diagnosis: Elevated unconjugated bilirubin may indicate excessive RBC destruction.
4. Monitoring Newborn Health: Newborn jaundice is common due to immature liver conjugation systems. High levels require monitoring to prevent kernicterus.

Methods of Bilirubin Estimation

Several laboratory techniques are used to measure bilirubin levels:

• Van den Bergh Reaction: A classical method using diazotized sulfanilic acid to form azobilirubin, which can be measured spectrophotometrically. Direct bilirubin reacts rapidly; indirect bilirubin reacts only in the presence of an accelerator.
• Colorimetric Methods: Based on the intensity of color formed by bilirubin-dye complexes.
• High-Performance Liquid Chromatography (HPLC): Provides highly accurate separation and quantification of bilirubin fractions.
• Automated Analyzers: Common in modern laboratories for routine bilirubin testing.

Factors Affecting Bilirubin Levels

• Hemolysis: Increases unconjugated bilirubin due to rapid breakdown of RBCs.
• Liver Diseases: Impair conjugation or excretion, affecting both bilirubin forms.
• Biliary Obstruction: Increases conjugated bilirubin due to blocked excretion pathways.
• Genetic Disorders: Such as Gilbert’s syndrome, Crigler-Najjar syndrome, and Dubin-Johnson syndrome, which affect bilirubin metabolism.

Clinical Conditions Associated with Abnormal Bilirubin

1. Hemolytic Anemias: Sickle cell anemia, thalassemia, autoimmune hemolytic anemia.
2. Liver Diseases: Viral hepatitis, alcoholic liver disease, cirrhosis.
3. Obstructive Diseases: Gallstones, cholangiocarcinoma, pancreatic carcinoma.
4. Neonatal Jaundice: Physiological or pathological causes.

Management and Monitoring

Bilirubin estimation is used not only for diagnosis but also for monitoring treatment effectiveness in liver and hematological disorders. In newborns, phototherapy and exchange transfusion may be used to lower bilirubin levels. In adults, treatment focuses on addressing the underlying cause.

Conclusion

The estimation of bilirubin is an essential tool in clinical biochemistry, providing critical information about liver function, bile duct patency, and red blood cell turnover. Abnormal bilirubin levels are a hallmark of various diseases, making its measurement indispensable in diagnostic medicine. Timely detection and interpretation of bilirubin abnormalities can significantly improve patient outcomes by guiding early and appropriate therapeutic interventions.

For more units of Ocular Biochemistry click below on the text 👇 

👉 Unit 1

👉 Unit 2

👉 Unit 3

👉 Unit 4