• Metabolism - carbohydrates, lipids, proteins, nucleotides in animals
• Clinical biochemistry - serum enzymes, blood gases, electrolytes in veterinary diagnostics
• Nutrition & digestion - ruminant fermentation, monogastric vs. polygastric differences
• Hormones & signaling - species-specific endocrinology
• Molecular biology - DNA, RNA, genetics in animals
• Pathological biochemistry - ketosis in cattle, hepatic lipidosis in cats, azotemia, etc.
• Comparative biochemistry - how biochemical processes differ across species

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QUESTION 5 - Definitions (1×6 = 6 marks)

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i) Lipoproteins

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ii) pH

pH = -log₁₀[H⁺]

• pH < 7.0 = acidic; pH = 7.0 = neutral; pH > 7.0 = alkaline
• Normal blood pH in domestic animals:

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iii) pKa

pKa = -log₁₀Ka

pH = pKa + log₁₀([A⁻]/[HA])

• A low pKa = stronger acid (dissociates readily)
• A high pKa = weaker acid

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iv) Heteropolysaccharide

• Hyaluronic acid - alternating D-glucuronic acid and N-acetyl-D-glucosamine; found in synovial fluid and vitreous humor
• Heparin - alternating glucuronic acid and glucosamine (sulfated); anticoagulant in mast cells
• Chondroitin sulfate - galactosamine + glucuronic acid; cartilage matrix component
• Agar - D-galactose + 3,6-anhydro-L-galactose; from seaweed

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v) Mucopolysaccharide

• A hexuronic acid (glucuronic or iduronic acid)
• An N-acetylhexosamine (glucosamine or galactosamine), often sulfated

• Mucopolysaccharidoses (MPS) are lysosomal storage diseases in animals (cats, dogs) caused by deficiency of enzymes that degrade GAGs - e.g., MPS I (alpha-L-iduronidase deficiency) in cats and dogs.
• Mucopolysaccharides are deposited in Hurler syndrome (neutrophil Adler bodies).

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vi) Mutarotation

1. The α-D-glucopyranose ring opens to form the straight-chain aldehyde
2. Ring closure can form either α (OH below plane, [α]D = +112.2°) or β (OH above plane, [α]D = +18.7°)
3. At equilibrium: ~36% α-form + ~64% β-form → specific rotation = +52.5°

• Catalyzed by mutarotase (aldose 1-epimerase) in cells
• Once a glycosidic bond forms, mutarotation is prevented
• Enzymes are stereospecific - glucose oxidase acts only on β-D-glucose

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QUESTION 6 - Classification of Lipids Based on Structure (5 marks)

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A. Simple Lipids

1. Fats (Triacylglycerols / Triglycerides)

• Esters of glycerol with three fatty acids
• Most abundant storage lipid in animals
• Saturated (solid at room temperature) or unsaturated (oils, liquid at room temperature)
• Vet example: Fat depots in cattle (tallow), poultry (lard), fatty acids mobilized in ketosis as NEFA (non-esterified fatty acids)

2. Waxes

• Esters of long-chain fatty acids with long-chain monohydric alcohols (not glycerol)
• Examples: Beeswax (myricyl palmitate), lanolin (wool wax from sheep), sebum in skin
• Function: Waterproofing, protection
• Vet example: Wool wax (lanolin) from sheep fleece; cerumen (ear wax) in dogs

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B. Compound (Complex) Lipids

1. Phospholipids

• Contain glycerol + 2 fatty acids + phosphate + nitrogenous base
• Major component of all cell membranes (lipid bilayer)
• Types:
  • Phosphatidylcholine (Lecithin) - most abundant; forms bilayer of RBC membrane
  • Phosphatidylethanolamine (Cephalin) - important in brain and nervous tissue
  • Phosphatidylserine - involved in apoptosis
  • Phosphatidylinositol - second messenger (IP₃) signaling
  • Plasmalogens - important in heart and brain
• Vet example: Lecithin in egg yolk (hen); pulmonary surfactant (dipalmitoylphosphatidylcholine) in fetal lungs - important in neonatal foal respiratory distress

2. Glycolipids (Glycosphingolipids)

• Contain sphingosine + fatty acid + carbohydrate (no phosphate)
• Types:
  • Cerebrosides - sphingosine + fatty acid + single sugar (glucose or galactose)
  • Gangliosides - contain sialic acid (N-acetylneuraminic acid); abundant in brain neurons
  • Globosides - contain multiple sugars
• Function: Cell surface recognition, neural signal transduction
• Vet example: GM1 gangliosidosis is a lysosomal storage disease in dogs (Beagles, Portuguese Water Dogs), cats, and cattle

3. Sphingomyelin

• Sphingosine + fatty acid + phosphocholine
• Major myelin sheath component in neurons
• Vet example: Niemann-Pick disease (sphingomyelinase deficiency) reported in cats and dogs

4. Lipoproteins

• Lipids non-covalently complexed with proteins (apolipoproteins)
• Transport lipids in blood (see Q5.i above)

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C. Derived Lipids

1. Fatty Acids

• Long-chain monocarboxylic acids
• Saturated: Palmitic (C16:0), Stearic (C18:0) - found in tallow
• Unsaturated: Oleic (C18:1), Linoleic (C18:2, ω-6), Linolenic (C18:3, ω-3), Arachidonic (C20:4)
• Essential fatty acids (EFAs): Linoleic and α-linolenic acid - must be supplied in diet; cats additionally require arachidonic acid (cannot synthesize from linoleic acid due to low Δ6-desaturase activity)

2. Steroids (Sterols)

• Based on cyclopentanoperhydrophenanthrene nucleus (4 fused rings)
• Cholesterol - precursor for bile acids, steroid hormones, vitamin D
• Bile acids - cholate, deoxycholate; emulsify fats in digestion
• Steroid hormones - cortisol, aldosterone, testosterone, estrogen, progesterone
• Vitamin D - cholecalciferol; derived from 7-dehydrocholesterol under UV light
• Vet example: Vitamin D₃ toxicosis (calcinosis) in dogs from rodenticides; cholesterol metabolism critical in ruminants

3. Fat-Soluble Vitamins

• Vitamins A, D, E, K - stored in fat; absorbed with dietary fat
• Vet example: Vitamin K deficiency (warfarin toxicity in dogs) causes coagulopathy; Vitamin E/selenium deficiency causes white muscle disease in calves and lambs

4. Prostaglandins and Eicosanoids

• Derived from arachidonic acid (C20 polyunsaturated FA)
• Include prostaglandins, thromboxanes, leukotrienes
• Function: Inflammation, fever, platelet aggregation, reproductive physiology
• Vet example: NSAIDs (aspirin, meloxicam) in animals inhibit cyclooxygenase → reduce prostaglandin synthesis; PGF2α used for luteolysis in cattle/mares

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QUESTION 7 - Levels of Protein Structure (5 marks)

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1. Primary Structure

• The linear sequence of amino acids in the polypeptide chain
• Amino acids are linked by peptide bonds (covalent bonds between α-carboxyl and α-amino groups)
• Read from N-terminus → C-terminus
• This sequence is determined by the gene (DNA codons)
• Stabilized by: Covalent peptide bonds
• Significance: The primary structure determines all higher orders of structure. A single amino acid change can alter function (e.g., sickle cell-like disorders in animals)
• Vet example: Feline hemoglobin has a different primary structure than canine or human hemoglobin, giving it different oxygen affinity and susceptibility to Heinz body formation with oxidants

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2. Secondary Structure

• Local, regularly repeating spatial arrangement of the polypeptide backbone, stabilized by hydrogen bonds between backbone NH and C=O groups
• Main types:

α-Helix

• Right-handed coil; 3.6 amino acids per turn
• H-bonds between every 4th amino acid (residue n and n+4)
• Side chains project outward
• Found in: Keratin (hair, wool, hooves, horn in ruminants), myoglobin, hemoglobin

β-Pleated Sheet

• Extended, zigzag strands aligned parallel or antiparallel
• H-bonds between adjacent strands
• Found in: Silk fibroin (insect silk), immunoglobulins

β-Turns and Loops

• Allow polypeptide chain to reverse direction
• Common on protein surfaces
• Stabilized by: Hydrogen bonds
• Vet example: Keratin α-helix structure in wool (sheep) can be altered to β-sheet by stretching (wool's elastic property)

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3. Tertiary Structure

• The overall three-dimensional folding of a single polypeptide chain, including all secondary structure elements
• Determined by interactions among side chains (R-groups)

• Globular proteins (e.g., enzymes, antibodies) fold into compact shapes with a hydrophobic core and hydrophilic surface
• Vet example: Myoglobin tertiary structure in horse/dog muscle; immunoglobulin G tertiary structure in bovine colostrum

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4. Quaternary Structure

• Association of two or more polypeptide subunits (each with its own tertiary structure) into a functional multisubunit complex
• Subunits held together by the same non-covalent forces as tertiary structure (+ sometimes disulfide bonds)
• Examples:
  • Hemoglobin - tetramer (2α + 2β subunits); allows cooperative O₂ binding
  • Antibodies (IgG) - 4 chains (2 heavy + 2 light)
  • Collagen - triple helix of 3 α-chains

• Vet example: Bovine hemoglobin has different subunit composition and oxygen affinity than equine or canine hemoglobin. Fetal hemoglobin (HbF) in neonatal foals/calves has higher O₂ affinity than adult HbA.

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QUESTION 8 - Watson-Crick DNA Model + DNA vs RNA (5+4 = 9 marks)

A. Watson-Crick Double Helix Model of DNA (1953)

Key Features:

• Two polynucleotide strands wound around each other in a right-handed double helix
• The two strands run antiparallel (one 5'→3', the other 3'→5')
• The exterior (backbone) consists of deoxyribose sugar + phosphate groups
• The interior consists of nitrogenous bases

• Purines pair with pyrimidines:
  • Adenine (A) ↔ Thymine (T) - joined by 2 hydrogen bonds
  • Guanine (G) ↔ Cytosine (C) - joined by 3 hydrogen bonds
• This is complementary, antiparallel base pairing
• G-C rich DNA is more stable (higher melting temperature, Tm)

• Diameter: 2 nm (20 Å)
• Pitch (length per turn): 3.4 nm
• Base pairs per turn: 10 bp
• Rise per base pair: 0.34 nm
• The planes of bases are perpendicular to the helical axis

• The double helix has an asymmetric wrapping creating a major groove (wider) and a minor groove (narrower)
• Regulatory proteins and transcription factors bind in the major groove

• Hydrogen bonds between complementary base pairs (hold strands together)
• Hydrophobic stacking interactions between bases (major stabilizing force)
• Phosphodiester backbone (covalent, connects nucleotides within each strand)

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B. Differences Between DNA and RNA

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QUESTION 9 - Types of Membrane Transport (5 marks)

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1. Passive Transport (No energy required - moves down concentration/electrochemical gradient)

A. Simple Diffusion

• Movement of small, nonpolar, lipid-soluble molecules directly through the lipid bilayer
• Driven by concentration gradient (high → low)
• Rate depends on: Lipid solubility, molecular size, concentration gradient, surface area
• Examples: O₂, CO₂, N₂, urea, ethanol, steroid hormones, fatty acids
• Vet example: O₂/CO₂ exchange across alveolar membranes in horse/dog lungs; CO₂ diffusion across rumen epithelium

B. Facilitated Diffusion

• Movement of polar or charged molecules down concentration gradient, aided by specific membrane transport proteins (carriers or channels)
• No energy required; saturatable (has Km and Vmax like enzymes)
• Types of transport proteins:
  • Channel proteins - form aqueous pores (e.g., aquaporins for water, ion channels for Na⁺, K⁺, Ca²⁺, Cl⁻)
  • Carrier proteins - bind solute, undergo conformational change (e.g., GLUT transporters for glucose)
• Examples: Glucose via GLUT1-4, amino acids, ions
• Vet example: GLUT2 in ruminant mammary gland for milk lactose synthesis; aquaporin channels in renal tubules of dogs/cats for water reabsorption

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2. Active Transport (Energy required - can move against gradient)

A. Primary Active Transport

• Directly uses ATP hydrolysis to move substances against concentration or electrochemical gradient
• Transport protein acts as an ATPase pump
• Examples:
  • Na⁺/K⁺-ATPase (Sodium pump) - pumps 3 Na⁺ out, 2 K⁺ in; maintains resting membrane potential in all animal cells
  • Ca²⁺-ATPase - pumps Ca²⁺ out of cytoplasm (SERCA in SR of muscle)
  • H⁺/K⁺-ATPase - pumps H⁺ into stomach (parietal cells); target of proton pump inhibitors (omeprazole in dogs/horses)
• Vet example: Na⁺/K⁺-ATPase inhibition by digitalis glycosides improves cardiac contractility in dogs with heart failure; impaired Ca²⁺-ATPase in muscle causes exertional rhabdomyolysis in horses

B. Secondary Active Transport (Co-transport)

• Uses the electrochemical gradient created by primary active transport (usually Na⁺ gradient) as energy source - no direct ATP use
• Symport - substrate moves in same direction as Na⁺ (e.g., Na⁺-glucose cotransporter SGLT1/2 in intestine and kidney)
• Antiport - substrate moves opposite to Na⁺ (e.g., Na⁺/H⁺ exchanger, Na⁺/Ca²⁺ exchanger)
• Vet example: SGLT2 inhibitors (e.g., velagliflozin in horses) used for equine metabolic syndrome/laminitis; SGLT1 in bovine small intestine for glucose absorption

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3. Vesicular Transport (Bulk Transport)

A. Endocytosis (into cell)

• Phagocytosis - engulfment of large particles (bacteria, debris) by pseudopods; performed by neutrophils and macrophages
• Pinocytosis - engulfment of extracellular fluid/small solutes in small vesicles
• Receptor-mediated endocytosis - ligand binds specific receptor → clathrin-coated pit → endosome; e.g., LDL uptake via LDL receptor; colostral IgG absorption in neonatal calves/foals via Fc receptor

B. Exocytosis (out of cell)

• Vesicles fuse with plasma membrane and release contents
• Constitutive - continuous secretion (e.g., mucus secretion)
• Regulated - triggered by signal (e.g., insulin release from pancreatic β-cells in response to glucose; neurotransmitter release at neuromuscular junction)
• Vet example: Colostrum IgG uptake by neonatal calves through pinocytosis/receptor-mediated endocytosis is time-limited (gut closure at 24-36 hours) - critical for passive immunity

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Summary Diagram

MEMBRANE TRANSPORT
├── Passive (no ATP)
│   ├── Simple Diffusion (O₂, CO₂, lipids)
│   └── Facilitated Diffusion (glucose via GLUT, ions via channels)
├── Active (uses energy)
│   ├── Primary Active (Na⁺/K⁺-ATPase, Ca²⁺-ATPase)
│   └── Secondary Active (SGLT1/2 symport, Na⁺/H⁺ antiport)
└── Vesicular Transport
    ├── Endocytosis (phagocytosis, pinocytosis, receptor-mediated)
    └── Exocytosis (insulin release, neurotransmitter release)

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Sources: Basic Medical Biochemistry (Lieberman & Peet, 6e); Biochemistry, Lippincott Illustrated Reviews (8e); Harper's Illustrated Biochemistry (32e); Tietz Textbook of Laboratory Medicine (7e). All applied to veterinary species context.

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QUESTION 14 - Definitions (1×5 = 5 marks)

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i) Glycosuria

1. Blood glucose exceeds the renal threshold (hyperglycemic glycosuria), OR
2. Renal tubular reabsorption is impaired (normoglycemic/renal glycosuria)

• Dogs: ~180-220 mg/dL
• Cats: ~280-300 mg/dL (higher threshold - glycosuria less common in stressed cats)
• Cattle: ~100 mg/dL (lower threshold - glycosuria in ketosis rare)

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ii) Proteinuria

• Urine Protein:Creatinine ratio (UPC): Key test in dogs and cats
  • Normal: UPC <0.2 in dogs; <0.4 in cats
  • Borderline: 0.2-0.5 (dogs), 0.4-0.5 (cats)
  • Proteinuric: >0.5 (both species)
• Greyhounds and Bernese Mountain Dogs are predisposed to glomerulonephropathy
• Persistent proteinuria in dogs/cats is a major risk factor for progressive chronic kidney disease (CKD)
• Amyloidosis in Shar-Peis and Abyssinian cats causes severe glomerular proteinuria

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iii) Pregnancy Toxaemia

• In late pregnancy, energy demands of rapidly growing fetus/fetuses exceed dietary energy intake
• The body mobilizes NEFA (non-esterified fatty acids) from fat depots
• Liver is overwhelmed with NEFA → excess acetyl-CoA → ketone body production (acetoacetate, β-hydroxybutyrate, acetone)
• Glucose deficiency compounds neurological signs

• Multiple fetuses (twins/triplets in ewes and does)
• Sudden change in feed or underfeeding in late gestation
• Obesity before pregnancy
• Concurrent disease/stress

• Blood glucose: <40 mg/dL (hypoglycemia)
• Blood ketones: >3 mmol/L (hyperketonemia)
• Blood β-hydroxybutyrate (BHBA): >3 mmol/L
• Ketonuria (positive urine dipstick for ketones)
• Elevated NEFA
• Elevated liver enzymes (hepatic lipidosis)

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iv) Xenobiotics

• Air, water, food, soil
• Absorbed via inhalation, ingestion, or skin contact

• Most xenobiotics are lipophilic - absorbed through membranes, transported by lipoproteins
• Metabolized primarily in the liver by cytochrome P-450 enzymes
• Species differences in P-450 enzymes affect drug metabolism:
  • Cats lack efficient glucuronyl transferase → aspirin, acetaminophen, phenol compounds are toxic
  • Dogs are slow acetylators → sulfonamide toxicity
  • Ruminants have extensive rumen microbial biotransformation before systemic absorption

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v) Acute Phase Proteins (APPs)

• SAA and CRP are rapid, sensitive markers of inflammation/infection in dogs and cats
• Haptoglobin in cattle: marker of mastitis, pneumonia, metritis
• APPs are used to monitor response to treatment, predict prognosis in sepsis, FIP in cats
• Rise within 6 hours of stimulus; more sensitive than WBC count for early inflammation

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QUESTION 15 - Liver and Kidney Function Tests (5+5 = 10 marks)

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A. LIVER FUNCTION TESTS (LFTs)

1. Tests of Hepatocellular Integrity (Enzyme Leakage Tests)

a) Serum Aminotransferases (Transaminases)

• Normal range (dog): 10-100 U/L; (cat): 10-100 U/L
• Liver-specific in dogs and cats (cytosolic enzyme of hepatocytes)
• Elevated in: hepatocellular necrosis, hepatitis, hepatic lipidosis, copper storage disease
• Not liver-specific in horses and cattle (also in muscle)
• Considered MORE specific for liver disease than AST

• Present in liver, heart, skeletal muscle, RBCs
• Less liver-specific than ALT
• Useful in horses and cattle where ALT is not hepatic-specific
• Normal range (horse): 100-350 U/L
• AST:ALT ratio >2 suggests alcoholic hepatitis in humans; in animals, suggests muscle damage component

b) Alkaline Phosphatase (ALP / SAP)

• Membrane-bound enzyme in hepatocyte canalicular membranes, bone, kidney, intestine
• Elevated in cholestasis (obstructive jaundice) and bone disease
• Dog has multiple isoenzymes including a steroid-induced ALP (SALP) - elevated with glucocorticoids (Cushing's syndrome, steroid administration)
• Cat - ALP elevation is very significant even when mildly elevated (short half-life ~6 hours)
• Foals and calves - physiologically high ALP due to active bone growth
• Very high in biliary obstruction, cholangiohepatitis

c) Gamma-Glutamyl Transferase (GGT / GGT)

• Membrane enzyme; useful in cattle, horses (more sensitive than ALP for hepatic disease)
• Elevated in: biliary obstruction, hepatocellular disease
• Neonatal ruminants: Elevated GGT from colostrum absorption (first 24-48 hrs) - NOT a sign of liver disease
• Equine: Valuable for detecting hepatic lipidosis, pyrrolizidine alkaloid toxicity

d) Lactate Dehydrogenase (LDH)

• Widely distributed - liver, heart, muscle, RBC
• Isoenzymes help localize the organ
• Less specific; used in combination with other tests

e) Sorbitol Dehydrogenase (SDH)

• Highly liver-specific in cattle, sheep, and horses (unlike ALT)
• Short half-life - excellent marker of acute hepatocellular damage
• Valuable in ruminant liver disease diagnosis

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2. Tests of Hepatic Synthetic Function

a) Serum Albumin

• Synthesized exclusively by liver
• Half-life ~7-10 days (dog), ~18-19 days (cattle)
• Low albumin (hypoalbuminemia): chronic liver failure, cirrhosis, protein-losing nephropathy, PLE
• Normal: dogs 2.5-4.0 g/dL; cattle 3.0-3.8 g/dL

b) Serum Total Protein and Globulins

• Globulins increase in chronic inflammation, liver disease (reverse albumin:globulin ratio)
• A:G ratio <1.0 suggests chronic disease

c) Prothrombin Time (PT) and Clotting Factors

• Liver synthesizes factors I, II, V, VII, VIII, IX, X, XI, XII
• PT prolonged in severe acute liver disease (cannot wait for albumin to drop)
• Vitamin K deficiency (rodenticide toxicity in dogs) also prolongs PT - respond to Vit K injection

d) Blood Urea Nitrogen (BUN) / Urea

• Liver converts ammonia to urea via urea cycle
• Low BUN in liver failure = impaired urea synthesis
• Portosystemic shunts (dogs, cats) → ammonia not converted → hepatic encephalopathy + low BUN

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3. Tests of Bilirubin Metabolism

a) Serum Total Bilirubin

• Normal: dogs <0.3 mg/dL; cats <0.2 mg/dL; horses <2-3 mg/dL
• Elevated (icterus/jaundice) when >2 mg/dL (dogs/cats)

b) Fractionation: Conjugated vs. Unconjugated Bilirubin

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4. Tests of Excretory Function

a) Serum Bile Acids (SBA)

• Most sensitive indicator of hepatic function and portal circulation in small animals
• Done as pre- and 2-hour post-prandial bile acids (fasting + fed sample)
• Normal: preprandial <10 µmol/L; postprandial <20 µmol/L (dogs)
• Elevated in: portosystemic shunts (PSS), liver failure, hepatitis
• Test of choice for detecting PSS in dogs and cats

b) Blood Ammonia

• Elevated (hyperammonemia) in: portosystemic shunts, hepatic failure, urea cycle defects
• Causes hepatic encephalopathy - neurological signs in dogs/cats with PSS

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B. KIDNEY FUNCTION TESTS (RENAL FUNCTION TESTS)

1. Glomerular Function Tests

a) Blood Urea Nitrogen (BUN) / Blood Urea

• Urea is the end product of protein catabolism; excreted by kidneys
• Normal: dogs 10-28 mg/dL; cats 20-40 mg/dL; cattle 10-25 mg/dL

b) Serum Creatinine

• More reliable than BUN - less affected by diet, muscle breakdown rate is constant
• Freely filtered at glomerulus, minimally secreted by tubules
• Reflects GFR (inversely related)
• Normal: dogs 0.5-1.5 mg/dL; cats 0.8-1.8 mg/dL; horses 1.2-1.9 mg/dL
• IRIS staging of CKD in dogs/cats based on creatinine
• Limitation: Creatinine rises only after >75% nephron loss (not sensitive for early CKD)

c) Symmetric Dimethylarginine (SDMA)

• Newer biomarker of GFR in dogs and cats
• Detects CKD when 40% of nephrons are lost (earlier than creatinine)
• Not affected by muscle mass (unlike creatinine)
• Normal: dogs <14 µg/dL; cats <14 µg/dL
• Used in IRIS CKD staging

d) Creatinine Clearance / GFR Measurement

• GFR = (Urine creatinine × Urine volume) / (Plasma creatinine × Time)
• Gold standard: inulin clearance (exogenous compound, freely filtered, not reabsorbed/secreted)
• Practical alternative: iohexol clearance (used in dogs/cats)

e) BUN:Creatinine Ratio

• Normal: 15-25:1
• >25:1 suggests prerenal cause (dehydration) or GI bleeding
• <15:1 suggests hepatic failure (reduced urea synthesis) or low protein diet

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2. Tubular Function Tests

a) Urinalysis - Urine Specific Gravity (USG)

• Measures concentrating ability of renal tubules
• Normal USG for dogs: 1.020-1.060; cats: 1.035-1.060
• Isosthenuria (USG 1.008-1.012): Fixed dilute urine = severe tubular damage
• Hyposthenuria (USG <1.008): Dilute urine = intact tubular function (DI, psychogenic polydipsia)

b) Urine Protein:Creatinine Ratio (UPC)

• Assesses glomerular integrity and tubular protein handling
• Normal: <0.2 (dogs), <0.4 (cats)

c) Fractional Excretion of Electrolytes

• FE(Na) = (Urine Na × Plasma Cr) / (Plasma Na × Urine Cr) × 100
• Used to differentiate prerenal from renal causes of oliguria

d) Urine Gamma-Glutamyl Transferase (GGT)

• Marker of tubular damage
• Urine GGT:Creatinine ratio used in horses and cattle

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Summary Table: Key Veterinary Reference Values

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QUESTION 16 - Metabolic Acidosis and Alkalosis with Compensatory Mechanisms (6 marks)

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ACID-BASE FUNDAMENTALS

pH = 6.1 + log₁₀([HCO₃⁻] / (0.03 × PaCO₂))

• HCO₃⁻: 18-24 mEq/L (dogs), 17-22 mEq/L (cats)
• PaCO₂: 35-45 mmHg
• pH: 7.35-7.45

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A. METABOLIC ACIDOSIS

Causes in Animals:

• Ketoacidosis: DM in dogs/cats; pregnancy toxaemia in ewes/does; bovine ketosis
• Lactic acidosis: Sepsis, shock, grain overload in ruminants (D-lactic acidosis), exhaustion
• Uremic acidosis: CKD - failure to excrete H⁺ and regenerate HCO₃⁻
• Toxins: Ethylene glycol (antifreeze) poisoning in dogs/cats; salicylate toxicity
• Rumen acidosis: Excess lactic acid production from Lactobacillus spp. in grain overload

• Severe diarrhea (loss of HCO₃⁻ - esp. calves/foals)
• Renal tubular acidosis (RTA)
• Urethral obstruction (cats - uroabdomen)

Biochemical Changes:

Compensatory Mechanism for Metabolic Acidosis:

• Decreased pH stimulates peripheral and central chemoreceptors
• Increases respiratory rate and depth (hyperventilation)
• CO₂ is blown off → PaCO₂ falls
• This raises pH back toward normal (never fully corrects)
• Expected PaCO₂ compensation: PaCO₂ = (1.5 × HCO₃⁻) + 8 ± 2 (Winter's formula - dogs)
• Also: PaCO₂ ≈ HCO₃⁻ + 15

• Kidneys increase H⁺ secretion (into tubular lumen)
• Increase NH₄⁺ excretion (ammonia buffer system)
• Increase HCO₃⁻ reabsorption and regeneration
• (Note: If kidneys are the cause, this compensation is absent/impaired)

• Bicarbonate buffer: H⁺ + HCO₃⁻ → H₂CO₃ → H₂O + CO₂
• Intracellular buffering: H⁺ moves into cells in exchange for K⁺ (→ hyperkalemia)
• Hemoglobin buffer: Hb- + H⁺ → HHb
• Bone carbonate: Mobilized in chronic acidosis

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B. METABOLIC ALKALOSIS

Causes in Animals:

• Abomasal displacement/volvulus (Left DA, Right DA) in cattle: Most important cause in veterinary medicine
  • Abomasal sequestration of HCl → hypochloremic, hypokalemic metabolic alkalosis
  • Classic: pH ↑, HCO₃⁻ ↑, Cl⁻ ↓, K⁺ ↓
• Vomiting (loss of HCl): Dogs/cats with pyloric obstruction, gastric dilation-volvulus (GDV)
• Nasogastric suctioning

• Excessive NaHCO₃ administration (iatrogenic)
• Sodium bicarbonate overdose in treating acidosis

• Diuretic therapy (furosemide) → loss of Cl⁻ and K⁺-rich fluid → HCO₃⁻ relatively concentrated

Biochemical Changes:

Compensatory Mechanism for Metabolic Alkalosis:

• Elevated pH inhibits respiratory chemoreceptors
• Causes hypoventilation → CO₂ retention → PaCO₂ rises
• This lowers pH back toward normal
• Expected compensation: PaCO₂ = (0.9 × HCO₃⁻) + 9 ± 2
• Limitation: Hypoxia (from hypoventilation) eventually overrides the respiratory inhibition

• Normally kidneys can very efficiently excrete HCO₃⁻
• However, in hypochloremic metabolic alkalosis (as in abomasal volvulus), renal compensation is impaired because:
  • Cl⁻ depletion drives HCO₃⁻ reabsorption (to maintain electroneutrality)
  • Hypovolemia stimulates aldosterone → Na⁺ retention + K⁺/H⁺ loss → paradoxical aciduria
• Treatment: IV isotonic NaCl (to provide Cl⁻) + KCl supplementation

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Summary Comparison Table

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QUESTION 17 - Diabetes Mellitus and Its Diagnosis (5+1 = 6 marks)

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A. Diabetes Mellitus

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Types of Diabetes Mellitus

1. Type 1 DM (Insulin-Dependent DM)

• Caused by immune-mediated or idiopathic destruction of pancreatic β-cells
• Results in absolute insulin deficiency
• Prone to diabetic ketoacidosis (DKA)
• Requires exogenous insulin to sustain life
• In animals: Most common form in dogs (similar to human T1DM)
  • Immune-mediated insulitis, chronic pancreatitis, or idiopathic β-cell loss
  • Also occurs in horses, cattle, cats (less common)

2. Type 2 DM (Non-Insulin-Dependent / Insulin-Resistant DM)

• Characterized by insulin resistance in peripheral tissues + relative β-cell insufficiency
• Patients may have normal, low, or elevated insulin levels
• Usually non-ketotic (unless stressed)
• Associated with obesity
• In animals: Most common form in cats (85-95% of feline DM)
  • Amyloid deposition in islets (IAPP - islet amyloid polypeptide) contributes to β-cell loss
  • Diet change (low-carb) and weight loss can lead to diabetic remission in cats
  • Also common in horses (Equine Metabolic Syndrome / PPID)

3. Secondary DM

• DM secondary to another condition:
  • Diestrus-associated DM in intact female dogs - progesterone stimulates GH → insulin resistance
  • Hyperadrenocorticism (Cushing's) - cortisol antagonizes insulin
  • Acromegaly (cats) - excess GH → insulin resistance; most severe insulin-resistant DM in cats
  • Chronic pancreatitis - destruction of islets (common cause in dogs)
  • Exocrine pancreatic insufficiency (dogs)
  • Glucagonoma - rare
  • Drugs: corticosteroids, megestrol acetate (cats), thiazide diuretics

4. Gestational DM

• Occurs during pregnancy (progesterone, placental lactogen → insulin resistance)
• Can occur in bitches during diestrus (equivalent of luteal phase)

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Pathophysiology and Metabolic Consequences

• Glucose cannot enter cells (muscle, fat, liver) → hyperglycemia
• Fat mobilization → elevated NEFA → ketogenesis in liver → ketones (acetoacetate, β-OHB, acetone)
• Protein catabolism → gluconeogenesis → worsens hyperglycemia
• Osmotic diuresis (glucose exceeds renal threshold) → polyuria → polydipsia
• Classic signs: PU/PD, polyphagia, weight loss, cataracts (dogs)

• Life-threatening complication - mostly dogs and cats
• Severe insulin deficiency + stress hormones (glucagon, cortisol, epinephrine)
• Massive ketone production → anion gap metabolic acidosis
• Signs: vomiting, anorexia, weakness, dehydration, fruity breath (acetone), coma

• Hyperglycemia → sorbitol accumulation in lens (aldose reductase pathway) → osmotic damage → cataracts
• Occurs in ~70% of diabetic dogs within 5-6 months of diagnosis

• Severe insulin deficiency → massive fat mobilization → hepatic fat accumulation
• Diabetic cats very prone to hepatic lipidosis

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B. Diagnosis of Diabetes Mellitus

In Dogs and Cats:

• Stress hyperglycemia is acute → fructosamine remains normal
• True DM → fructosamine elevated (reflects weeks of chronic hyperglycemia)

Oral Glucose Tolerance Test (OGTT):

• 1 g/kg glucose orally → measure blood glucose at 0, 30, 60, 120 minutes
• Diabetic: glucose >200 mg/dL at 2 hours; fails to return to baseline
• Used in prediabetes detection and horses with metabolic syndrome

Urine Ketones:

• Positive ketones = DKA (more severe/acute form)
• Dogs with DKA require hospitalization with insulin + fluid therapy

Additional tests in animals:

• Abdominal ultrasound: Pancreatic changes (pancreatitis), adrenal size (Cushing's)
• ACTH stimulation test / LDDST: Rule out Cushing's in dogs
• Serum progesterone: Rule out diestrus-associated DM in intact females
• IGF-1 levels: Screen for acromegaly in cats (caused by pituitary tumor)

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QUESTION 18 - Detoxification Mechanism of Xenobiotics (3 marks)

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Phase I Reactions (Functionalization)

• A large family of heme-containing mixed-function oxidases
• CYP3A4 - most important (metabolizes ~50% of drugs)
• Located in ER of hepatocytes; also in intestinal mucosa, lungs, skin
• Reaction: Drug + O₂ + NADPH → Oxidized drug + H₂O + NADP⁺

• Carbon tetrachloride → CCl₃· (hepatotoxic free radical) via CYP2E1
• Acetaminophen → NAPQI (reactive quinone) → hepatotoxicity (especially cats - lack GSH)
• Aflatoxin B₁ → epoxide metabolite → binds DNA → hepatocarcinogen in cattle, dogs

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Phase II Reactions (Conjugation/Biosynthetic)

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Phase III - Excretion

• Excreted in urine (small conjugates, <500 Da)
• Excreted in bile (larger conjugates, >500 Da) → enterohepatic circulation possible

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Species Differences in Xenobiotic Detoxification (Critical for Veterinary Practice)

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Diagram of Xenobiotic Biotransformation

LIPOPHILIC XENOBIOTIC (absorbed)
         ↓
[PHASE I - CYP450]
Oxidation / Reduction / Hydrolysis
         ↓
Reactive Intermediate (±toxic)
         ↓
[PHASE II - Conjugation]
Glucuronide / Sulfate / Glutathione / Acetyl / Methyl conjugate
         ↓
WATER-SOLUBLE, NON-TOXIC CONJUGATE
         ↓
    Urine / Bile → Feces

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Sources: Harper's Illustrated Biochemistry (32e); Robbins & Kumar Pathologic Basis of Disease (10e); Mulholland & Greenfield's Surgery (7e); Tietz Textbook of Laboratory Medicine (7e); Rosen's Emergency Medicine (10e); Katzung's Basic & Clinical Pharmacology (16e). All content framed in veterinary species context.

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QUESTION 5 - Definitions (1×11 = 11 marks)

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a) Mutarotation

• At equilibrium in glucose: ~36% α + ~64% β, with specific rotation = +52.5°
• Catalyzed by mutarotase in cells
• Vet relevance: Glucose oxidase-based analyzers measure only β-glucose; mutarotase is added to commercial kits for accurate results in veterinary diagnostics

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b) Glycosidic Bond

• Ruminants can digest cellulose (β-1,4 glycosidic bonds) via rumen microbial cellulases - monogastrics cannot
• Lactase deficiency in neonatal animals = inability to hydrolyze β(1→4) bond of lactose → lactose intolerance, diarrhea
• Glycogen phosphorylase cleaves α-1,4 bonds in glycogen for energy mobilization in animals

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c) Lipoproteins

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d) Albumin-Globulin (A:G) Ratio

A:G ratio = Albumin (g/dL) / Globulin (g/dL)

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e) Hyperinsulinism

• Blood glucose <60 mg/dL (dogs) → neurological signs
• Neuroglucopenia → weakness, seizures, collapse
• Counter-regulatory hormones activated: glucagon, epinephrine, cortisol, GH

1. Clinical signs of hypoglycemia
2. Blood glucose <60 mg/dL during episode
3. Relief of signs with glucose administration

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f) Dysproteinemia

• Hypoalbuminemia: liver failure (reduced synthesis), PLN, PLE, malnutrition
• Hypoglobulinemia: failure of passive transfer (neonates), immunodeficiency
• Vet example: Failure of passive transfer (FPT) in foals/calves = insufficient colostral IgG absorption → low IgG → immunodeficiency → septicemia

• Dehydration (relative increase)
• Hypergammaglobulinemia: chronic infection, FIP, multiple myeloma, ehrlichiosis in dogs
• Monoclonal gammopathy: single clone of plasma cells producing one immunoglobulin → M-spike on electrophoresis = multiple myeloma, lymphoma, ehrlichiosis

• Abnormal immunoglobulin produced by malignant plasma cells
• Vet example: Bence-Jones proteins (free light chains) in urine of dogs with multiple myeloma → damage renal tubules → cast nephropathy

• Serum protein electrophoresis (SPE) separates proteins into: Albumin, α1, α2, β, γ-globulins
• Polyclonal gammopathy: broad γ-peak = chronic infection, ehrlichiosis, FIP
• Monoclonal gammopathy: sharp M-spike = myeloma

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g) Emulsification

• Bile salts (e.g., sodium taurocholate, sodium glycocholate) are amphipathic - have both hydrophilic and hydrophobic regions
• They coat fat globules → reduce surface tension → break large globules into tiny droplets (~1 µm)
• Creates a large surface area for pancreatic lipase to act upon
• Also form micelles with fatty acids and monoglycerides for absorption

• Cats: taurine conjugation only (unlike dogs that use both glycine and taurine) → taurine deficiency impairs bile salt synthesis + causes dilated cardiomyopathy
• Bile duct obstruction (pancreatitis in dogs, cholangiohepatitis in cats) → failure of emulsification → steatorrhea (fat malabsorption, greasy stool)
• Ruminants: bile delivered continuously (no gallbladder) vs. dogs/cats/horses (have gallbladder); pigs/horses also lack bile storage in some circumstances

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h) Peptide Bond

-CO-NH- (peptide linkage)

• Has partial double-bond character (resonance) → rigid, planar structure
• Trans configuration is preferred (side chains on opposite sides of the plane)
• NOT freely rotatable (unlike the Cα-N and Cα-CO bonds which can rotate, defining backbone conformation)
• Hydrolyzed by: proteases (trypsin, chymotrypsin, pepsin), strong acid/base, heat

• 2 amino acids = dipeptide
• 3-10 amino acids = oligopeptide

• 10 amino acids = polypeptide

• 100 amino acids (generally) = protein

• Trypsin and chymotrypsin (pancreatic proteases) hydrolyze peptide bonds during digestion in all animals
• Exocrine Pancreatic Insufficiency (EPI) in German Shepherds → insufficient protease → undigested protein in feces (polyphagia + weight loss)
• Proline and hydroxyproline in collagen cause unique kinking of the polypeptide - critical for tendon and ligament strength in horses

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i) pKa Value

• In amino acids: each ionizable group has its own pKa (pKa1 for α-COOH, pKa2 for α-NH₃⁺, pKaR for side chain)
• At the isoelectric point (pI) = (pKa1 + pKa2)/2 → net charge on amino acid = 0

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j) Oxidative Stress

• Mitochondrial electron transport chain (normal metabolism)
• Inflammatory cells (NADPH oxidase in neutrophils - respiratory burst)
• CYP450 reactions (xenobiotic metabolism)
• Ischemia-reperfusion injury

• Lipid peroxidation of cell membranes
• Protein oxidation and carbonylation
• DNA strand breaks, base oxidation (8-OHdG)

• Vitamin E/Selenium deficiency in ruminants → white muscle disease (nutritional myodegeneration) - oxidative damage to cardiac and skeletal muscle
• Heinz body hemolytic anemia in cats (onion, garlic, acetaminophen) → oxidation of hemoglobin
• Equine rhabdomyolysis - oxidative damage to muscle
• Aflatoxin → oxidative liver damage in poultry and cattle
• Selenium supplementation in livestock (selenium-deficient soils in many regions) prevents white muscle disease and retained placenta in cattle

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k) Pregnancy Toxaemia

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QUESTION 6 - Classification of Amino Acids Based on R-Group Solubility in Water (5 marks)

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CLASS 1: Nonpolar, Hydrophobic Amino Acids (R groups are insoluble in water)

• These R groups are insoluble in water
• Cluster in hydrophobic core of globular proteins - major force of protein folding
• BCAAs (Val, Leu, Ile): primary fuel for skeletal muscle during exercise and starvation
• Vet relevance: BCAA-enriched diets used in horses for muscle maintenance; proline + hydroxyproline constitute 1/3 of amino acids in collagen (tendons, bones, ligaments)

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CLASS 2: Polar, Uncharged (Neutral) Amino Acids (R groups are soluble but neutral at physiological pH)

• R groups are soluble in water (form H-bonds with water)
• Located on surface of proteins facing aqueous environment
• Serine and threonine: key sites of phosphorylation in signal transduction cascades
• Cysteine: forms disulfide bonds that stabilize extracellular proteins (immunoglobulins, keratin)
• Vet relevance:
  • Cystine urolithiasis in dogs (Newfoundlands, English Bulldogs) - defect in cystine reabsorption → cystine crystals/stones in urine
  • Tyrosine deficiency → reduced melanin → coat color changes; precursor of thyroxine (critical for metabolic rate in all animals)
  • Glutamine is the most abundant amino acid in blood; critical fuel for rapidly dividing cells (enterocytes, lymphocytes) → glutamine supplementation in critically ill veterinary patients

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CLASS 3: Charged Polar Amino Acids - Acidic (Negatively Charged at pH 7.4)

• Highly soluble in water (charged, interact strongly with water)
• Located on protein surfaces, form ionic bonds (salt bridges) with basic amino acids
• Aspartate and Glutamate are central to transamination reactions (ALT, AST enzymes)
• Glutamate is the major excitatory neurotransmitter in the CNS
• Vet relevance:
  • AST (Aspartate aminotransferase) is the primary liver enzyme used in cattle and horses for liver disease diagnosis
  • Glutamate participates in urea cycle (glutamate → α-ketoglutarate → enters TCA cycle)
  • Glutamate + ammonia → Glutamine (ammonia detoxification)

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CLASS 4: Charged Polar Amino Acids - Basic (Positively Charged at pH 7.4)

• All are soluble in water (charged)
• Form ionic bonds with negatively charged molecules (DNA, acidic lipids, acidic amino acids)
• Histidine has a pKa near physiological pH → acts as a proton acceptor/donor → key in enzyme catalysis, hemoglobin's Bohr effect, and intracellular buffering
• Arginine: precursor of NO (nitric oxide) via NO synthase → vasodilation; intermediate in urea cycle
• Lysine: essential amino acid; cross-links collagen and elastin via lysyl oxidase → important in connective tissue
• Vet relevance:
  • Lysine deficiency in poultry and pig diets → growth retardation (most limiting AA in grain diets); lysine supplementation of swine rations
  • Arginine is essential in cats (cannot synthesize ornithine from glutamate) → short arginine-free meal → life-threatening hyperammonemia
  • Histidine in hemoglobin: Bohr effect (H⁺ binds to His residues at high CO₂ → promotes O₂ release in tissues)

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Summary Table

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QUESTION 7 - Lipoproteins and Their Functions (5 marks)

Structure of Lipoproteins

• Outer shell: Phospholipids (monolayer, polar heads outward), free cholesterol, apolipoproteins
• Hydrophobic core: Triglycerides, cholesterol esters, fat-soluble vitamins

Classification by Density and Function

1. Chylomicrons

• Origin: Intestinal enterocytes (from dietary fat)
• Core lipid: Dietary triglycerides (>85%)
• Key apoprotein: ApoB-48, ApoC-II, ApoE
• Function: Transport exogenous (dietary) triglycerides from intestine to peripheral tissues (muscle, adipose) via lymph → blood
• Metabolism: ApoC-II activates lipoprotein lipase (LPL) on capillary endothelium → TG hydrolyzed → FFA released to tissues → chylomicron remnants taken up by liver (via ApoE-receptor)
• Vet note: Postprandial lipemia (milky plasma) in dogs/cats = normal after fatty meal; pathological chylomicronemia in lipoprotein lipase deficiency (cats, miniature Schnauzers)

2. VLDL (Very Low-Density Lipoprotein)

• Origin: Liver hepatocytes
• Core lipid: Endogenous triglycerides (55%)
• Key apoprotein: ApoB-100, ApoC-II, ApoC-III, ApoE
• Function: Transport endogenous triglycerides synthesized in liver to peripheral tissues
• Metabolism: LPL cleaves TG → releases FFA → VLDL becomes IDL, then LDL
• Vet note: Elevated VLDL in bovine fatty liver syndrome; hypothyroidism in dogs; equine metabolic syndrome

3. IDL (Intermediate-Density Lipoprotein)

• Origin: Formed from VLDL after TG removal
• Function: Transient intermediate; taken up by liver (via ApoE) OR converted to LDL
• Short-lived; not usually measured in veterinary diagnostics

4. LDL (Low-Density Lipoprotein) - "Bad Cholesterol"

• Origin: Formed from VLDL/IDL after most TG removed
• Core lipid: Cholesterol esters (50%)
• Key apoprotein: ApoB-100 only
• Function: Deliver cholesterol to peripheral cells via LDL receptor (ApoB-100 binds LDL receptor) → endocytosis → cholesterol released for membrane synthesis, steroid hormone production
• Vet note: In dogs (unlike humans), LDL is NOT the predominant lipoprotein - HDL is. LDL elevated in hypothyroid dogs, chronic kidney disease

5. HDL (High-Density Lipoprotein) - "Good Cholesterol"

• Origin: Liver and intestine; also formed from surface remnants of chylomicrons/VLDL
• Core lipid: Cholesterol esters (largest proportion relative to its size)
• Key apoprotein: ApoA-I, ApoA-II
• Function: Reverse cholesterol transport - removes excess cholesterol from peripheral tissues → transport to liver for excretion as bile acids
• ApoA-I activates LCAT (Lecithin-Cholesterol Acyl Transferase) → esterifies free cholesterol → incorporated into HDL core
• Vet note: HDL is the major plasma lipoprotein in dogs, cats, horses, cattle (unlike humans where LDL predominates). Cats completely lack CETP (cholesteryl ester transfer protein).

6. Lipoprotein(a) [Lp(a)]

• LDL-like particle with an extra apoprotein: Apo(a) linked to ApoB-100 by disulfide bond
• Structurally similar to plasminogen → may inhibit fibrinolysis → thrombotic tendency
• Measured in dogs; significance unclear in veterinary medicine

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Functions of Lipoproteins

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QUESTION 8 - Watson-Crick DNA Model + DNA vs RNA (5+4 = 9 marks)

A. Watson-Crick Double Helix Model (1953)

Features of the B-DNA Double Helix:

• DNA consists of two complementary strands wound around each other
• Strands run antiparallel: one 5'→3', the other 3'→5'
• The backbone (exterior) = alternating deoxyribose + phosphate groups (hydrophilic)
• The bases (interior) face inward (hydrophobic stacking)

• A=T: Adenine pairs with Thymine via 2 hydrogen bonds
• G≡C: Guanine pairs with Cytosine via 3 hydrogen bonds
• This ensures [A]=[T] and [G]=[C] in any double-stranded DNA (Chargaff's rule)
• G-C pairs = more stable (3 H-bonds); G+C content determines melting temperature (Tm)

• Unequal wrapping of the two strands creates:
  • Major groove (wider, ~22 Å): binding site for regulatory proteins, transcription factors
  • Minor groove (narrower, ~12 Å): binding site for some drugs (e.g., berenil in trypanosomiasis treatment in animals)

• Hydrogen bonds between bases (hold strands together)
• Base stacking interactions (π-π stacking of aromatic rings above/below each other - the strongest stabilizing force)
• Phosphodiester bonds (covalent, within each strand)

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B. Differences Between DNA and RNA

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QUESTION 9 - Membrane Transport & Membrane Lipids (5 marks each)

OPTION A: Types of Membrane Transport

1. Passive Transport (No energy - moves down gradient)

• Simple diffusion: O₂, CO₂, lipid-soluble molecules directly through bilayer
• Facilitated diffusion: Polar/charged molecules via carrier proteins (GLUT transporters for glucose) or channel proteins (aquaporins, ion channels)

2. Active Transport (Energy required - against gradient)

• Primary active transport: Direct ATP hydrolysis by ATPase pumps (Na⁺/K⁺-ATPase, Ca²⁺-ATPase, H⁺/K⁺-ATPase)
• Secondary active transport: Uses Na⁺ gradient (created by Na⁺/K⁺-ATPase) as energy source - SGLT1/2 (glucose symport), Na⁺/Ca²⁺ exchanger (antiport)

3. Vesicular Transport

• Endocytosis: Pinocytosis, phagocytosis, receptor-mediated endocytosis (colostral IgG in neonatal calves/foals)
• Exocytosis: Insulin secretion, neurotransmitter release

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OPTION B: Membrane Lipids and Their Functions

1. Phospholipids (Major component - ~50% of membrane lipids)

• Form the lipid bilayer barrier (hydrophilic heads outward, hydrophobic tails inward)
• Lung surfactant: Dipalmitoylphosphatidylcholine (DPPC) reduces alveolar surface tension - deficiency in neonatal foals/premature lambs causes respiratory distress syndrome
• Platelet activation: Externalized phosphatidylserine on activated platelets binds coagulation factors
• Signal transduction: PIP₂ → IP₃ (releases Ca²⁺) + DAG (activates PKC)
• Emulsification: Bile contains phosphatidylcholine for fat emulsification

2. Cholesterol (25-30% of membrane lipids in animal cells)

• Cell membranes of ruminants adjust cholesterol content seasonally for temperature adaptation
• Cholesterol toxicosis in horses (hyperlipemia): Ponies, donkeys in NEB → massive VLDL release → hyperlipidemia + hepatic lipidosis

3. Glycolipids (5-10% of membrane lipids, outer leaflet only)

• GM1/GM2 gangliosidosis: Lysosomal storage diseases in dogs (Beagles, Portuguese Water Dogs), cats, cattle - deficiency of ganglioside-degrading enzymes → accumulation in neurons → progressive neurodegeneration
• Feline blood types (A, B, AB): Determined largely by glycolipid antigens on RBCs - critical for blood transfusions (Type B cats have strong anti-A antibodies → neonatal isoerythrolysis in Type A kittens of Type B mothers)

Membrane Asymmetry

• Outer leaflet: Phosphatidylcholine, sphingomyelin, glycolipids, gangliosides
• Inner leaflet: Phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol

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QUESTION 10 - Methods of Urine Analysis and Diagnostic Applications (5+5 = 10 marks)

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I. Physical Examination of Urine

1. Color

2. Turbidity (Clarity)

• Normal: clear to slightly cloudy
• Turbid: Pyuria (pus), crystalluria, bacteriuria, lipiduria
• Vet note: Healthy horse urine is normally turbid due to calcium carbonate crystals and mucus (not pathological)

3. Volume (Urine Output)

• Normal urine output: dogs ~20-40 mL/kg/day; cats ~22-30 mL/kg/day
• Polyuria (PU): >50 mL/kg/day → DI, CKD, diabetes, hyperadrenocorticism, hypercalcemia
• Oliguria: <0.5 mL/kg/hr → acute kidney injury, severe dehydration, urethral obstruction
• Anuria: No urine production → severe AKI, complete obstruction

4. Specific Gravity (USG)

• Measures solute concentration (urine-concentrating ability of renal tubules)
• Measured by: Refractometer (most accurate), urinometer, dipstick (less accurate)

5. Odor

• Ammoniacal: urease-positive bacteria (UTI), alkaline urine, urea degradation
• Fruity/sweet: ketonuria (diabetes, pregnancy toxaemia, starvation)

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II. Chemical Examination (Urine Dipstick + Specific Tests)

1. pH

• Normal range: dogs/cats pH 6.0-7.5; pH varies with diet
• Alkaline urine (pH >7.5): UTI with urease bacteria, post-prandial alkaline tide, diet (plant-based), metabolic alkalosis
• Acidic urine (pH <6.0): Protein-rich diet, metabolic acidosis, respiratory acidosis
• Clinical use:
  • Struvite uroliths form in alkaline urine (cats, dogs) → treat by acidifying diet
  • Urate and oxalate uroliths form in acidic urine

2. Protein (Proteinuria)

• Detected by dipstick (reaction with tetrabromophenol blue - detects albumin primarily)
• Quantified by UPC (urine protein:creatinine ratio) - gold standard
• Normal: UPC <0.2 dogs, <0.4 cats
• Confirmed with SSA (sulfosalicylic acid) precipitation test - detects all proteins including Bence-Jones
• Causes: Glomerulonephritis, amyloidosis, UTI, CKD, post-exercise (physiological)

3. Glucose (Glycosuria)

• Detected by dipstick (glucose oxidase/peroxidase method) - specific for glucose
• Benedict's test: Non-specific reducing sugar test (also detects fructose, galactose, lactose) - useful in ruminants and neonates
• Normal: absent/trace
• Positive → diabetes mellitus, stress hyperglycemia (cats), Fanconi syndrome, renal glycosuria

4. Ketones (Ketonuria)

• Detected by dipstick - Rothera's nitroprusside test (detects acetone + acetoacetate; NOT β-hydroxybutyrate)
• Positive in: Diabetic ketoacidosis (dogs, cats), starvation ketosis, pregnancy toxaemia (ewes, does, dairy cows), bovine ketosis
• Cowside test for ketosis: Keto-Test strips for β-hydroxybutyrate in milk (most sensitive in cattle)

5. Bilirubin (Bilirubinuria)

• Only conjugated (direct) bilirubin appears in urine
• Dogs: Trace bilirubinuria may be normal (especially in concentrated urine of male dogs - kidney tubules conjugate some bilirubin)
• Cats: Any bilirubinuria is significant (low renal threshold)
• Detected by dipstick (diazonium salt reaction)
• Positive → hepatic disease, biliary obstruction, severe hemolysis

6. Urobilinogen

• Colorless product of bilirubin metabolism by intestinal bacteria; partially reabsorbed and excreted in urine
• Increased: Hemolysis, hepatocellular disease
• Absent: Complete biliary obstruction (no bilirubin reaching intestine → no urobilinogen → also absent in urine and stool)

7. Blood (Occult Blood / Hematuria / Hemoglobinuria)

• Dipstick detects heme (in RBCs, free hemoglobin, or myoglobin)
• Must differentiate: Hematuria (intact RBCs) vs. Hemoglobinuria vs. Myoglobinuria
• Hematuria confirmed by microscopy (RBCs in sediment)
• Causes: UTI, urolithiasis, trauma, tumor, coagulopathy, immune-mediated hemolytic anemia

8. Nitrite

• Produced by nitrate-reducing bacteria (Gram-negative)
• Positive → bacteriuria (UTI)
• Less reliable in veterinary species (urine often doesn't sit long enough for bacterial conversion)

9. Leukocyte Esterase

• Detects WBCs (neutrophils); indicates pyuria
• Not reliable in dogs and cats - species difference in WBC enzyme; confirmed by microscopy

10. Urine Creatinine

• Used to calculate UPC ratio and fractional excretion of electrolytes

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III. Microscopic Examination of Urine Sediment

Preparation:

• Centrifuge urine (400g × 5 min) → pour off supernatant → resuspend pellet → examine wet mount

A. Cells

B. Casts (Indicate Renal Origin)

C. Crystals

D. Bacteria, Fungi, Parasites

• Bacteria: Must distinguish contamination vs. true infection (quantitative culture)
• Candida sp.: Immunocompromised animals, prolonged antibiotic use
• Capillaria plica (Pearsonema plica): Bladder worm in dogs/cats → hematuria

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QUESTION 11 - Metabolic Acidosis and Alkalosis + Compensatory Mechanisms (6 marks)

• Metabolic Acidosis: definition, causes (DKA, lactic acidosis, diarrhea in calves, rumen acidosis, uremia), biochemical changes, respiratory and renal compensation
• Metabolic Alkalosis: definition, causes (abomasal displacement in cattle, GDV in dogs, vomiting), biochemical changes, respiratory and renal compensation, paradoxical aciduria
• Summary comparison table

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QUESTION 12 - Ketosis and Its Diagnosis (5+1 = 6 marks)

Ketosis

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Biochemistry of Ketone Body Formation (Ketogenesis)

1. Lipolysis in adipose → NEFA released into blood
2. NEFA → Liver → undergoes β-oxidation → Acetyl-CoA
3. When energy demand is high (starvation, NEB, uncontrolled DM) and oxaloacetate (OAA) is depleted (diverted to gluconeogenesis), Acetyl-CoA cannot enter TCA cycle
4. Excess Acetyl-CoA → Ketogenesis:
   • 2 Acetyl-CoA → Acetoacetyl-CoA
   • Acetoacetyl-CoA + Acetyl-CoA → HMG-CoA (via HMG-CoA synthase)
   • HMG-CoA → Acetoacetate (via HMG-CoA lyase)
   • Acetoacetate → β-hydroxybutyrate (via BHBA dehydrogenase; reduced by NADH)
   • Acetoacetate → Acetone (spontaneous decarboxylation - fruity smell in breath)

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Ketone Bodies and Their Fates

• Acetoacetate/BHBA → Acetyl-CoA → TCA cycle → ATP
• During prolonged starvation, brain adapts to use ketones as primary fuel (normally uses glucose only)
• Liver CANNOT use its own ketones (lacks succinyl-CoA transferase/thiophorase)

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Types of Ketosis in Veterinary Species

1. Bovine Ketosis (Acetonaemia)

• Occurs 2-6 weeks post-partum (peak milk production)
• High-producing cows cannot consume enough feed → Negative Energy Balance (NEB)
• Fat mobilization → elevated NEFA → hepatic ketogenesis
• BHBA >1.0 mmol/L = subclinical; >3.0 mmol/L = clinical

• Overconditioned cows at calving (BCS >3.5)
• Massive fat mobilization post-partum → fatty liver → impaired hepatic function → impaired gluconeogenesis → worsening ketosis
• High NEFA + high BHBA + elevated liver enzymes (AST, GLDH)

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2. Pregnancy Toxaemia (Small Ruminants)

• Ewes/does with multiple fetuses in late gestation
• Signs: blindness, teeth grinding, recumbency, coma
• BHBA >3 mmol/L; blood glucose <40 mg/dL

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3. Diabetic Ketoacidosis (DKA) - Dogs and Cats

• Severe insulin deficiency + stress hormones (glucagon, cortisol, epinephrine)
• Massive NEFA mobilization + ketogenesis
• Metabolic acidosis (anion gap) + dehydration + electrolyte imbalance
• Life-threatening; requires intensive care

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4. Equine Hyperlipaemia/Hyperlipidaemia

• Particularly in ponies, donkeys, and miniature horses under stress/NEB
• Massive fat mobilization → VLDL overload → hypertriglyceridemia + ketosis + hepatic lipidosis
• Triggers: late pregnancy, lactation, starvation, illness

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Diagnosis of Ketosis

1. Clinical Diagnosis

• History and clinical signs (milk drop, anorexia, neurological signs)
• Breath smell: fruity/acetone odor (acetone is volatile)

2. Bedside/Cowside Tests

3. Laboratory Tests

4. Herd Monitoring (Dairy Cattle)

• Routine milk BHBA testing at 5-7 days post-partum identifies subclinical ketosis
• BHBA >0.10 mmol/L in milk (Keto-Test) = subclinical ketosis → intervention

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QUESTION 13 - Phase I Metabolism of Xenobiotics (3 marks) OR Carbohydrate Classification (5 marks)

OPTION A: Phase I Metabolism of Xenobiotics

1. Oxidation (Most Important)

• Catalyzed by Cytochrome P-450 (CYP) enzyme system in hepatocyte ER
• Reaction: Substrate + O₂ + NADPH → Oxidized product + H₂O + NADP⁺
• Types: Hydroxylation (aromatic/aliphatic), epoxidation, N-oxidation, S-oxidation, deamination
• Examples in animals:
  • Barbiturate hydroxylation → more water-soluble (cats are slow at this)
  • Acetaminophen → NAPQI (N-acetyl-p-benzoquinoneimine) via CYP2E1 - toxic intermediate especially in cats
  • Carbon tetrachloride → CCl₃· free radical → hepatotoxicity
  • Aflatoxin B₁ → epoxide metabolite → carcinogen in cattle, poultry

2. Reduction

• Reduces -NO₂ (nitro) and -N=N- (azo) groups
• Nitrate reduction by rumen bacteria → nitrite → oxidizes hemoglobin → methemoglobin → nitrite poisoning in ruminants (grazing on nitrate-rich forages, preserved silage)

3. Hydrolysis

• Cleaves ester, amide, or glycoside bonds
• Atropine hydrolysis by atropinase (rabbits, goats can eat deadly nightshade safely - they have plasma atropinase)
• Organophosphate hydrolysis by paraoxonase (PON1) - species differences (horses have very low PON1 activity)

Key Enzyme: Cytochrome P-450 (CYP)

• Heme-containing mixed-function oxidases
• Located in ER of hepatocytes (also intestine, lung, skin)
• CYP1A2, CYP2B6, CYP2C, CYP2D6, CYP2E1, CYP3A4 - different substrate specificities
• Species differences: Cats have reduced CYP activity for certain substrates → drug toxicity (aspirin, acetaminophen, phenols, NSAIDs)

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OPTION B: Classification of Carbohydrates

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Classification of Carbohydrates

I. MONOSACCHARIDES (Simple Sugars - Cannot be hydrolyzed further)

• Aldoses: Have aldehyde group (-CHO) at C1 → e.g., Glucose, Galactose, Mannose, Ribose
• Ketoses: Have ketone group (C=O) at C2 → e.g., Fructose, Ribulose, Xylulose

• Exist in ring forms (pyranose = 6-membered; furanose = 5-membered) in solution
• All are reducing sugars (free anomeric carbon can reduce Cu²⁺ in Benedict's test)
• Show mutarotation and optical activity (D- and L- forms)

• Glucose (D-glucose): Primary blood sugar; universal fuel; normal blood glucose: dogs 70-120 mg/dL; cats 70-150 mg/dL; cattle 45-75 mg/dL; horses 70-110 mg/dL
• Galactose: From lactose digestion; converted to glucose-1-phosphate (Leloir pathway) in liver
• Fructose: From sucrose; fruit sugar; phosphorylated by fructokinase in liver (bypasses PFK regulation)
• Ribose/Deoxyribose: Pentoses; components of RNA/DNA nucleotides

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II. DISACCHARIDES (2 monosaccharides joined by glycosidic bond)

• Lactose intolerance = deficiency of intestinal lactase → undigested lactose → osmotic diarrhea. Common in adult carnivores (cats/dogs lose lactase after weaning)
• Sucrose is not normally present in significant amounts in animal feeds; excess sucrose → equine metabolic disorders

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III. OLIGOSACCHARIDES (3-10 monosaccharides)

• Short chains of 3-10 sugar units linked by glycosidic bonds
• Often attached to proteins (glycoproteins) or lipids (glycolipids) via N- or O-glycosidic bonds
• Examples: Raffinose (3 units, in legumes → flatulence in monogastrics), Stachyose (4 units), Human milk oligosaccharides (HMO), Blood group antigens

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IV. POLYSACCHARIDES (>10 monosaccharides - most with thousands of units)

A. Homopolysaccharides (one type of monosaccharide)

• Two forms: Amylose (linear, α-1,4 bonds) + Amylopectin (branched: α-1,4 + α-1,6 bonds every 24-30 glucose units)
• Digested by amylase (salivary + pancreatic) → maltose → glucose
• Major carbohydrate source in monogastric rations (pigs, poultry, dogs, cats)
• Ruminants: Starch digested predominantly by ruminal microbes (amylolytic bacteria) → VFAs (acetate, propionate, butyrate); excess starch → rumen acidosis

• More branched than starch: α-1,4 bonds + α-1,6 branches every 8-12 glucose units
• Stored in liver (up to 5-6% of liver weight; ~100g in adult dog) and muscle (1-2% of muscle weight)
• Liver glycogen: Maintains blood glucose during fasting (glycogenolysis)
• Muscle glycogen: Provides energy for muscle contraction; CANNOT release glucose to blood (no glucose-6-phosphatase)
• Vet relevance:
  • Glycogen storage diseases (GSD): GSD Type I (von Gierke's) - glucose-6-phosphatase deficiency - reported in dogs
  • Exertional rhabdomyolysis in horses (Polysaccharide Storage Myopathy/PSSM): Abnormal glycogen accumulation in muscle; treat with low-starch, high-fat diet
  • Neonatal hypoglycemia in piglets/puppies: limited glycogen stores → rapid hypoglycemia when not feeding

• Linear polymer of glucose with β(1→4) glycosidic bonds
• Cannot be digested by any vertebrate enzyme
• Ruminants: Rumen microbes (Fibrobacter succinogenes, Ruminococcus sp.) produce cellulases → digest cellulose → VFAs → energy (up to 70% of ruminant energy needs)
• Horses: Hindgut fermentation (cecum + large colon) of cellulose
• Monogastrics (dogs, cats, pigs): Cannot digest cellulose; acts as dietary fiber → bulks stool, feeds colonic microbiome

• Polymer of N-acetyl-D-glucosamine with β(1→4) bonds
• Major component of insect exoskeletons, crustacean shells, fungal cell walls
• Digested by chitinase (acid mammalian chitinase in gastric juice of insectivores)

• Fructose polymer (β-2,1 bonds)
• Found in roots of chicory, Jerusalem artichoke
• Exogenous use: Gold standard for GFR measurement (inulin clearance) in all veterinary species
• Dietary use: Prebiotic fiber in some pet foods

B. Heteropolysaccharides (two or more types of monosaccharides)

• Long unbranched chains of repeating disaccharide units
• One unit = amino sugar (GlcNAc or GalNAc) + uronic acid (GlcUA or IdUA)
• Often sulfated and linked to proteins = proteoglycans

• Hyaluronic acid injections used in equine joint disease (osteoarthritis)
• Chondroitin sulfate + glucosamine supplements widely used for joint disease in dogs, horses
• Mucopolysaccharidoses (MPS): Lysosomal storage diseases in cats (MPS I, VI, VII) and dogs (MPS III, VII) due to enzyme deficiency → GAG accumulation in tissues

• Short oligosaccharide chains covalently attached to proteins via N- or O-glycosidic bonds
• Examples: Mucins (airway mucus), blood group antigens, collagen, immunoglobulins
• Mucus in respiratory and GI tract is a glycoprotein gel - protects epithelium from pathogens

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Sources: Basic Medical Biochemistry (Lieberman & Peet, 6e); Biochemistry, Lippincott Illustrated Reviews (8e); Harper's Illustrated Biochemistry (32e); Robbins & Kumar Pathologic Basis of Disease (10e); Henry's Clinical Diagnosis and Management by Laboratory Methods. All content applied to veterinary species throughout.