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Practical Biochemistry

A young infant, who was nourished with a synthetic formula, had a sugar in the blood and urine. This compound gave a positive reducing sugar test but was negative when measured with glucose oxidase (specific test for detection or estimation of Glucose). Treatment of blood and urine with acid (which cleaves glycosidic bonds) did not increase the amount of reducing sugar measured. Which of the following compounds is most likely to be present in this infant’s blood and urine?
A. Glucose
B. Fructose
C. Maltose
D. Sorbitol
E. Lactose

The right answer is fructose.

Reducing sugars are usually detected by Benedict’s reagent, which contains copper sulphate, sodium citrate and sodium carbonate. Sodium carbonate makes the medium alkaline. Copper sulphate furnishes Cu2+ ions and sodium citrate prevents the precipitation of cupric ions as cupric hydroxide by forming a loosely bound cupric- sodium –citrate complex which on dissociation gives a continuous supply of cupric ions.

Benedict’s test


Carbohydrates with free aldehyde or ketone groups have the ability to reduce solutions of various metallic ions. Reducing sugars under alkaline conditions tautomerise and form enediols. Enediols are powerful reducing agents. They reduce cupric ions to cuprous form and are themselves converted to sugar acids. The cuprous ions combine with OH- ions to form yellow cuprous hydroxide which upon heating is converted to red cuprous oxide.


Take 5 ml of Benedict’s reagent. Add 8 drops of carbohydrate solution. Boil over a flame or in a boiling water bath for 2 minutes. Let the solution cool down.


Benedict’s test is a semi quantitative test. The color of the precipitate gives a rough estimate of a reducing sugar present in the sample (figure-1)

Green color- Up to 0.5 g %(+)

Green precipitate -0.5-1.0 g %(++)

Yellow precipitate -1.0-1.5 g %(+++)

Brick red precipitate- >2.0 G% (++++)

Negative benedict's testPositive benedict's test

(-ve)                (+ve)

Figure– The positive test is given by reducing sugars. The color of the precipitate determines the rough estimate of the reducing sugar present in the given sample.

Fehling test is an alternative to Benedict’s test. It differs from Benedict’s test in that it contains sodium potassium tartrate in place of Sodium citrate and potassium hydroxide as an alkali in place of sodium carbonate in Benedict’s reagent. It is not a preferred test over Benedict’s test since the strong alkali present causes caramelisation of the sugars; hence it is less sensitive than Benedict’s reagent.

Positive Benedict’s test for urine signifies Glycosuria.

Glycosuria is a non-specific term. Glucosuria, lactosuria, galactosuria, pentosuria and fructosuria denote the presence of specific sugars in urine.

Causes of Glycosuria are:

a. Renal glycosuria

b. Diabetes mellitus

c. Alimentary glucosuria

d. Hyperthyroidism, hyperpituitarism and hyperadrenalism

e. Stress, severe infections, increased intracranial pressure

Lactosuria– in lactose intolerance

Galactosuria– in galactosemia

Fructosuria– in hereditary fructose intolerance

Pentosuria – in essential pentosuria

Examples of non-carbohydrate substances which give a positive Benedict’s reaction are:

a) Creatinine

b) Ascorbic acid

c) Glucuronates

d) Drugs: Salicylates, PAS and Isoniazid.

Glucose oxidase test is a specific enzymatic method for the determination and estimation of glucose present in a given sample. True glucose can be estimated by this method.

As regards other options

Glucose cannot be present since specific test is negative.

Sorbitol is non reactive to reduction test.

Maltose and lactose would have caused increase in the amount of reducing sugar upon acid hydrolysis.

Hence it is fructose which is reducing in nature but non reactive to glucose oxidase.


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Q.1- What is the range of serum creatinine in normal health?

Answer- The serum creatinine ranges between

1) In children (<12 years) 0.25-0.85 mg/dl

2) Adult male 0.7-1.5 mg/dl

3) Adult female 0.4-1.2 mg/dl

Q.2- What are the conditions of high serum creatinine levels?

Answer- Higher levels are observed in – Renal failure (All causes) and in muscular dystrophies. Falsely high levels are observed in diabetic ketoacidosis.

Q.3- If in a patient, serum creatinine has been found to be higher than normal but blood urea is within the normal range, what is the likely possibility?

Answer- It can not be renal failure because in such a state both blood urea and serum creatinine should have been higher, since both are excreted by kidney through urine in normal health. But since blood urea is normal, it could be any other reason and the most likely cause is muscular dystrophy. The diagnosis can be made from the history and clinical symptoms. The other possibility can be of false high value as in diabetic keto acidosis.

Q.4- In a patient with normal serum creatinine level, blood urea has been found to be much higher than normal, what could be the possibility?

Answer- Both urea and creatinine should be higher than normal in renal failure, if creatinine is normal the possibility of renal failure can be ruled out. Blood urea can be higher than normal in conditions of- Advancing age, high protein diet, dehydration, catabolic state and in post renal obstructive conditions (Stone, stricture, growth etc)

Q.5- What is the principle of alkaline picrate method (Jaffe’s reaction) for the estimation of serum creatinine?

Answer- Creatinine under alkaline conditions reacts with Picric acid to form Creatinine picrate (An orange-red colored complex), the intensity of which is measured at 520 nm.

Q.- 6- Alkaline picrate method is considered a less sensitive method for creatinine estimation, what are the other substances which can give a positive reaction with alkaline picrate?

Answer- Jaffe’s reaction is not specific for creatinine. In serum up to 20% of the total chromogens (Color forming substances) can be substances other than creatinine which give a positive reaction with alkaline picrate, while in urine these are only 5%. Other non specific chromogens that react with Picric acid are – proteins, ketone bodies, pyruvate, glucose and Ascorbate.

Q.7- What is the difference between creatine and creatinine?

Answer- Creatinine is the anhydrous product of creatine. Creatine is converted to creatinine non enzymatically by the loss of one molecule of water. About 2 % of creatine is converted to Creatinine daily.

Q.8- Which form out of creatine and creatinine is present in urine in normal health?

Answer- In normal urine creatinine is mainly present, creatine is present only in trace amounts.

Q.9-Name the amino acids that contribute towards creatine synthesis

Answer- Creatine is synthesized from Glycine, Arginine and Methionine. In the first step, Glycine and Arginine combine together to form Guanido Acetic acid, this reaction takes place in kidney. In the second step, Guanido acetic acid is methylated by Methionine to form Methyl Guanido acetic acid (Creatine). This reaction takes place in liver. Creatine is transported to muscles, where it is phosphorylated and stored in the form of creatine-P. 98% of the total amount of creatine is present in muscles.

Q.10- What is Lohmann reaction?

Answer- Creatine is phosphorylated to creatine-P by the enzyme Creatine kinase, present in muscle, brain and myocardium. The stored creatine phosphate in the muscle serves as an immediate source of energy. During muscle contraction, the energy is first derived from ATP hydrolysis. Thereafter, the ATP is generated by the -hydrolysis of creatine-P. The high energy phosphate is transferred to ADP to form ATP. This reaction is called Lohmann reaction.  In the resting muscles the creatine-P is restored at the expense of ATP provided from glycolysis.

Q.11- What are the common causes of creatinuria?

Answer- Excretion of creatine in urine is called Creatinuria, which is observed under the following conditions-

1) In children- Probably due to impaired conversion of creatine to creatinine

2) Pregnancy

3) Febrile conditions

4) Thyrotoxicosis

5) Muscular dystrophies, myositis and Myasthenia gravis

6) Uncontrolled diabetes mellitus

7) Starvation

8) Wasting diseases- such as Malignancies.

Q.12- What is creatinine co-efficient? What is its significance?

Answer- It is the ratio of- mg of creatinine in urine in 24 hours/ Body weight in kg.

The value is 20-26 for males and 14 to 22 in females.

Significance- It depends on muscle mass and remains fairly constant. Since muscle mass remains constant in a given individual, the creatine coefficient serves as a reliable index of the adequacy of a 24 hour urine collection

Q.13- What is the reason for high creatinine level in males in normal health?

Answer- Creatine is synthesized in liver, passes in to circulation and is almost taken entirely by the skeletal muscle for conversion to creatine-P, which serves as a storage form of energy in skeletal muscles. About 2% of creatine is converted to Creatinine daily. Since its concentration is related to the muscle mass and males have more muscle mass that is why the level of serum creatinine is higher in males in normal health.

Q.14- What is the normal range of creatinine clearance? What is the significance of measuring creatinine clearance?

Answer- The normal values of creatinine clearance are-

Males- 95-140 ml/minute

Females-85-125 ml/minute

These values are close to GFR (Glomerular filtration rate). Clearance values are decreased in impaired renal functions and so provide a rough measure of renal damage.

Q.15- Out of urea and creatinine clearance, the estimation of which clearance is preferred to assess renal functional status and why?

Answer- Unlike urea, serum Creatinine level is not affected by diet, age, dietary factors or by fluid depletion. Creatinine is filtered but is not absorbed by the tubules (unlike urea), hence it is a better predictor of GFR. (The values are slightly higher than GFR due to tubular secretion). The methodology is also simple, due to all these reasons; creatinine clearance is preferred over urea clearance for determining the functional status of the kidney.

Q.16-   Calculate the creatinine clearance of a patient with serum creatinine of 3 mg/dL, volume of urine excreted 1500 ml/ day and urinary creatinine of 0.75 G/L 

Answer- Creatinine clearance(C)= UV/P

Where U= Urinary creatinine (mg/dl)

V= Volume of urine excreted (ml/day)

P= Serum Creatinine (mg/dl)

Thus applying the values-

V= 1500 ml/day, convert to ml/ minutes

i.e. = 1500/24×60= 1.1 ml/minute (Approximately)

U= 0.75 G/L, Convert it to mg/dl

i.e. – 75 mg/dl

Creatinine clearance (C) = 75×1.1/3

   = 27.5 ml/minute

It is much below the physiological range; hence it is a case of impaired renal functions.

Q.17- Comment upon the functional status of the kidney, if the serum creatinine is 4.5 mg/dL, blood urea- 86 mg/dL  and serum uric acid as 12 mg/dl.

Answer- Urea, creatinine and uric acid are normally excreted by kidney through urine, since the levels of three of them are higher than normal in the given case that means kidney is failing to clear out these substances from blood that is why they are accumulating in blood, hence it is a case of impaired renal functions, possibly renal failure.

Q.18- A patient with long-standing diabetes mellitus has reported to emergency with generalized swelling of the body. Blood biochemistry reveals-

Hb- 8 G/dL

F.B.S- 260 mg/dl

Blood urea- 98 mg/dl

Serum creatinine- 3.4 mg/dL

Serum Uric acid- 10.8 mg/Dl

Urine analysis- Sugar ++++


Urea clearance 32 ml/minute

Creatinine clearance- 68 ml/minute

Comment on the findings and provide a provisional diagnosis.

Answer- It is a case of renal failure due to long-standing diabetes mellitus. Blood urea, serum creatinine and uric acid are high, clearance values are low, Hb is low due to decreased Erythropoietin, sugar and protein in urine are suggestive of Diabetic nephropathy which has progressed to renal failure.

Q.19- Out of serum creatinine and blood urea, which is more sensitive indicator of falling renal functions?

Answer- Serum creatinine is more sensitive indicator of falling renal functions than blood urea.  Urea level is affected by non renal causes also, while creatinine is a relatively a stable parameter, hence its measurement carries more significance to assess falling functional status of the kidney.

Q.20- In a patient with diabetic ketoacidosis, creatinine is high while blood urea is normal, what is the possibility?

Answer- Jaffe’s reaction for estimation of serum creatinine gives positive reaction with Glucose and ketone bodies also, which are high in diabetic ketoacidosis. It is falsely high level due to other chromogens and not due to creatinine. Normal blood urea level indicates normal renal functions.

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                    [Blood (B), Serum (S), Plasma (P),Urine (U)]
Blood Gases
(B- Arterial) 7.35 – 7.45 (H+44.7-45.5 nmol/L)
 Partial pressure of CO
 ( PaCO2)
(B-Arterial) 35 – 45mm Hg(4.7-6 kPa
Partial pressure of O2(PaO2 )
(B-Arterial 80) – 100mm Hg(10.67-13.33kPa)
Bicarbonate (HCO3)
24-28 meq/L(24-28 mmol/L)
O2 Sat %( Arterial)
O2 Sat %( Venous)
CO2 content
Total Serum  21-30 mmol/L
<5% of Hemoglobin
Anion gap
7–16 mmol/L
(B)Men-14-18g/dl (2.09-2.79mmol/L)
Women-12 – 16 g/dL (1.86-2.48 mmol/L)
(S) 2-3mg/dl
Haemtocrit (PCV)
Men 40 – 52% (0.4-0.52)
Women 37 – 47%(0.37-0.47)
 Red Blood Count (RBC)
0.2-2% of red cells
White blood Count(WBC)
5000-10,000/ μl (5-10×109/L)
  • Polymorph nuclear (PMN): 35-80%
  • Immature Polys (Bands): 0-10%
  • Lymphocytes (Lymp): 20-50%
  • Monocytes (Mono): 2-12%
  • Eosinophils (Eos): 0-7%
  • Basophils (Bas): 0-2%
150,000-400,000/ μL(0.15-0.4×1012/L)
Mean Corpuscular volume(MCV)
Men-80-94f L
Women-81-99f L (By coulter counter)
Mean Corpuscular Hemoglobin(MCH)
Mean Corpuscular Hemoglobin Concentration(MCHC)
32-36g/dl, red blood cells (32-36%)
Average diameter of red cell
7.3μm(5.5-8.8 μm)
Bleeding time
Ivy method, 1-7 minutes(60-420 seconds)
Template method, 3-9 minutes (180-540seconds).
Clot retraction
Begins in 1-3 hrs: complete in 6-24 hrs. No clot lysis in 24 hours.
Fragility of red cells
Begins at 0.45-0.38%NaCl, complete at 0.36-0.3% NaCl
Partial Thromboplastin time
Activated, 25-37seconds
Prothrombin Time
(P)11-14.5 seconds, International Normalized Ratio(INR)- (P)2.0-3.0
Erythrocyte Sedimentation Rate (ESR or Sed-Rate)
Male: 1 – 13 mm/hr
: 1 – 20 mm/hr
Blood Volume
8.5 – 9.1% of total body weight
Adrenals hormones
(P) 8.00 AM, 5-25μg/dl(138-690nmol/L)
8.00 PM<10 μg/dl(275 nmol/l)
(P)  Supine 2-9 ng/dl (56-250 pmol/L)Increased when upright
(P) <135 pg/ml
(P)Supine,<100 pg/ml(<550pmol/l)
Nor Epinephrine
(P)Supine,<500 pg/ml(<3nmol/l)
Adrenal hormones and metabolites
(U)2-26μg/24 hrs (5.5-72 nmol/d), values vary with sodium and potassium intake
(U)Total,<100 μg//24 hrs
(U)<10 μg//24 hrs(<100 nmol/d)
Nor Epinephrine
(U)<100 μg//24 hrs(<590nmol/d)
Free(U)-20-100 μg//24 hrs(0.55-2.76μmol/d)
11,17 OH corticoids
(U)Men-4-12mg/24 hrs
Women-4-8mg/24 hrs
17- keto steroids
(U)< 8 Years 0-2mg/24 hrs
Adolescents-2-20mg/24 hrs(1mg=3.5μmol)
(U)<1.3 mg/24 hrs(<6.6 μmol/d) or <2.2μg/mg creatinine
Vanillyl Mandelic acid(VMA)
(U)Up to 7 mg/24 hrs(< 35 μmol/d)
Growth hormone(GH)
(S) Adults, 1-10 ng/ml, (46-465 pmol/L) by RIA
Thyroid Stimulating hormone(TSH)
(S) < 10μU/ ml
Follicle stimulating hormone (FSH)
(S) Pre pubertal-, 2-12 m IU/ml
Adult men- 1-15 m IU/ml
Adult Women-1-30 m IU/ml
Luteinizing hormone(LH)
(S) Pre pubertal-, 2-12 m IU/ml
Adult men- 1-15 m IU/ml
Adult Women-< 30 m IU/ml
Corticotropin (ACTH)
(P) 8.00-10.00 AM, up to 100pg/ml, (22pmol/L)
(S) 1-25ng/ml (0.4-10 nmol/L)
Somatomedin C
(P) 0.4-2U/ml
Anti diuretic Hormone (ADH, Vasopressin)
(P)Serum osmolality285 mosm/kg, 0-2 pg/ml; >290 mosm/kg. 2-12 + pg/ml
(S) Men and non pregnant women <0.2μg/dl (<7nmol/L) by RIA
Chorionic Gonadotropin
(S) Beta subunit; Men-< 9 mIU/ mL
Pregnant Women->10 mIU/ mL
Testosterone, free
(S) Men, 10-30 ng/dl; Women 0.3-2 ng/dl. (1ng/dl=0.035nmol/L)
Testosterone, Total
(S) Pre pubertal , < 100 ng/dl ; Adult men, 300-1000 ng/dl; Adult women 20-80 ng/dl, luteal phase up to 120 ng/dl
(S) Men- 12-34 pg/ml;
women, menstrual cycle 1-10 days 24-68 pg/ml
11-20 days, 50-300 pg/ml
21-30 days, 73-149 pg/ml by RIA, (Ipg/ml=3.6 pmol/L)
(S) Follicular phase, 0.2-1.5 ng/ml
luteal phase, 6-32 ng/ml;
Pregnancy,>24 ng/ml,
Men, < 1ng/ml= 3.2 nmol/L
Thyroxin, free T4 (FT4)
(S) 0.8-2.4ng/dl (10-30 p mol/L)
Thyroxin, Total (TT4)
(S) 5-12 μg/dl(65-156nmol/L) by RIA
Thyroxin binding globulin capacity
(S) 12-28 μg T4/dl (150-360 nmol T4/dl)
Tri iodo thyronine (T3)
(S) 80-220 ng/dl (1.2-3.3 nmol/L)
Reverse Tri iodo thyronine (r T3)
(S) 30-80 ng/dl (0.45-1.2 nmol/L)
Tri iodo thyronine uptake(RT3U)
(S) 25-36% as TBG assessment(RT3U ratio) 0.85-1.15
(S)<100 pg/ml (< 29.2pmol/L)
(S) 4-25μU/mL(29-181pmol/L)
C- peptide
(S) 0.9-4.2ng/mL
(S), fasting) 20-100pg/mL
Parathyroid hormone (intact)
(S)8–51 pg/mL.
(S) up to 100 pg/ml (47 p mol/L).
Elevated, > 200 pg/ml
Pepsinogen I
(S) 25-100 ng/ml

Iron Studies

Adult Women- 20-120ng/ml
Child-15 years-7-140ng/ml
(S) 50-175 μg/dl(9-31.3 μmol/L)
Iron Binding capacity
(S)Total- 250-410 μg/dl(44.7-73.4μmol/L)
% saturation- 20-55%
(S) 200-400 mg/dl(23-45μmol/L)
(S) 40-170mg of Hb binding capacity
Blood Chemistry
Alanine Aminotransferase (ALT) SGPT
0-45 IU/L at 37°C
Aspartate Aminotransferase (AST) SGOT
0-41 IU/L at 37°C
Alkaline Phosphatase (ALP)
(S) Adults- 5-13 units(KA), 0.8-2.3(Bessey- Lowry):SMA 30-85 IU/L at 37°C: SMAC 30-115IU/L at 37°C
(S)- 80-180 units/dl (Somogyi)
Gamma Glutamyl Transpeptidase
(S) <30 units/L at 30°C
(S) <150 units/L
Acid phosphatase
(S)- 1-5 U(KA),0.1-0.63U(Bessey- Lowry)
Total Creatine Kinase (Total CK)
(S)-10-50 IU/L at 30°C
Creatine Kinase-MB (CK-MB)
Creatine Kinase-MM (CK-MM)
Creatine Kinase-BB (CK-BB)
Lactate dehydrogenase
(S)-55-140 IU/L at 30°C
(S) 1.5-2.0 U/L
<0.4 ng/ml
5′ Nucleotidase
0–11 U/L
Anti-streptolysin O Titer (ASO)
(S or P) 65-110 mg/dl(3.6-6.1 mmol/l)
Impaired glucose tolerance
111–125 mg/dL (6.2–6.9 mmol/L)
Diabetes mellitus
>125 mg/dL(>7.0 mmol/L)
Glucose, 2 h postprandial
70–120 mg/dL(3.9–6.7 mmol/L)
<285 μ mol/L
Hemoglobin Alc
Urea Nitrogen
(S or P) 8-25 mg/dl (2.9-8.9 mmol/)
(S or P) 0.7-1.5 mg/dl (62-132μmol/)
Uric acid
(S or P) Men-3-9 mg/dl(0.18-0.54 mmol/)
Women-2.5-7.5 mg/dl(0.15-0.45 mmol/)
(S) 24-28 meq/L(24-28mmol/L)
(S or P) 3.5-5 meq/L (3.5-5 mmol/L)
(S or P) 136-145 meq/L (136-145mmol/L)
(S or P) 96-106 meq/L( 96-106mmol/L)
(S or P)1.8-3 mg/dl(0.75- 1.25mmol/L)
(S or P) 100-200μg/dl(16-31 μmol/L)
(S) 50-150 μg/dl (7.65-22.95 μmol/L)
Phosphorus, inorganic
(S –fasting) 3-4.5 mg/dl (1-1.5mmol/L)
(S) 8.5-10.3 mg/dl (2.1-2.6mmol/L)
Calcium (ionized)
(S) 4.25-5.25 mg/dl; 2.1-2.6 meq/L (1.05-1.3 mmol/L)
(U)< 80 μg/24 hrs (< 0.4μmol/d)
(B) 0.6-1 mg/dl (70-114 μmol/L)
(B) Venous-, 4-16 mg/dl, (0.44-1.8mmol/L)
Ketone (acetone)
Acetone and Acetoacetate
(S) 0.3-2 mg/dl (3-20 mg/L)
(S)0–3 mg/dL(0–290 μmol/L)
Serum Total Protein
(S) 6-8g/dl (60-80 g/L)
(S) 3.5 -5.5 g/dl (35-55 g/L)
170–340 mg/L
(S)2-3.6 g/dl ( 20-36 g/L)
(P) 0.2-0.6 g/dl (2-6 g/L)
4.4–10.8 μmol/L
α1 –Antitrypsin
(S)> 180 mg/dl
C-reactive protein
0.2–3.0 mg/L
Prostate-Specific Antigen (PSA)
0 – 4 ng/mL (likely higher with age)
(S) 25-43mg/dl (1.7-2.9μ mol/L)
(S) Total, 0.2-1.2 mg/dl (3.5-20.5 μ mol/L)
Direct- (Conjugated), 0.1-0.4 mg/dl (<7 μ mol/L)
Indirect, 0.2-0.7 mg/dl (<12 μ mol/L)
(U) 0-2.5mg/24 hrs (70-470 μ mol/d)
40-280 mg/24 hrs (70-470 μ mol/d)
Delta Amino Levulinic acid (U)
Prophobilinogen (U)
1.5-7.5 mg/24 hrs (11-57 μ mol/d)
< 2 mg/24 hrs (<8.8 μ mol/d)
(P) 10-80 μg/dl
(S) 280-296mosm/kgwater (280-296 mmol/kg water)
Specific Gravity
(B) 1.056(Varies with Hb and protein concentration)
(S) 1.0254- 1.0288(varies with protein concentration)
Fecal Fat
<30% dry weight
Lipid profile
Cholesterol mg/dL (mmol/L) 
(S or P) 150-250 mg/dl (3.9- 5.72 mmol/L)
<200 (<5.17)- desirable
 200–239 (5.17–6.18) –borderline high
>240 (>6.21) – high
(S)< 165 mg/dl (1.9 mmol/L)
Lipid fractions
Desirable levels
HDL Cholesterol mg/dL(mmol/L)  
<40 (60 (>1.55)- high
LDL Cholesterol mg/dL(mmol/L) 
i) <70 (<1.81)
-Therapeutic option for very high risk patients
 ii) <100 (<2.59)
iii) 100–129
    (2.59 –3.34)
 –  Near optimal/above   optimal
 iv) 130–159
   (3.36– 4.11)
-Borderline high
 v)  160–189               (4.14–4.89)
  vi) >190 (4.91)
-Very high
VLDL Cholesterol
<40 mg/dl
Free fatty acid
(P) 200-800μmol/L
Lipoprotein (a)
0–30 mg/dL
Vitamin A
(S) 15-60 μg/dl (0.53-2.1 μmol/L)
β Carotene
(S) Fasting) 50-300 μg/dl
Vitamin B12
(S)> 200 pg/ml (>148 pmol/L)
Vitamin D
(S) Cholecalciferol (D3); 25- hydroxy cholecalciferol, 8- 55 ng/ml (19.4-137 nmol/L);
1,25 dihydroxy cholecalciferol, 26-65 pg/ml (62-155pmol/L);24,25-dihydroxy cholecalciferol,
1-5ng/ml (2.4-12 nmol/L).
Vitamin C (Ascorbic acid)
(P) 0.4-1.5 mg/dl (23-85μmol/L)
Folic acid
(S) 14-34 nmol/L
Vitamin E
(S) 5–18 μg/mL(12–42μ mol/L)
Vitamin K
(S) 0.13–1.19 ng/mL(0.29–2.64 nmol/L)
Vitamin B6
(P) 5–30 ng/mL(20–121 nmol/L)
Ig A
9.0-33 g/L
Ig G
7.2-15.0 g/L
Ig M
0.5-2.5 g/L
Ig D
0-0.4 g/L
Ig E
100-200 μg/L
Renal function Tests
p- Amino Hippurate(PAH)  clearance (RPF)
Men, 560-830 ml/min,
Women, 490-700 ml/min.
Creatinine Clearance, endogenous (GFR)
Men, 110-150 ml/min.
Women, 105-132 ml/min.
Inulin clearance
Approximately same as creatinine( corrected to 1.73 m2 surface area
Filtration fraction(FF)
Men-17-21%; women, 17-23 %(FF=GFR/RPF)
(U)On normal diet and fluid intake: Range 500-850 mOsm/kg water. Achievable range, normal kidney: Dilution 40-80 mOsm; concentration(dehydration) up to 1400 mOsm/kg water(At least three to four times plasma osmolality)
Specific gravity of urine
Urine volume
 0.4 -2.0 L/day.   
Urine pH
Acidity, titratable
20–40 meq/d(20–40 mmol/d)
30–50 meq/d(30–50 mmol/d)
Specific Gravity
Urine Ketone
Urine blood
Urine Proteins
Urine Nitrites
Negative, Traces
Urine Bilirubin
0-2.5mg/24 hrs
Urine Micro
  • RBCs: 0-2/HPF
  • WBCs: 0-2/HPF
  • RBC Casts: 0/HPF
Urine Glucose
< 250mg/dl
Urine Creatinine
Men-1.0-2.0 g/d
Women-0.8-1.8 g/d
Urea nitrogen
6–17 g/d(214–607 mmol/d)
Uric acid (normal diet)
250–800 mg/d(214–607 mmol/d)
Sodium (varies with intake)
100–260 meq/d(100–260 mmol/d)
Potassium (varies with intake)
25–100 meq/d(25–100 mmol/d)
Phosphate (phosphorus) (varies with intake)
400–1300 mg/d(12.9–42.0 mmol/d)
Micro albumin
0–30 mg/d(0.0–0.03 g/d)
30–300 mg/d(0.03–0.30 g/d)
Clinical albuminuria
>300 mg/d(>0.3 g/d)
Micro albumin/creatinine ratio
0–30 μg/mg creatinine(0–3.4 g/mol creatinine)
30–300 μg/mg creatinine(3.4–34 g/mol creatinine)
Clinical albuminuria
>300μg/mg creatinine(>34 g/mol creatinine)

Cerebral Spinal Fluid


60-150 mm Hg

Specific gravity

1.006 to 1.007

p H

CSF glucose
45-100 mg/dl
CSF Proteins
10-45 mg/dl
CSF Chlorides
700-760 mg/dl (120-130 meq/L) as NaCl
CSF urea
20-40 mg/dl
CSF Calcium
5.5-6 mg /dl
CSF cells
0-4 mononuclear per
Gram:common measurement of weight. Used in this table: pg (picograms), g (grams), mg(milligrams), etc. per liter
micrometer (µm) : a unit of length. Mean Corpuscular Volume is expressedin cubic micrometers
mole: also “gram molecular weight,” a quantity based on the atomicweight of the substance. Many test results in the System Internationale areexpressed as the number of moles per liter. In US units, these measurements areusually in grams per liter. Used in this table: mmol (millimoles), µmol,(micromoles), nmol (nanomoles), pmol (picomoles) per liter

Some units of measurement include the following fractions and multipliers:
mega (M) : 10 6 or x1,000,000
kilo (k) : 10 3 or x1,000
deca or deka : 10 1 or x10
deci (d) : 10 -1 or ÷10
milli (m) : 10 -3 or ÷1,000
micro (µ) : 10 -6 or ÷1,000,000
nano (n) : 10 -9 or ÷1,000,000,000
pico (p) : 10 -12or ÷1,000,000,000,000

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Q.1- What is the normal range of blood urea?

Answer- Blood urea in normal health ranges between 20-40 mg/dl.

 Q.2- How is urea formed in the body?

 Answer- Urea is the end product of protein metabolism. It is formed in the liver from ammonia and carbon dioxide;both of them are considered to be waste products of the body. Ammonia is highly toxic, it is detoxified through conversion to urea, which is non toxic and water-soluble and is excreted through urine by the kidneys. In higher concentration urea is also toxic. Hence formation and excretion of urea is dependent on liver and kidney functions. In liver disorders urea formation is impaired hence blood urea decreases while in disorders of kidney; excretion of urea is impaired resulting in high blood urea levels. Hence blood urea level can be considered a predictor of hepatic or renal functional status.

 Q. 3-What is the difference between Blood Urea and Blood Urea Nitrogen (BUN)?

 Answer-Blood urea is sometimes expressed in terms of nitrogen. Such expression is very common in clinical practice. Molecular weight of urea is 60 and each gram mol of urea contains 28 gram of nitrogen. Thus a serum concentration of 28 mg/dl of BUN is equivalent to 60mg/dl of urea. Any value of BUN can be converted to urea by multiplying the figure by 2.14.

Blood urea= BUN x2.14

For example if BUN for a patient is 30 mg/dl,

Blood urea = 30x 2.14=64.20mg/dl.

 Q.4- What is meant by NPN (Non protein Nitrogen)?

Answer- NPN includes Urea, uric acid and Creatinine. The major route for excretion of these compounds is urine. In kidney dysfunction the levels of these compounds are elevated in plasma. Of the three, creatinine estimation is the most specific index of renal function. Urea level depends on the protein intake and protein catabolism and also on age of an individual. It is also affected by volume of plasma.

 Other minor components of NPN are urobilinogen, Indican, ammonia and amino acids.

 Q.5- What is Uremia? What are the conditions causing uremia?

 Answer- The clinical state with blood urea higher than normal is called Uremia. The conditions causing high urea level are as follows-

 A) Physiological – Advancing age and high protein diet.

 B) Pathological-Classified in to three categories-

 a) Pre Renal– Primarily there is reduction of plasma volume, leading to lowering of blood pressure with consequent reduction of renal blood flow and Glomerular filtration rate(GFR). This leads to urea retention. Reduced plasma volume is seen in

 i) Dehydration- as in severe vomiting, intestinal obstruction, pyloric stenosis, severe prolonged diarrhea, fluid depletion associated with Diabetic keto acidosis, shock, burns and hemorrhages.

ii) Increased  protein catabolism- as in High fever, toxic state, metabolic response to injury, hemorrhage in to the alimentary tract, digestion of protein passing along the intestine and later deamination of amino acids.

 b) Renal Uremia-In renal diseases there is reduction of GFR resulting in urea retention. Acute renal failure, acute glomerulonephritis, Malignant Hypertension and Pyelonephritis, all of them produce increase in the blood urea levels.

 c) Post Renal Uremia– Obstruction to the out flow of urine after it leaves the kidney leads to back pressure on the renal pelvis,diminished glomerular filtration of urea with consequent increase in the blood urea level.

Renal stone, stricture,Enlargement of prostate and malignant tumors may produce post renal uremia.

 Q.6- Under what conditions blood  urea is lower than normal?

 Answer- Blood urea level is low in liver diseases (Since urea is synthesized in liver),in pregnancy, growing period ,recovery from illness and in tissue healing, since  in all these later conditions, there is positive nitrogen balance and amino groups of the amino acids are not available for urea formation. In disorders of urea cycle also there is impaired urea formation, hence blood urea is low.

 Q.7-What is nitrogen balance?

 Answer- A normal healthy adult is said to be in nitrogen balance, because the dietary intake equals the loss through urine, feces and skin. When the excretion exceeds intake, it is negative nitrogen balance. When the intake exceeds excretion, it is said to be positive nitrogen balance.

 a)Positive nitrogen balance is observed in- Pregnancy- due to enlargement of uterus and fetal growth, growing period, during convalescence(recovery from illness or surgery due to active regeneration of tissues) and under the influence of Growth hormone, Insulin and androgens.

 b) Negative nitrogen Balance is observed in- acute illness,surgery, trauma, burns, malignancy, diabetes mellitus and chronic, debilitating diseases and in protein energy malnutrition. Corticosteroids cause a negative nitrogen balance.

 Q.8- Define clearance, what is its significance?

 Answer- Clearance is defined as the volume of plasma completely cleared of a substance per unit time and is expressed in ml/minute. In other words it is the ml of plasma which contains the amount of a substance excreted by the kidney in one minute. For example Urea clearance of 75 ml/minute means, 75 ml of plasma gets completely cleared of urea in one minute by excretion of urea through urine by kidney.

 Q.9- If a person has blood urea as 54 mg/dl and urea clearance 25 ml/minute, comment on the functional status of the kidney.

 Answer- Urea clearance is affected by volume of urine excreted per minute. If the excretion of urine is at the rate of 2 ml or more the clearance is designated as Maximum urea clearance (Cm) and if the rate of excretion is less than 2ml/minute, the clearance is designated as Standard urea clearance (Cs). The average normal values are 75 ml/minute for Maximum urea clearance (Cm)and 54 ml/minute for Standard urea clearance (Cs).

 Now in the given patient, the urea clearance is 25 ml/minute means only 25 ml of plasma is getting completely cleared of urea per minute, this is very low in comparison to the normal clearance values, that means kidney is failing to excrete urea in urine , hence it is  getting accumulated in  blood to cause high urea level, as is apparent from the high level of 54 mg/dl in this patient. So overall interpretation is that there is functional impairment of the kidney, this deficit seems to be a mild to  moderate functional deficit.

 Q.10- Calculate the urea clearance for the following three patients from the values given below and comment on the functional status of the kidney.

Patient                  Blood urea  Urinary Urea         Volume of urine excreted/ minute   

                               (mg/dl)                 (G/L)                     (ml/24 hours)

1)                                  40               12                         1500

2)                                  80               10                         1000

3)                                  64               18                         1200


Patient 1)

Blood urea (P)          =40 mg/dl

Urinary urea (U)      = 12 G/L =12,000mg/L = 1200 mg/dl

( Value should be in mg/dl)

Volume of urine excreted/day     – 1500 ml/24 hours

Convert it into ml/minute=          1500/24×60 ml/minute

 Or 1500/1440= 1.04ml/minute

 Since V is less than 2 ml/minute, hence standard urea clearance is to be considered

Cs    =                  U√V



=          1200 x√1.04



    = 30.6 ml/min (approx)

 The value of urea clearance is expressed in terms of percentage of the normal (Which is 54 ml/min– normal Cs)

 Hence Percentage urea clearance =    30.6 x 100



                                                   =    56.6 %

 It is mild renal functional deficit (The value between 50-70% of the normal is considered mild renal deficit)

 (Calculate for the other two patients- self exercise)

 Q.11- If blood urea is high and serum Creatinine is normal what is the probable diagnosis?

 Answer- In renal failure both blood urea and creatinine should be high, but in the given case normal creatinine rules out the possibility of renal failure. High blood urea can be due to physiological factors like high protein diet or advancing age or it could be pre renal uremia. In post renal uremia if the obstruction is not relieved renal damage does occur and in that case creatinine also rises after a time.

 Q.12- In a patient suffering from cirrhosis of liver, what results  do you expect from the following biochemical parameters-

i) Blood Urea   ii) Blood Ammonia iii) CSF Glutamine   iv)Urinary Urea

Answer- In  Cirrhosis of liver, blood urea is low ( since urea is formed in the liver), blood ammonia is high(since ammonia is not getting converted to urea), CSF glutamine is high (Ammonia couples with glutamate to form Glutamine) and urinary urea is low since blood urea is also low.

 Q.13-  A known diabetic patient has been brought to emergency with nausea, vomiting, extreme weakness, puffiness of face and mental confusion. His biochemical profile is as follows

Blood urea-80 mg/dl

Serum Creatinine- 3.4 mg/dl

Random blood glucose- 234 mg/dl

 Serum total proteins- 4.5 G/dl

Serum Total Cholesterol- 335mg/dl

Urine analysis – Proteins and glucose are present

 Make a probable diagnosis.

Answer- The patient is suffering from Diabetic nephropathy which has progressed to renal failure, High blood glucose,cholesterol, low  serum total proteins,proteinuria and glycosuria go in favor of diagnosis of diabetic nephropathy and high blood urea and creatinine signify Renal failure.

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Q.1- What is the normal range of serum total proteins? What is A: G ratio?

 Answer- The total plasma protein concentration normally ranges between 6.0 to 8.0 G/dl. Albumin ranges between 3.5 to 5.5 G/dl and Globulins range between 2.5-3.5G/dl . The ratio of albumin to Globulin concentration is called A: G ratio and it varies from 1.5-2.5:1.

 Q.2 – What are the causes of hypoproteinemia?

 Answer- Hypoproteinemia,i.e., a generalized decrease in plasma proteins occurs in –

 (1) Liver diseases,because hepatic protein synthesis is depressed (Mainly albumin is low)

 (2) Renal disorders like the Nephrotic syndrome, in which glomerular membrane permeability increases markedly.

 (3) Malnutrition and starvation- reduced dietary availability

 (4) Protein-losing enteropathy- excessive loss through intestine

 (5) Wide spread burns- loss from skin

 (6) Severe hemorrhage- increased protein catabolism

 (7) Defective digestion or malabsorption as in carcinoma of stomach or pancreas, peptic ulcer and steatorrhea.

 (8) Fever- increased protein catabolism

 (9) Pregnancy-hemodilution and increased requirement

 (10) Acute infections in-untreated diabetes mellitus and hyperthyroidism

 Q.3- What are the causes of hyperproteinemia?

 Answer- Increase in plasma proteins are seen in

 (1) Acute inflammatory states. Several plasma proteins increase sharply during any acute inflammation. These are called acute phase proteins. They include C-reactive proteins (CRP), so-called because it reacts with C- polysaccharide of pneumococci.

(2) Multiple myeloma. In this condition, the plasma cells secrete large amounts of immunoglobulin resulting in hypergammaglobulinemia

(3) Dehydration –due to hemoconcentration

 (4) Chronic infections

 (5) Leukemias

 (6) Lymphomas

 (7) Tuberculosis

 (8) Kala Azar

 (9) HIV infection

 Increase in plasma protein concentration is generally due to an increase in total globulins (Gamma globulins) and the concentration of albumin remains the same or decreases marginally. A decrease in total protein concentration is due to fall in albumin and some times globulins. In these conditions A:G ratio changes due to either reduction of Albumin or increase of Globulins.

 Q.4-What is the effect of plasma volume on total protein concentration?

 Answer- Decrease in the volume of plasma water –Hemoconcentration, is reflected as relative hyperproteinemia and the concentration of all individual plasma proteins are increased to the same degree.

Hyperproteinemia is observed in dehydration due to-inadequate water intake or due to excessive water loss as in –Diarrhea, vomiting, Addison’s disease, diabetic ketoacidosis and diabetes Insipidus.

 Hemodilution occurs with water intoxication and salt retention syndromes, during massive intravenous infusions and physiologically when a recumbent position is achieved. A recumbent position decreases plasma protein concentration by 03-0.5 G/dl. In hemodilution individual plasma proteins are decreased by the same degree.

 Q.5- What are the functions of plasma proteins?

Answer- Functions of the plasma proteins include:

  • Intravascular osmotic effect for maintaining fluid and electrolyte balance
  • Contribute to the viscosity of the plasma
  • Transport of insoluble substances around the body by allowing them to bind to protein molecules. Blood plasma proteins like albumin functions as carrier proteins that help in the translocation of different biomolecules in body.
  • Protein reserve for the body
  • Clotting of blood
  • Inflammatory response
  • Protection from infection- The gamma globulins function as antibodies
  • Maintenance of the acid-base balance
  • Blood plasma contains the protease inhibitor enzymes like alpha-1 antitrypsin that help in the reduced proteolytic activity in the blood.

Q.6-What is the cause of hyperproteinemia in cirrhosis of liver or other liver disorders?

 Answer-Although the concentration of serum Albumin is reduced in severe liver diseases (Since albumin is synthesized in liver), that of globulins is usually increased (For compensation as they are synthesized in spleen and bone marrow) so that the total plasma protein concentration is rarely low and is often high.

 Q.7- What is the cause of edema in hypoproteinemia?

 Answer- Plasma proteins contribute to plasma colloidal osmotic pressure, counteracting the effects of capillary blood pressure, which tends to force water in to tissue spaces. The lowered plasma protein concentration causes a decrease in the plasma osmotic pressure and water is forced in to tissue spaces resulting in edema. Edema is probable when the albumin concentration falls below 2G/dl.

 Q.8- State your diagnosis from the following Blood and Urinary findings

Biochemical Parameter   Patient 1                         Patient 2

Blood                             Hypoproteinemia                  Hypoproteinemia

Urine                              Proteinuria                                 Normal Urine                          

Answer-Patient 1- With Hypoproteinemiaand proteinuria is most probably suffering from Nephrotic syndrome, while Patient 2 with hypoproteinemia without proteinuria might be suffering from Kwashiorkor.

 Q.9- What is Nephrotic syndrome? What are the laboratory findings in this syndrome?

Answer-Nephrotic syndrome(NS), also known as Nephrosis, is defined by the presence of nephrotic-range proteinuria, edema, hyperlipidemia, and hypoalbuminemia. 

In a healthy individual, less than 0.1% of plasma albumin may traverse the glomerular filtration barrier.Nephrotic-range proteinuria in adults is characterized by protein excretion of 3.5 g or more per day.

 Biochemical basis

1) An increase in glomerular permeability leads to albuminuria and eventually to hypoalbuminemia. In turn, hypoalbuminemia lowers the plasma colloid osmotic pressure, causing greater trans capillary filtration of water throughout the body and thus the development of edema.

2) In the nephrotic syndrome, levels of serum lipids are usually elevated. This can occur via (i) hypoproteinemia that stimulates protein,including lipoprotein, synthesis by the liver, and (ii) diminution of lipid catabolism caused by reduced plasma levels of lipoprotein lipase.

 Laboratory findings

1) Urinalysis- Proteinuria is observed

2) Blood chemistries

i) Serum total and differential protein- A decreased serum albumin (< 3 g/dL) and total serum protein less than 6 g/dL may be observed.

ii)-Lipid Profile-Hyperlipidemia occurs in over 50% of those with early nephrotic syndrome. The degree of hyperlipidemia increases with the degree of protein loss.

iii) ESR and Serum Fibrinogen levels-Patients can have an elevated erythrocyte sedimentation rate as a result of alterations in some plasma components such as increased levels of fibrinogen.

iv) Renal Function Tests- BUN (Blood urea nitrogen) and serum creatinine rise in the setting in of renal failure.

v) Renal biopsy- For confirmation of diagnosis

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Q.1-What is the level of Fasting blood glucose in normal health?

Answer- Fasting blood glucose (FPG) is measured after an overnight fast of 10 hrs. (1) FPG 7.0mmol/L (126 mg/dL) warrants the diagnosis of Diabetes Mellitus (DM).

Q.2- What is Random blood Glucose? What is its significance?

Answer-Random is defined as without regard to time since the last meal. Random Blood Glucose (RBG) measurement is required only during emergency. The current criteria for the diagnosis of DM emphasize that the FPG is the most reliable and convenient test for identifying DM in asymptomatic individuals. A random plasma glucose concentration >11.1 mmol/L (200 mg/dL) accompanied by classic symptoms of DM (polyuria, polydipsia, weight loss) is sufficient for the diagnosis of DM.

Q.3- What is the difference between Post load blood glucose and postprandial blood glucose?

Answer-Post load blood glucose means blood glucose level 2 hours after a glucose load as it is done in Oral glucose tolerance test. Post prandial means blood glucose 2 hours after a normal meal.

Q.4- Why is plasma sample preferred over serum sample for estimation of blood glucose?

Answer- Plasma has the advantage over serum since the blood can be immediately added to a mixture of anticoagulant and preservative and can be processed immediately. If serum is used, there is a delay in processing the sample since it takes nearly 20-30minutes for the serum to get separated from the blood and in that span some glucose is metabolized by anaerobic glycolysis and falsely low levels are obtained. Hence on practical grounds plasma is a preferred choice over serum.

Q.5- What is the purpose of adding preservative to the blood sample(collected for estimation of blood glucose) before processing?

Answer- Red blood cells possess glycolytic enzymes; hence glucose disappears fairy rapidly due to glycolysis from the whole blood. Glucose loss takes place approximately at a rate of 0-5 mmol/L/hour and it completely disappears within 6 hours. So blood is collected in to an anti glycolytic and anticoagulant mixture to get the accurate results.A fluoride and oxalate mixture (20 mg/5ml) in the ratio of 1:3 is used to prevent glucose loss. Sodium fluoride acts as a preservative since by acting as an inhibitor of Enolase enzyme, it inhibits the conversion of 2-phospho glycerate to phosphoenol pyruvate. Hence glycolysis is inhibited and glucose loss of the sample is prevented. Oxalate (Potassium oxalate) acts as an anticoagulant.

Q.6- What is the reason that for a given volume, the level of whole blood glucose is always lower than that of plasma glucose?

Answer-Water content of plasma is more than whole blood for a given volume and it has more dissolved glucose. Water content of red blood cells is 73% as compared to 93% of plasma.To convert from whole-blood glucose, multiplication by 1.15 has been shown to generally give the serum/plasma level.

Q.7- What are the conditions causing hyperglycemia?

Answer-The common conditions causing hyperglycemia are as follows-

i) Diabetes Mellitus

ii) Insulin Resistance

iii) Diabetes prone states– Gestational Diabetes, Impaired fasting glycemia, Impaired glucose tolerance    

iv) Diseases of the exocrine pancreas– cystic fibrosis, hemachromatosis, fibrocalculous pancreatopathy, pancreatitis, pancreatectomy, neoplasia,   

v) Endocrinopathies—Acromegaly, Cushing’s syndrome,glucagonoma, pheochromocytoma, hyperthyroidism, somatostatinoma, aldosteronomas

vi) Drug- or chemical-induced—Nicotinic acid, glucocorticoids,thyroid hormone, β -adrenergic agonists, thiazides,phenytoin etc.

vii) Genetic syndromes causing hyperglycemia—Down’s syndrome,Klinefelter’s syndrome, Turner’s syndrome, Wolfram’s syndrome, Friedreich’s ataxia, Huntington’s chorea, Laurence-Moon-Biedl syndrome, myotonic dystrophy,Porphyria, Prader-Willi syndrome

viii) Stress

ix) Chronic infections

Q.8- What are the common causes of Hypoglycemia?

Answer-Hypoglycemia is a laboratory ‘diagnosis’ which is usually considered a blood glucose level below 60 mg/dL. Symptoms begin at plasma glucose levels in the range of 60 mg/dL and impairment of brain function  occurs at approximately 50 mg/dL. The common causes of hypoglycemia are as follows-

1) Physiological- Prolonged fasting or starvation

2) Pathological-

i) Drugs -especially insulin, sulfonylureas, ethanol sometimes quinine, rarely Salicylates,sulfonamides, others.

ii) Critical illnesses– Hepatic, renal, cardiac failure or sepsis

iii) Hormone deficiencies- Cortisol, growth hormone, or both ,Glucagon and epinephrine (in insulin-deficient diabetes)

iv) Endogenous hyperinsulinism


vi) Insulin secretagogue (sulfonylurea, other)

vii) Ectopic insulin secretion

viii) Alimentary (Postgastrectomy)

ix) Hereditary fructose intolerance, Galactosemia

x) Glycogen Storage diseases

Q.9-What are the indications for performing glucose tolerance test?

Answer-If the fasting plasma glucose level is 126 mg/dL or higher on more than one occasion, further evaluation of the patient with a glucose challenge is unnecessary. However, when fasting plasma glucose is less than 126 mg/dL in suspected cases, a standardized oral glucose tolerance test may be done.

75 g of glucose dissolved in 300mL of water is given after an overnight fast to a person who has been receiving at least 150–200 g of carbohydrate daily for 3 days before the test. The data is interpreted as follows-

Normal Glucose Tolerance

Impaired Glucose Tolerance

Diabetes Mellitus

Fasting plasma glucose (mg/dL)

< 110



Two hours after

glucose load

< 140



*The Diabetes Expert Committee criteria for evaluating the standard oral glucose tolerance test.

For proper evaluation of the test, the subjects should be normally active and free from acute illness.Medications that may impair glucose tolerance include diuretics, contraceptive drugs, glucocorticoids, niacin, and phenytoin should be avoided on that day.

The common indications for Glucose Tolerance Test are as-

a) Family history of diabetes mellitus

 b) Typical Symptoms but normal biochemical profile

 c) Abnormal biochemical profile but no symptoms

 d) History of gestational diabetes mellitus

 e) History of large for size babies at the time of delivery

Q.10- What is the advantage of estimating Glycated hemoglobin over fasting blood glucose?

Answer- Glycated hemoglobin (HbA1c)comprises 4–6% of total hemoglobin A1.The hemoglobin A1c fraction is abnormally elevated in diabetic persons with chronic hyperglycemia.Since glycohemoglobins circulate within red blood cells whose life span lasts up to 120 days, they generally reflect the state of glycemia over the preceding 8–12 weeks, thereby providing an improved method of assessing diabetic control. Fasting blood glucose on the other hand is affected by many factors like meals, stress, infections and drugs,hence a true glycemic status can not be ascertained.

Q.11- What is meant by self monitoring of blood glucose?

Answer-Capillary blood glucose measurements performed by patients themselves, as outpatients, are extremely useful. In type 1 patients in whom”tight” metabolic control is attempted, they are indispensable. There are several paper strip (glucose oxidase, glucose dehydrogenase, or hexokinase methods for measuring glucose on capillary blood samples. A reflectance photometer or an amperometric system is then used to measure the reaction that takes place on the reagent strip.

Q.12- Apart from estimation of blood glucose, what are the other investigations to be carried out for the diagnosis of diabetes mellitus?

Answer- The laboratory investigations for the diagnosis of Diabetes mellitus are as follows-

1) Urine Analysis– shows Glucosuria, Ketonuria and Microalbuminuria

2) Blood Biochemistry includes Blood glucose estimation and apart from that the other tests are-

i) Glycated hemoglobin

ii) Serum fructosamine

iii) Lipid profile

iv) Plasma insulin

v) c-peptide

vi) Islet cell antibodies

3) Additional Tests– In addition to the standard laboratory evaluation, the patient should be screened for DM-associated conditions (e.g., kidney, liver and thyroid dysfunction).

Q.13- If the blood glucose is lower than 60 mg/dL, as in case of Starvation, what kind of abnormal compounds will be excreted in urine in excess?

Answer- Urine will have ketone bodies in excess, but glucose will not be there. In states of glucose deprivation, fatty acids are largely oxidized and the product acetyl coA is channeled towards ketogenesis. Ketone bodies are also used as alternative fuel molecules.

Q.14- What is the most important cause of hypoglycemia in Diabetes Mellitus?

Answer- One of the therapeutic goals of diabetes is to decrease blood glucose levels in an effort to minimize the development of the long term complications of the disease. However, appropriate dosage is difficult to achieve in all patients, and hypoglycemia caused by excess insulin is the most common complication of insulin therapy, occurring in more than 90 % of the patients. The frequency of hypoglycemic episodes, coma and seizures is particularly high with intensive treatment regimens designed to achieve tight control of blood glucose. In normal individuals, hypoglycemia triggers a compensatory secretion of counterregulatory hormones, most notably glucagon and epinephrine, which promote hepatic production of glucose. However patients with type 1 diabetes also develop a deficiency of glucagon secretion.This defect occurs early in the disease and is almost universally present four years after diagnosis. These patients thus rely on epinephrine secretion to prevent severe hypoglycemia. However as the disease progresses, type 1 diabetes patients show diabetic autonomic neuropathy and impaired ability to secrete epinephrine in response to hypoglycemia. The combined deficiency of glucagon and epinephrine secretion creates a condition sometimes called “Hypoglycemia unawareness”. Thus patients with long standing diabetes are particularly vulnerable to hypoglycemia. Hypoglycemia can also be caused by strenuous exercise. Exercise promotes glucose uptake into muscles and decreases the need for exogenous insulin. Patients should therefore check blood glucose levels before or after intensive exercise to prevent or abort hypoglycemia.

Q.15- What is meant by “Honeymoon period” in diabetes mellitus?

Answer- Around ¼ of all patients who get type 1 diabetes develop what is known as a ‘honeymoonperiod within days or weeks of the onset of treatment. It is as if the patient has gone into remission and it can be confusing for the patient as it would appear that the condition has corrected itself. Some patients actually require no insulin during this phase and this may last for weeks or months. It is usually best to keep treating with insulin even if the requirements are negligible, to avoid possible insulin allergy upon re-exposure and also to maintain a treatment regimen and not give false hope to the patient.

Q.16- What is the significance of non diabetic Glycosuria?

Answer- Non diabetic Glycosuria(renal Glycosuria) is a benign asymptomatic condition wherein glucose appears in the urine despite a normal amount of glucose in the blood, either basally or during a glucose tolerance test. Its cause may vary from an autosomally transmitted genetic disorder to one associated with dysfunction of the proximal renal tubule (Fanconi’s syndrome, chronic renal failure), or it may merely be a consequence of the increased load of glucose presented to the tubules by the elevated glomerular filtration rate during pregnancy. As many as 50% of pregnant women normally have demonstrable sugar in the urine, especially during the third and fourth months. This sugar is practically always glucose except during the late weeks of pregnancy, when lactose may be present.

Q.17- What is the significance of Microalbuminuria?

Answer-Microalbuminuria- may be defined as an albumin excretion rate intermediate between normality (2.5-25 mg/day) and macroalbuminuria (250 mg/day). The small increase in urinary albumin excretion is not detected by simple albumin stick tests and requires confirmation by careful quantization in a 24 hr. urine specimen. The importance of micro- albuminuria in the diabetic patient is that it is a signal of early reversible renal damage.

Unlike type 1 diabetes mellitus,in which microalbuminuria is a good indicator of early kidney damage,microalbuminuria is a common finding (even at diagnosis) in type 2 diabetes mellitus and is a risk factor for macrovascular (especially coronary heart) disease. It is a weaker predictor for future kidney disease in type 2 diabetes mellitus.

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Q.1- What are the components of Benedict’s qualitative reagent? What is the significance of each of the components?

Answer-Reducing sugars are usually detected by Benedict’s reagent,which contains copper sulphate, sodium citrate and sodium carbonate. Sodium carbonate makes the medium alkaline. Copper sulphate furnishes Cu2+ ions and sodium citrate prevents the precipitation of cupric ions as cupric hydroxide by forming a loosely bound cupric- sodium –citrate complex which on dissociation gives a continuous supply of cupric ions.

Reducing sugars  under alkaline conditions tautomerise and form enediols. Enediols are powerful reducing agents. They can reduce cupric ions to cuprous form which is the basis for Benedict’s reaction.

The alkaline conditions of this test also cause isomeric transformation of ketoses to aldoses, resulting in all mono-saccharides and most disaccharides to respond to this test by reducing the blue Cu2+ ion to cuprous oxide (Cu2O), a brick red-orange precipitate. This solution has been used in clinical laboratories for testing urine.

Q.2- Why is Benedict’s test considered a semi quantitative test?

Answer-In Benedict’s test, when fixed volume of urine (8 drops) and Benedict’s reagent(5 ml) are boiled for 2 minutes, different colored precipitated are obtained. The precipitate color can vary between green, yellow, orange or red. The color of the precipitate signifies the rough estimate of the amount of reducing sugar present in the given sample.The interpretations are made as follows-

Color of the precipitate       Amount of reducing sugar present in the given sample

Green color of solution               Up to 0.5 g%   (+)

Green precipitate                         0.5- 1.0 g%  (++)

Yellow precipitate                       1.0-1.5g%   (+++)

Orange precipitate                      1.5-2.0 g%   (++++)

Brick red precipitate                   more than 2 g%

Due to the fact that an approximate concentration of the reducing sugar can be determined by this test, it is called a semi-quantitative test.

Q.3- What is the alternative test for identification of reducing sugars?

Answer- Fehling test is an alternative to Benedict’s test. It differs from Benedict’s test in that it contains sodium potassium Tartrate in place of Sodium citrate and potassium hydroxide as an alkali in place of sodium carbonate in Benedict’s reagent. It is not a preferred test over Benedict’s test since the strong alkali present causes caramalizaton of the sugars; hence it is less sensitive than Benedict’s reagent.

Q.4 – Can Benedict’s test be given positive by non carbohydrate substances?

Answer- Any substance which is reducing in nature would give this test positive. For example- Ascorbates, Urates, Salicylates and Glucuronates give this test as positive, since they are all reducing in nature.

Q.5- Benedict’s and Barfoed’s both are reduction tests, they  why are they performed separately ?

Answer-Barfoed’s solution contains cupric ions in an acidic medium as compared to Benedict’s test where the medium is alkaline. The milder conditions allow oxidation of mono-saccharides but do not allow oxidation of disaccharides. If the time of heating is carefully controlled, disaccharides do not react while reducing mono-saccharides give the positive result (red Cu2O precipitate).Thus this test is carried  out to confirm the presence of mono-saccharides or to rule out the presence of disaccharides.

Q.6- If a sugar gives both Benedicts and Barfoed’s tests negative, which test should be performed to confirm the identity of the sugar?

Answer- Both negative tests signify that the sugar is a non reducing disaccharide and  hence it could be Sucrose. Seliwanoff and Inversion tests should be carried out to confirm the presence of sucrose

Q.7- If Benedict test is positive but Barfoed’s test is negative for an unknown sugar solution, which test should be carried out to confirm the identity of the sugar?

Answer- Benedict’s positive and Barfoed’s negative shows that the given sugar is reducing in nature but is not a monosaccharide, it could be a disaccharide. Osazone test should be carried out to confirm the identity of the reducing sugar. The characteristic shape of the osazone crystals would signify the identity of the reducing disaccharide.It could  be Maltose or Lactose. Maltose forms sun flower shaped crystals  while hedge hog shaped crystals are formed by Lactose.

Q.8- An unknown carbohydrate reacted positively with Benedict’s reagent formed a red precipitate with Barfoed’s reagent within 3 minutes and turned blue-green when mixed with Bial’s reagent. What carbohydrate it could be?

Answer- From the observations, it is clear that the given carbohydrate is a reducing monosaccharide (Positive Benedict’sand Barfoed’s tests) and since the Bial’s test is a confirmatory test for  pentoses so the given carbohydrate could be a pentose.

Q.9- A urine sample gives a positive reaction with Benedict’s and Barfoed’s reagents and forms rhombic plate shaped osazone crystals, what can be the nature of the sugar and what is that clinical state called when the above said sugar is excessively excreted in urine?

Answer- Positive reaction with Benedict’s and Barfoed’s reagent signifies  that the given carbohydrate is a reducing monosaccharide and rhombic crystals are given by Galactose. Hence it is Galactose and the clinical state is called Galactosuria which is seen in Galactosemia, a congenital disorder of Galactose metabolism.

Q.10- Urine sample of a patient suffering from hereditary fructose intolerance is to be analysed. Fructose is excessively excreted in urine in this condition. Name the conventional tests which should be carried out for its detection?

Answer- Fructose is a ketohexose,reducing in nature, hence Seliwanoff test and Benedict’s should be carried out to confirm the presence of fructose. Seliwanoff test is given positive by Keto hexoses in the free or bound form.

Q.11- What is the precaution while performing Barfoed’s test?

Answer- This test is used to distinguish reducing monosaccharides from reducing disaccharides by controlling p H and time of heating. Monosaccharides react fast whereas the reaction with disaccharides is slow. However if the heating is prolonged disaccharides can be hydrolysed by acid and the resultant monosaccharides give the test positive. Hence false positive result can be obtained upon prolonged heating of a disaccharide solution.

Q.12- What will be the result of Benedict test with the hydrolytic product of Starch?

Answer- The hydrolytic product of starch is maltose and if further hydrolyzed it is Glucose, both are reducing sugars, hence Benedict’s test will be positive in this situation.

Q.13- Sucrose is non reducing in nature but Benedict test shows positive reaction with the hydrolytic products,what is the reason?

Answer- Sucrose is a disaccharide having Glucose and Fructose linked together by α1->2 linkage. Both the functional groups of the components sugars are involved in the linkage, hence no free functional group is there to cause reduction, but upon hydrolysis the products  (glucose and fructose ) have free functional groups (Anomeric carbons) to carry out the process of reduction. Hence Benedict’s test is given positive by hydrolytic products.

Q.14- Raffinose is a trisaccharide, what is expected from Barfoed’s test ?

Answer- It will be negative; it is given positive only by monosaccharides.

Q.15- Which test should be carried out to confirm the presence of a glycoprotein present in a given solution?

Answer- Biuret test for the protein part and Molisch test for the carbohydrate part of the glycoprotein should be carried out for confirmation.

Q.16- Which test out of the followings will be negative with products of hydrolysis of starch Iodine,Benedict, Barfoed’s?

Answer- Iodine test will be negative,since iodine forms a coordinate complex between the helically coiled polysaccharide chains and iodine gets centrally located within the helix due to absorption. The color obtained with iodine depends upon the chain length of the polysaccharide (branched or unbranched) required for complex formation. Amylose (Linear chain component) gives a deep blue color; Amylopectin (Branched chain component) gives a purple color, while glycogen gives a reddish-brown color.The hydrolytic products (mono-saccharides) do not react with iodine due to the reasons mentioned above, hence test should be negative.

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Q.1-What is the difference between salting in and salting out?

Answer- A protein’s multiple acid base groups make its solubility properties dependent on the concentration of salts, the polarity of the solvent, the p H and the temperature. The solubility of a protein in aqueous solution is a sensitive function of the concentration of the dissolved salts. The solubility of a protein increases upon addition of low concentration of salt. This phenomenon is called salting in. The explanation for salting in is that as the salt concentration of the protein solution increases additional ions more effectively shield the protein molecules and thus promote protein water interactions, (protein -protein interactions are minimized as the like charges repel each other) resulting in increase in solubility of the proteins. 
At high concentration of salt, the solubility of the protein decreases. This effect is known as salting out and is primarily a result of the competition between added salt ions and protein molecules for the given solvent. At high salt concentration many of the added ions are solvated, so that the solvent available becomes insufficient to dissolve protein. Hence protein – protein interactions become stronger than protein water interactions and the protein is precipitate out. Salting out is the basis of one of the most common protein purification procedures. By adjusting the amount of salt concentration in a solution containing a mixture of proteins, the different proteins can be separated out.

Q.2- What is the difference between half saturation and full saturation?
Answer- Half saturation and full saturation signify the amount of salt required for precipitation of a protein. The amount of salt required for precipitation depends upon the molecular weight of a protein. There is generally an inverse relationship between amount of salt required and the molecular weight of the protein. The proteins with a large molecular weight need less salt. Globulins are precipitated by half saturation with ammonium sulphate, where as albumin with a low molecular weight is precipitated at full saturation. Fibrinogen is best precipitated by 1/5 th saturation with ammonium sulphate. The commonly used salt solutions are 
!) Ammonium sulphate  
ii) A mixture of sodium sulphate and sodium sulphite.
Q.3- Why is alcohol used as an antiseptic?

Answer- Water miscible organic solvents, such as Acetone and Ethanol are generally good protein precipitants because their lower dielectric constants lower the solvating power of their aqueous solutions for dissolved proteins. Hence the protein-protein interactions become stronger than protein solvent interactions and the proteins are precipitated out.

Alcohol rubbed over the injured site acts by precipitating membrane proteins of the micro-organisms and hence the multiplication of the micro-organisms is prevented at the site of tissue injury. Although alcohol cannot coagulate every single cell, it functions well to inhibit the growth and reproduction of many microorganisms, including bacteria, fungi, protozoa, and viruses. Human skin cells are more resistant to alcoholic coagulation than most microorganisms. This is why the human skin doesn’t coagulate if it comes into contact with alcohol.

Q.4- What is iso -electric p H precipitation?

Answer- Proteins generally bear numerous ionizable groups which have a variety of pK’s. At a p H characteristic of each protein, the positive charges on the molecule exactly balance the negative charges. At this Isoelectric point (p I)of the protein there is no net charge on it and the protein remains immobile in the electric field .The solubility of a protein is near a minimum near its p I. This phenomenon is the basis of a protein purification procedure known as isoelectric precipitation in which the pH of a protein mixture is adjusted to the pI of the protein to be isolated so as to selectively minimize its solubility. The precipitated protein is separated out from a mixture of proteins. Each protein has a specific Isoelectric pH (p I).

Q,5- What is the biochemical basis of curdling of milk ?

Answer- Curdling of milk is based on the principle of isoelectric pH precipitation. Addition of little amount of curd (Containing fermentation product of lactose in the form of lactic acid), lowers the pH of milk bringing it close to Isoelectric pH of Casein (milk protein). At this pH casein gets precipitated causing curdling of milk.

In normal conditions milk has a pH of about 6.5-6.7 and at this pH value the casein has a negative charge and there fore the casein micelles are relatively soluble, because they repel each others. Upon addition of curd ( lactic acid) casein acquires its Isoelectric point at 4.6 pH, that is at this pH value it has a quantity of positive charges equal to the quantity of negative charges and the positive part of each “micelle” is attracted by the negative part of the others, causing the formation of ionic bonds among the “micelle” working against the dipole-dipole bonds with water,i.e. protein-protein interactions get stronger than protein water interactions, so that the protein precipitates producing curdling of milk

Q.6- What is the benefit of using protein free filtrate for quantitative estimations?

Answer- Proteins present in the sample produce turbidity. The turbid solutions absorb more light-giving heightened optical density values. Since absorbance is proportional to the concentration of unknown substance. Hence the turbid solutions give falsely high levels of the substance present in the solution. Thus the proteins are eliminated from the solution before the quantitative procedures for the accuracy of the results.

Q.7- What is the principle of Heat coagulation test? What is the purpose of adding dilute acetic acid to the solution?

Answer-Heat causes denaturation of proteins by increasing the kinetic energy of the molecules and by breaking the weak bonds like electrostatic interactions, hydrogen bonds and hydrophobic interactions. The denatured protein becomes insoluble and precipitates out. Dilute acetic acid provides an acidic medium and brings the pH close to Isoelectric p H . At Isoelectric p H the protein has  a minimum solubility and is thus precipitated.
 In urinalysis, upon heating urine in an alkaline medium the phosphates produce the similar turbidity as observed with proteins but the turbidity disappears upon addition of acid if the phosphates are there but it persists if it was due to proteins.

Q.8-Why is milk or raw egg used as an antidote to heavy metal poisoning?

Answer- Heavy metal salts act to denature proteins in much the same manner as acids and bases. Heavy metal salts usually contain Hg+2, Pb+2,Ag+1 Tl+1, Cd+2 and other metals with high atomic weights. Since salts are ionic they disrupt salt bridges in proteins.The reaction of a heavy metal salt with a protein usually leads to an insoluble metal protein salt.
This reaction is used for its disinfectant properties in external applications. For example AgNO3 is used to prevent gonorrhea infections in the eyes of new-born infants. Silver nitrate is also used in the treatment of nose and throat infections, as well as to cauterize wounds.
Mercury salts administered as Mercurochrome or Merthiolate have similar properties in preventing infections in wounds.
This same reaction is used in reverse in cases of acute heavy metal poisoning. In such a situation, a person may have swallowed a significant quantity of a heavy metal salt. As an antidote, a protein such as milk or egg whites may be administered to precipitate the poisonous salt. Then an emetic is given to induce vomiting so that the precipitated metal protein is discharged from the body.

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Q.1- What are the conditions in which urine gives a positive reaction with Benedict’s qualitative reagent?

Answer- Benedict’s test gives a positive reaction with all reducing agents, which may be sugars or other agents. In urine, the commonly found reducing sugars are- Glucose, Fructose, Galactose, Lactose and Pentoses. The other non sugar reducing agents are Ascorbates,Urates, Glucuronates, Homogentisic acid, Salicylates etc which also give positive reaction with Benedict’s reagent. Hence positive Benedict’s test should be further explored to know the exact identity of the substance present in urine.

Q.2- A 23- year-old male was denied a job of a sales man on the ground of positive Benedict’s test. Further investigations revealed normal fasting blood glucose and normal oral glucose tolerance test. The specific test for glucose in urine was negative. There was no abnormality detected on physical examination of the patient. There was no family history of diabetes mellitus. What are the conditions in which such like findings can be observed?

Answer- There are two main possibilities-

1) It might be case of renal glycosuria due to transporter deficiency (SGLT-2) required for the reabsorption of Glucose from the filtrate. But such patients generally have polyuria as a result of glucosuria (depending upon the severity of defect) and Polydipsia also due to excessive polyuria. In this patient there are no such symptoms, so it can be ruled out.

2) The second possibility is of Pentosuria, which seems to be the most likely cause in the given patient. Pentosuria goes undiagnosed and it is always a chance finding.There are no symptoms and no treatment is required for this.  Essential Pentosuria occurs due to deficiency of Xylitol dehydrogenase, the enzyme of uronic acid pathway. The confirmation is done by Bial’s test and chromatography.

Q.3- In case of non availability of Benedict’s reagent, is there any alternative test for detection of reducing sugars in urine?

Answer- Fehling’s test can be undertaken,but that is less sensitive than Benedict’s test. The strong alkali (KOH) of Fehling test causes caramalizaton of sugars hence they can be left undetected.

Q.4- Enlist some of the common conditions of Glycosuria

Answer- Glycosuria (presence of detectable amount of any sugar in urine) includes the following:

1) Glucosuria: (presence of detectable amount of glucose in urine).

a) Uncontrolled DM: The concentration of glucose in plasma exceeds renal threshold.

b) Renal Glucosuria: Normal plasma glucose concentration with proximal tubular malfunction leads to decreased renal threshold (gestational diabetes, transported defect and Fanconi’s syndrome).

2).Fructosuria: (Presence of fructose in urine)

a) Alimentary: due to increased fructose intake.

b) Metabolic:deficiency of fructokinase or aldolase B enzyme in the liver.

3). Galactosuria: (Presence of galactose in urine)

a) Alimentary: increased galactose intake

b) Metabolic: deficiency of Galactokinase or galactose -1-phosphate   uridyl transferase in theliver.

4). Lactosuria: (Presence oflactose in urine)

a) Lactation

b) Lactose intolerance

5) Pentosuria :( Presence ofpentose in urine)

a) Alimentary- Excessive ingestion of fruits containing pentoses like Raspberries

b) Metabolic defect- Essential Pentosuria- Deficient Xylitol dehydrogenase

Q.5- What are the conditions causing Ketonuria?

Answer-(Presence of ketones”Acetone, acetoacetic acid and β- Hydroxybutyric acid” in urine)

a) Diabetic ketoacidosis.

b) Glycogen storage diseases

c) Starvation

d) Prolongedvomiting

e) Unbalanced diet:high fat and low carbohydrate diet.

Q.6- What is the possibility if Rothera’s test for ketone bodies in urine is negative but blood ketone level is high ?

Answer- Rothera test is given by acetone and Acetoacetate. Beta hydroxy butyrate, which is the main ketone body of urine does not give this test positive. Hence it is possible despite a good concentration of ketone bodies in urine the test is still negative. The indirect test is performed, by converting beta hydroxy butyrate to Aceto acetate to get the positive result.

Q.7- What are the common causes of proteinuria?

Answer- Normally less than 200 mg protein is excreted in the urine daily; More than this level leads to a condition called “proteinuria”.
Proteinuria is either glomerular or tubular-

1) Glomerular proteinuria is due to increased glomerular permeability leading to filtration of high molecular weight proteins (e.g. glomerulonephritis)

2) Tubular proteinuria occurs as a result of decreased reabsorption with normal glomerular permeability leading to excretion of low molecular weight proteins (e.g. chronic nephritis).

Proteinuria is classified into prerenal,renal and post renal.

1) Pre-renal proteinuria:

a) Dehydration

b) Congestive Heart failure with passive congestion of kidneys

c) Increased intra abdominal pressure as in ascites, tumor

d)  Severe anaemia, fever

e) Bence-Jones protein: This abnormal gamma globulin (light chains only)is synthesized by malignant plasma cells (multiple myeloma). It precipitates at60oC, redissolves on boiling and reprecipitates on cooling to 60oC.

2. Renal proteinuria:

a) Nephrotic syndrome


c)Gestational (in the 3rd trimester of pregnancy)

d) Glomerulonephritis

e) Diabetic nephropathy 

f)  Renalischemia or neoplasia

g) Nephrotoxins(aminoglycosides; gentamicin, streptomycin…etc.)

h) Overflow proteinuria (haemoglobin due to intravascular haemorrhage, microglobulin in leukaemia and lymphoma, and myoglobin in rhabdomyolysis

3)- Post-renal proteinuria

a)  Lower urinary tract infection, tumors or stones

b) Vaginal bleeding

c) Mixing of urine with semen or vaginal secretions (False proteinuria)

Besides these cause functional  proteinuria may be there in conditions like prolonged standing, exposure to cold, severe exercise and in extreme stress.

Q.8- What is Acholuric jaundice?

Answer- Acholuric jaundice means- jaundice without bilirubin in urine Haemolytic jaundice or Prehepatic jaundice is called Acholuric jaundice, since there is Unconjugated hyperbilirubinemia. Unconjugated bilirubin can not appear in urine due to two reasons –

1) Water insoluble,only water soluble substances are excreted in urine.

2) Unconjugated bilirubin is bound to albumin making it a macro molecule; hence can not be filtered.

Thus In haemolytic jaundice test for bilirubin (Fauchet test) is negative for urine but test for urobilinogen(Ehrlich’s test) is positive since urobilinogen is there in excess.Dark color of urine in haemolytic jaundice is due to Urobilinogen and not due to Bilirubin.

Q.9- In which type of jaundice both Bilirubin and urobilinogen are there in urine ?

Answer- In hepatic jaundice both conjugated and unconjugated hyperbilirubinemia is there, hence urine contains both urobilinogen and bilirubin.

Q.10- A patient has reported to Medical OPD with jaundice. There is a previous history of several episodes of biliary colic.  The patient is in acute distress and there is pain and tenderness in the right hypochondrium. His blood and urine samples have been sent for biochemical investigations. What should be the urinary findings for this patient in terms of Bilirubin and urobilinogen?

Answer- The patient has obstructive jaundice due to gall stones. Hence there should be conjugated hyperbilirubinemia. Urine will be positive for bilirubin while negative for urobilinogen.

Q.11- What are the causes of Hematuria and hemoglobinuria?

Answer- 1) Hematuria (Presence of detectable amount of blood in urine):

a) Acute and chronic glomerulonephritis.

b) Local disorders of kidney and genitourinary tract (trauma, cystitis, renal calculi,  tumors etc). 

c) Bleeding disorders(haemophilia). 

d) Malignant hypertension

2) Hemoglobinuria(Presence of hemolysed blood in urine):

a)Hemoglobinopathies (sickle-cell anaemia and Thalassemia)

b) Transfusion reaction (blood incompatibility).

c) Malaria(plasmodium falciparum)

d) Snake bite

Q.12- What is chyluria? 

Answer- Chyluria (Presence of lymph/ fat in urine):

a)The urine acquires a milky appearance which disappears on shaking with ether.  

b)Due to abnormal connection between the intestinal lymphatic system and urinary   tract 

c) It  may be congenital or acquired.

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 Q1. –What are the solid components of urine in normal health?

Answer-Urine is a fluid composed of water (95%), inorganic and organic solids (5%) which include:

A) Chief inorganic solids :- Sodium, Potassium, Chlorides,

In addition, smaller amounts of calcium,magnesium, sulphate and phosphates, and traces of iron, copper, zinc and iodine.

B) Chief organic solids :

1. Non-protein nitrogen (NPN) compounds like-urea,uric acid, creatinine and amino acids

2. Organic acids   

3. Sugars.

4. In addition, traces of proteins, vitamins, hormones and pigments are also present in the urine.

Q.2- What kind of abnormal components are present in urine in different diseased  states?

Answer- Under different conditions proteins, reducing sugars, ketone bodies, blood, bile pigments, bile salts, lymph or fat may be there in urine. These substances are present in urine in normal health also but only in very minute concentration and hence cannot be detected by routine laboratory procedures. In different diseases their excretion is grossly increased and hence can be easily detected by conventional methods. So it is abnormal concentration which makes them abnormal constituents.

Q.3- What is the amount of urea excreted per day? What are the causes of decreased urea excretion in urine?

Answer- Approximately 15-40 g of urea is excreted per day in normal health. Urinary urea is decreased mainly in renal failure, but it can also be decreased in patients on low protein diet, in growing period (amino acids are needed for active protein synthesis in growing period, so amino group is not available for urea formation, in fact all conditions of positive nitrogen balance like pregnancy, recovery from illness, repair etc. will lower down urinary urea levels). Since urea is formed in liver and excreted through kidneys, hence in liver diseases, urea formation can be impaired, and thus urinary urea level will also be low.

In renal failure blood urea is high but urinary urea is low due to failure of excretion, while in other conditions of positive nitrogen balance and cirrhosis of liver, blood urea is also low.

In contrast urinary urea is high in conditions of negative nitrogen balance or when the body is in a state of catabolism as in starvation, chronic infections, hyperthyroidism, cancers etc. Most importantly decreased urea excretion in the presence of high blood urea level carries most practical attention, since these two features depict impaired functional status of the kidney.

Q.4- What do you expect the level of urinary uric acid in a patient suffering from gout?

Answer- Uric acid is a normal component of urine. Its concentration may be decreased or may be normal in gout depending upon the type of gout. Hyperuricemia in gout may be due to increased formation or due to impaired excretion of uric acid. In renal disease or when there is excessive load of lactate or ketone bodies uric acid is retained (Lactate and ketone bodies share a common transporter- organic acid transporter in the kidney for reabsorption in the renal tubule, hence when excess lactate or ketone bodies are there in filtrate, uric acid is reabsorbed while lactate and ketone bodies are excreted out)), Excess formation of uric acid takes place in various congenital or metabolic defects. In these conditions excretion is not much affected.

Q.5- A 67- year-old male has been hospitalized due to chronic renal failure. He is a known diabetic and hypertensive.  His blood and urine samples have been sent to the laboratory for confirmation of diagnosis. What are the markers of renal failure in blood and urine?

Answer- In renal failure the substances which are normally excreted by the kidney in urine are not excreted and are retained in blood. It can be well presumed that urea, uric acid and creatinine (normal urinary solid components) would be low in urine but would be high in blood. These three parameters – urea, uric acid and creatinine serve as markers of renal failure. Based on the same facts of impaired excretion in renal failure- urea and creatinine clearance are undertaken to determine the functional status of the kidney.

Q.6-What is the source of sulphates in urine?

Answer- Urinary sulphates are derived from sulphur-containing vitamins and amino acids. The sulphur-containing vitamins are-Lipoic acid, Thiamine and Biotin, while Cysteine, Cystine, Methionine and Homocysteine are sulphur-containing amino acids.

Q.7- What are the sources of phosphates in urine?

Answer- Phospholipids, phosphoproteins, nucleic acids and nucleotides contribute phosphates to urine. Phosphates are also derived from demineralization of bones. In hyperparathyroidism and rickets urinary phosphates are increased, while in hypoparathyroidism and renal diseases urinary phosphates are decreased.

Q.8-What is the source of Hippuric acid in urine?

Answer- Hippuric acid is formed in the liver by conjugation reaction. Benzoic acid conjugates with Glycine to form Hippuric acid which is excreted in urine. This reaction takes place exclusively in liver. Hippuric acid excretion test is undertaken by giving a loading dose of sodium benzoate to determine the functional status of the liver. Kidney functions should be normal for this test. In liver diseases Hippuric acid excretion is deceased.

Q.9- Whatis creatinuria?

Answer-Creatine is a precursor of creatinine and upon losing one molecule of water, creatine is converted to creatinine. So creatinine is an anhydrous from of creatine. This is a nonenzymatic spontaneous conversion. Creatine is not normally excreted in urine,it is creatinine which is there in urine. But in certain conditions, creatine is excreted excessively in urine and this state is termed as creatinuria. The causes for this are- Hyperthyroidism, Myasthenia gravis, pregnancy, infancy,uncontrolled diabetes mellitus, growing period, starvation and muscular  diseases.

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