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Practical Biochemistry
Geetha Damodaran K
SECTION ONE: QUALITATIVE ANALYSIS
1:
Reactions of Carbohydrates
1A. REACTIONS OF MONOSACCHARIDES
INTRODUCTION
CLASSIFICATION
REACTIONS OF MONOSACCHARIDES
1. Molisch Test (α-Naphthol Reaction) (Fig. 1A-1)
2. Benedict's Test (Fig. 1A-3)
3. Barfoed's Test (Fig. 1A-5)
4. Rapid Furfural Test
5. Seliwanoff's Test
6. Osazone Test
1B. REACTIONS OF DISACCHARIDES
INTRODUCTION
REACTIONS OF DISACCHARIDES
1. Molisch Test
2. Benedict's Test
3. Barfoed's Test
4. Osazone Test
5. Seliwanoff's Test
6. Rapid Furfural Test
7. Specific Sucrose Test (Fig. 1B-6)
1C. REACTIONS OF POLYSACCHARIDES
INTRODUCTION
1. Homopolysaccharide
2. Heteropolysaccharide
REACTIONS OF STARCH
1. Molisch Test
2. Iodine Test
3. Benedict's Test
4. Starch Hydrolysis Test
1D. IDENTIFICATION OF UNKNOWN CARBOHYDRATES
1E. QUESTIONS
1F. REAGENT PREPARATION
2:
Reactions of Proteins
2A. GENERAL REACTIONS OF PROTEINS
INTRODUCTION
1. PRECIPITATION REACTIONS OF PROTEINS
(i) Precipitation by Salts
(ii) Precipitation by Heavy Metals
(iii) Precipitation by Anionic Reagents (Alkaloids)
(iv) Precipitation by Organic Solvents
(v) Precipitation by Heat
(vi) Precipitation by Strong Mineral Acids
TEST TO DEMONSTRATE DENATURATION AND COAGULATION
2. COLOR REACTIONS OF PROTEINS AND AMINO ACIDS
SPECIFIC COLOR REACTIONS USED TO IDENTIFY THE SIDE CHAIN (R) GROUPS OF AMINO ACIDS
2B. REACTIONS OF ALBUMIN(TABLE 2B-1)
2C. REACTIONS OF CASEIN(TABLE 2C-1)
2D. REACTIONS OF GELATIN(TABLE 2D-1)
2E. IDENTIFICATION OF UNKNOWN PROTEINS
2F. QUESTIONS
2G. REAGENT PREPARATION
3:
Reactions of Lipids
3A. GENERAL REACTIONS OF LIPIDS
INTRODUCTION
REACTIONS OF FATS AND FATTY ACIDS
1. SOLUBILITY TEST
2. Grease Spot Test
3. Acrolein Test (Fig. 3A-1)
4. Emulsification Test
5. Saponification (Table 3A-2)
6. Halogenation Test (Fig. 3A-2)
CHOLESTEROL
COLOR REACTION OF CHOLESTEROL
1. Salkowski's Reaction (H2SO4 Test)(Fig. 3A-3)
2. Liebermann-Burchard Reaction (Acetic Anhydride Sulfuric Acid Test) (Fig. 3A-4)
3B. QUESTIONS
3C. REAGENT PREPARATION
4:
Reactions of Urea
4A. GENERAL REACTIONS OF UREA
INTRODUCTION
Physical Properties of Urea Solution
CHEMICAL REACTIONS
1. Alkaline Hypobromite Test
2. Specific Urease Test (Fig. 4A-4)
4B. QUESTIONS
4C. SAMPLE PREPARATION
5:
Reactions of Creatinine
5A. GENERAL REACTIONS OF CREATININE
INTRODUCTION
Physical Properties
CHEMICAL PROPERTIES
1. Nitroprusside Acetic Acid Test
2. Jaffe' Test (Picric Acid Reaction)(Fig. 5A-2)
5B. QUESTIONS
5C. QUESTIONS
6:
Uric Acid
6A. GENERAL REACTIONS OF URIC ACID
INTRODUCTION (Fig. 6A-1)
PHYSICAL PROPERTIES
CHEMICAL REACTIONS
1. Benedict's Uric Acid Test
2. Schiff's Test
3. Murexide Test
6B. QUESTIONS
6C. REAGENT PREPARATION
7:
Scheme for Identification of Biologically Important Substance in a Given Solution
SCHEME
8:
Urine Analysis
8A. ANALYSIS OF NORMAL CONSTITUENTS OF URINE
INTRODUCTION
SPECIMEN COLLECTION
PHYSICAL EXAMINATION OF URINE (Table 8A-1)
Causes of Oliguria
Polyuria
Causes of Polyuria
Causes of Cloudiness
Measurement of pH
DEMONSTRATION OF INORGANIC CONSTITUENTS OF URINE (TABLE 8A-2)
1. Test for Chloride
2. Test for Sulfates
3,4. Test for Calcium and Phosphates(Fig. 8A-1)
5. Test for Ammonia
ORGANIC CONSTITUENTS OF URINE
DEMONSTRATION OF ORGANIC CONSTITUENTS OF URINE (Table 8A-3)
1. Test for Urea
2. Test for Uric Acid
3. Test for Creatinine
4. Test for Ethereal Sulfates
5. Test for Urobilinogen—Ehrlich's Test
ANALYSIS OF NORMAL CONSTITUENTS OF URINE
8B. ANALYSIS OF ABNORMAL CONSTITUENTS OF URINE
1. GLUCOSE
Benedict's Test (see Chapter 1—Reactions of Carbohydrates)
2. PROTEIN
Heat Coagulation Test (Fig. 8B-2)
3. KETONE BODIES
Rothera's Test (Fig. 8B-3)
4. BLOOD
Benzidine Test
5. BILE SALT
Hay's Test
6. BILE PIGMENT
Modified Fouchet's Test (Fig. 8B-4)
7. UROBILINOGEN
Ehrlich's Test
8C. QUESTIONS
8D. REAGENT PREPARATION
9:
Spectroscopy
9A. SPECTROSCOPIC EXAMINATION OF HEMOGLOBIN PIGMENTS
SPECTROSCOPY AND SPECTROSCOPE
Identification of Different Hb Derivatives
HEMIN CRYSTALS
9B. QUESTIONS
9C. REAGENT PREPARATION
10:
Reactions of Milk
10A. REACTIONS OF MILK
INTRODUCTION
REACTIONS OF MILK
Precipitation of Casein from Milk
TESTS WITH THE PRECIPITATE
TESTS WITH THE FILTRATE
10B. QUESTIONS
10C. REAGENT PREPARATION
SECTION TWO: QUANTITATIVE ANALYSIS
11:
Principles of Colorimetry
11A. INTRODUCTION TO QUANTITATIVE ANALYSIS
COLLECTION OF BLOOD SPECIMEN
Serum
Plasma
COLLECTION OF URINE SAMPLE
COLORIMETRY
PHOTOELECTRIC COLORIMETRY
Transmittance
Extinction
Molar Extinction Coefficient
Specific Extinction Coefficient
Limitations of Beer-Lambert's Law
PHOTOELECTRIC COLORIMETER (SEE FIG. 11A-2)
Measurement in a Photoelectric Colorimeter
11B. QUESTIONS
12:
Determination of Blood Sugar
12A. DETERMINATION OF GLUCOSE CONCENTRATION
SPECIMEN
METHODS
1. Folin-Wu Method
2. Orthotoluidine Method
3. Glucose Oxidase (GOD/POD) Method
12B. GLUCOSE TOLERANCE TEST
12C. QUESTIONS
12D. REAGENT PREPARATION
13:
Determination of Urea
13A. DETERMINATION OF UREA CONCENTRATION
13B. QUESTIONS
13C. REAGENT PREPARATION
14:
Determination of Creatinine
14A. DETERMINATION OF CREATININE CONCENTRATION
INTERPRETATION
CREATININE CLEARANCE TEST
URINE CREATININE DETERMINATION
14B. QUESTIONS
14C. REAGENT PREPARATION
15:
Determination of Total Protein and Albumin
15A. DETERMINATION OF SERUM TOTAL PROTEIN AND ALBUMIN
DETERMINATION OF ALBUMIN
INTERPRETATION
SERUM TOTAL PROTEINS
SERUM ALBUMIN
GLOBULINS
Albumin globulin ratio
15B. QUESTIONS
15C. REAGENT PREPARATION
16:
Determination of Cholesterol
16A. DETERMINATION OF TOTAL CHOLESTEROL
DETERMINATION OF TOTAL CHOLESTEROL
1. Photometric Method
2. Enzymatic Method
16B. QUESTIONS
16C. REAGENT PREPARATION
17:
Determination of Uric Acid
17A. DETERMINATION OF URIC ACID CONCENTRATION
Interpretation
Causes of Hyperuricemia
HYPOURICEMIA
17B. QUESTIONS
17C. REAGENT PREPARATION
18:
Determination of Bilirubin
18A. DETERMINATION OF SERUM BILIRUBIN
DETERMINATION OF SERUM BILIRUBIN
INTERPRETATION
Reference Range
18B. QUESTIONS
18C. REAGENT PREPARATION
19:
Determination of Transaminases
19A. DETERMINATION OF SERUM TRANSAMINASES
INTERPRETATION
ALT and AST in Hepatic Disorders
AST in Ischemic Heart Disease
19B. QUESTIONS
19C. REAGENT PREPARATION
20:
Determination of Alkaline Phosphatase
20A. DETERMINATION OF SERUM ALKALINE PHOSPHATASE
ASSAY OF ALKALINE PHOSPHATASE
INTERPRETATION
High Alkaline Phosphatase Activity
ISOENZYMES OF ALP
20B. QUESTIONS
20C. REAGENT PREPARATION
21:
Determination of Calcium
21A. DETERMINATION OF CALCIUM CONCENTRATION
DETERMINATION OF CALCIUM
1. Photometric Method—O-Cresolphthalein Method
2. Titration Method—Method of Clarkand Collip (1925)
INTERPRETATION
Reference Range in Adults
Alterations in Serum Calcium Levels
Low Serum Calcium Levels
High Serum Calcium Levels
21B. QUESTIONS
21C. REAGENT PREPARATION
22:
Determination of Phosphorus
22A. DETERMINATION OF INORGANIC PHOSPHORUS
DETERMINATION OF INORGANIC PHOSPHORUS
1. Photometric Method—Method of Fisk and Subba Rao
INTERPRETATION
Reference Range
ALTERATIONS IN INORGANIC PHOSPHORUS IN SERUM
Hypophosphatemia
Hyperphosphatemia
22B. QUESTIONS
22C. REAGENT PREPARATION
23:
Determination of Titrable Acidity and Ammonia in Urine
23A. DETERMINATION OF AMMONIA AND TITRABLE ACID IN URINE
DETERMINATION OF TITRABLE ACIDITY AND AMMONIA
Method Used: Titration Method
INTERPRETATION
23B. QUESTIONS
23C. REAGENT PREPARATION
24:
Determination of Urine Chloride
24A. DETERMINATION OF URINE CHLORIDE
DETERMINATION OF URINE CHLORIDE
Method Used: Titration Method
INTERPRETATION
24B. QUESTIONS
24C. REAGENT PREPARATION
SECTION THREE: CHARTS
25:
Charts
25A. ACID-BASE DISORDERS
25B. JAUNDICE
25C. DIABETES
25D. IN BORN ERRORS OF METABOLISM
25E. PORPHYRIAS
25F. VITAMINS AND MINERALS
25G. TUMOR MARKERS
25H. WATER AND ELECTROLYTES
25I. BIOCHEMICAL PARAMETERSOF RENAL FUNCTIONS
25J. BIOCHEMICAL DIAGNOSIS OF MYOCARDIAL INFARCTION
25K. ENZYMOLOGY
SECTION FOUR: SPOTTERS
26:
Spotters
26A. INSTRUMENTS
26A-1. ESBACH'S ALBUMINOMETER
Spotter-1
Details of Esbach's Albuminometer
26A-2. ELECTROPHORETIC TANK
Spotter-2
Details of Electrophoretic Tank
26A-3. RYLE'S STOMACH TUBE
Spotter-3
Details of Ryle's Stomach Tube
26A-4. FOLIN-WU SUGAR TUBE
Spotter-4
Details of Folin-WU Sugar Tube
26A-5. URINOMETER (HYDROMETER)
Spotter-5
Spotter-6
Spotter-7
Details of Urinometer (Hydrometer)
Specific Gravity of the Urine of Normal Individuals: 1.015–1.025
High Specific Gravity
Low Specific Gravity Leading to Specific Gravity Less than 1010 (Fixed Specific Gravity)
26A-6. pH METER
Spotter-8
Details of pH Meter
26A-7. SEMIAUTOMATIC BIOCHEMISTRY ANALYZER
Spotter-9
Details of Photoelectric Colorimetry
26A-8. DRY CHEMISTRY STRIP TEST FOR URINE GLUCOSE
Spotter-10
26B. REAGENTS
26B-1
Spotter-11
26B-2
Spotter-12
26B-3
Spotter-13
26B-4
Spotter-14
26B-5
Spotter-15
26B-6
Spotter-16
26B-7
Spotter-17
26B-8
Spotter-18
26C. INDICATORS
26C-1
Spotter-19
26C-2
Spotter-20
26C-3
Spotter-21
26C-4
Spotter-22
26D. CRYSTALS
26D-1
Spotter-23
26D-2
Spotter-24
26D-3
Spotter-25
26D-4
Spotter-26
26D-5
Spotter-27
26E. SEPARATION TECHNIQUES
26E-1
Spotter-28
26E-2
Spotter-29
26E-3
Spotter-30
26E-4
Spotter-31
More about Paper Chromatography
26F. GRAPHS
26F-1
Spotter-32
26F-2
Spotter-33
26F-3
Spotter-34
26F-4
Spotter-35
26F-5
Spotter-36
26F-6
Spotter-37
26F-7
Spotter-38
26F-8
Spotter-39
26F-9
Spotter-40
26F-10
Spotter-41
26F-11
Spotter-42
26F-12
Spotter-43
26G. TESTS
26G-1
Spotter-44
26G-2
Spotter-45
26G-3
Spotter-46
26G-4
Spotter-47
26G-5
Spotter-48
26G-6
Spotter-49
26G-7
Spotter-50
26G-8
Spotter-51
26G-9
Spotter-52
26G-10
Spotter-53
26G-11
Spotter-54
26G-12
Spotter-55
26G-13
Spotter-56
26G-14
Spotter-57
26G-15
Spotter-58
26G-16
Spotter-59
26G-17
Spotter-60
26G-18
Spotter-61
26G-19
Spotter-62
26H. NUTRITION
26H-1
Spotter-63
26H-2
Spotter-64
26H-3
Spotter-65
26H-4
Spotter-66
26H-5
Spotter-67
26H-6
Spotter-68
26H-7
Spotter-69
26H-8
Spotter-70
26H-9
Spotter-71
26H-10
Spotter-72
26H-11
Spotter-73
26I. SPECTROSCOPY
26I-1
Spotter-74
26I-2
Spotter-75
26I-3
Spotter-76
26I-4
Spotter-77
26I-5
Spotter-78
26I-6
Spotter-79
26J. CONCEPTUAL QUESTIONS
26J-1
Spotter-80
26J-2
Spotter-81
26J-3
Spotter-82
26J-4
Spotter-83
26J-5
Spotter-84
26J-6
Spotter-85
26J-7
Spotter-86
26J-8
Spotter-87
26J-9
Spotter-88
26J-10
Spotter-89
26J-11
Spotter-90
26J-12
Spotter-91
26J-13
Spotter-92
26J-14
Spotter-93
26J-15
Spotter-94
26J-16
Spotter-95
26J-17
Spotter-96
26J-18
Spotter-97
26J-19
Spotter-98
26J-20
Spotter-99
SECTION FIVE: OSPE (OBJECTIVE STRUCTURED PRACTICAL EXAMINATION) QUESTIONS
27:
OSPE (Objective Structured Practical Examination) Questions
27A. OSPE - QUALITATIVE EXPERIMENTS
27B. OSPE-QUANTITATIVE EXPERIMENTS
INDEX
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