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Chemical – Bio Estimation and Analysis of Urine

Chemical – Bio Estimation and Analysis of Urine. By Dr.G.Srinivasagan Rajapalayam Rajus College. Urine consists of:. (96%) water. Inorganic: Cl - , Na, K. trace amounts of: sulfate, HCO 3 etc.). Urine:

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Chemical – Bio Estimation and Analysis of Urine

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  1. Chemical – Bio Estimation and Analysis of Urine By Dr.G.Srinivasagan Rajapalayam Rajus College

  2. Urine consists of: (96%) water Inorganic: Cl-, Na, K. trace amounts of: sulfate, HCO3 etc.) • Urine: • Is an ultrafiltrate of plasma from which glucose, amino acids, water and other substances essential to body metabolism have been reabsorbed. • Urine carries waste products and excess water out of the body. (2%) Urea: (half) (2%) Other compounds (4%) dissolved solids: Organic:creatinine uric acid Routine Urine Analysis

  3. Urine Analysis: Routine Urinalysis (Routine-UA): • It consists of a group of tests performed as part of physical examination. It involves macroscopic and microscopic analysis. Type of analysis: • macroscopic analysis: • microscopic examination: urine sediment is examined under microscope to identify the components of the urinary sediments. physical characteristics chemical analysis

  4. Steps in basic urine analysis Three steps analysis: First:physical characteristics of urine are noted and recorded. Second:series of chemical tests is run. Third: urine sediment is examined under microscope to identify the components of sediments.

  5. Urine Collection: Types of urine specimens: • type of specimen and collection procedure are determined by physician and depend on the tests to be performed. There are basically four types of urine specimens: • First morning specimen • Random urine specimen • Fractional collection • Timed collection • Composition and concentration of urine changes during 24hr • Urine conc. vary according: to water intake and physical activities.

  6. Truly representative sample: • it is necessary to regulate: time of collection, length of collection period, patient's dietary, medical intake and method of collection. • Initial morning sample is preferred (particularly for protein analysis) because they are more concentrated from overnight retention in bladder. • Time of analysis: - must analyzed within 1h at room temp. or within 8hr at 2oC- 8oC - If not assayed within these time limits, several changes will occur. • sample should collected in a clean container. • urine container must be sterile if the urine is to be cultured. • For microscopic examination, the urine must be fresh.

  7. I- Physical Characteristics: • direct visual observation. • Normal fresh urine: Color: pale or dark yellow-amber, clear. • Vol:750 - 2000 ml/24hr. • Physical examination involves: • Color • Transparency • Odour • Volume • pH • Specific gravity

  8. Normal Chemical Constituents of Urine: Organic: urea, uric acid, creatinine Inorganic: Cl-, PO4-3, HBO3, NH4, SO4-2 1- Urea: 1ml urine + 3ml NaOCL(sodium hypochlorite) ==>Evolution of N2 gas. 2- Uric acid UA: 1ml urine + 0.5 ml 10% NaOH +1ml Folinsreagent ===> Blue color. 3- Creatinine: - 1ml urine + drops Picric acid + drops NaOH ====> red color ppt. Note: if reaction is acidified with HCL, the color changes to yellow.

  9. 4- Chloride: • 1ml urine + drops HNO3 +1 ml AgNO3===> white ppt of AgCL 5- Phosphate: 1ml urine + 1ml conc. HNO3 + 1ml NH4-molybdate===>Yellow color. 6- Carbonate: 1ml urine + drops conc. HCL ==> Na2CO3+ 2 HCL ==> H2O + 2NaCL + CO2 7- Ammonia: - Make urine alkaline with NaOH. Close the tube with a cork containing another side tube dipped in Nessler's reagent. Heat the urine and then notice the evolving of NH3 in Nessler's reagent - Detect NH3 by its odour. - 1ml urine + 1ml phenol + 1ml NaBr =======> Blue color. 8- Sulphates: - 1ml urine + 2 drops conc. HCL + few drops BaCL2 ===> White ppt of BaSO4. SO4 + BaCL2 =====> BaSO4 + 2CL- (effervescence)

  10. Chemical characterstics • Protein. Protein is normally not found in the urine. Fever, hard exercise, pregnancy, and some diseases, especially kidney disease, may cause protein to be in the urine. • Glucose. Glucose is the type of sugar found in blood. Normally there is very little or no glucose in urine. When the blood sugar level is very high, as in uncontrolled diabetes, the sugar spills over into the urine. Glucose can also be found in urine when the kidneys are damaged or diseased. • Nitrites. Bacteria that cause a urinary tract infection (UTI) make an enzyme that changes urinary nitrates to nitrites. • Leukocyte esterase (WBC esterase). Leukocyte esterase shows leukocytes (white blood cells [WBCs]) in the urine. • Ketones. When fat is broken down for energy, the body makes substances called ketones (or ketone bodies). These are passed in the urine. Large amounts of ketones in the urine may mean a very serious condition, diabetic ketoacidosis, is present. A diet low in sugars and starches (carbohydrates), starvation, or severe vomiting may also cause ketones to be in the urine.

  11. Identification of Pathological Physical and Chemical UrineConstituents Abnormal (Pathological) constituents of urine: 1- Macroscopic analysis: • physical tests • chemical tests 2- Microscopic analysis: • Pathological urine constituents are substances which are not usually present in urine such as glucose, protein, ketones, RBCs, Hb, bilirubin…. etc.

  12. Glucose Bilirubin Ketones Specific Gravity Blood pH Protein Urobilinogen Nitrite Leukocyte How to detect abnormal constituents: Urine strip: • Strip is filter paper or plastic which has chemical substance (reagent) coated on it on different pads. • It gives color when react with substance in urine. • The produced color is compared with chart color visually or mechanically assessed.

  13. Abnormal Urine Constituents include: Proteinurea: • is the presence of abnormal amount of protein in urine. • Urine of healthy individual contains no protein due to: • In normal physiology, small M.wt. proteinsis reabsorbed by kidney tubules (proximal tubule) • large M.wt of protein so it can't pass through kidney tubule to urine. unless kidney tubule has damage. • The main protein in urine is albumin therefore, proteinuria=albuminuria

  14. Microalbuminuria: • Is the presence of small amounts of albumin in urine. • It is very important in detection of early stage of nephronpathy and in diagnosis of DM complication (nephropathy). • High protein in urine makes urine looks foamy. • Associated with face or feet abnormal odema, due to disturbance of liquid balance in body due to protein loss.

  15. Detection: • Qualitative test: using a reagent test strip. • Quantitative test: depends on volume and time of urine (protein conc. in urine may vary with time and volume) • Most assays are performed on urine sample of 12-24h. Reference value: • Quantitative for 24-h urine: Male:1-4 mg/dl Female: 3-10 mg/dl Child: 1-10mg/dl • Qualitative reference value: Normal = Negative

  16. Thank you

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