Clinical Biochemistry I (Sheet 1) PDF 2024

# Chemical Biochemistry I ## 13 ورقة **Course Lecturer:** Dr. Saleh Bilgasem **Semester:** 5th Semester **Year:** 2024 ## Clinical biochemistry Clinical biochemistry (Clinical chemistry or chemical pathology) is the division of medical biochemistry that deals with physiological chemical meas...

# Chemical Biochemistry I ## 13 ورقة **Course Lecturer:** Dr. Saleh Bilgasem **Semester:** 5th Semester **Year:** 2024 ## Clinical biochemistry Clinical biochemistry (Clinical chemistry or chemical pathology) is the division of medical biochemistry that deals with physiological chemical measurements of both natural & unnatural. ## Purpose of clinical chemistry tests: Clinical chemistry tests are the application part of biochemistry for the diagnosis of the clinical condition by determining various chemical constituents in different body fluids (both natural & unnatural) in blood, urine and other body. Determination can be qualitative or quantitative: * **Qualitative** determination of urea and creatinine in blood and urine helps in the assessment of kidney functioning or * **Quantitative** determination of inorganic substances like Na, K, Ca, Cl...etc. or may be enzymes or hormones in order to diagnose diseases, by many methods such as manual, kit and automated. ## Types of specimens for chemical analysis: Whole blood, serum or plasma. The most common specimen is serum, collected in a tube with no anticoagulant so that the blood will clot. * **Urine** - often 24 hour collection * **Others:** Pleural fluid, Cerebrospinal Fluid (CSF) and other fluids. ## 1. Blood Blood could be obtained from three different sources: * **Capillary blood:** It is obtained from the finger of the thumb after cleaning with spirit. Such sample is used for hematological analysis like RBC & WBC count, Hb...etc. * **Arterial blood:** It is restricted for blood gases analysis. CO2, O2, HCO3, pH ...etc. * **Venous blood:** Used for analysis of all chemical constituents in blood, like glucose, urea, cholesterol...etc. ## Avoiding hemolysis: Syringe and needle should be clean and dry. Minimum amount of constriction should be applied to the arm. Blood should be flow slowly and steadily into the syringe. Needle should be removed from the syringe before expelling blood into the tube. Gentle rotation should be used when the tube contains anticoagulant. ## Anticogulants: * **Heparin:** It is the most convenient anticoagulant because it does not produce any change in the chemical constituents of the blood. It acts as antithrombin, preventing the transformation of prothrombin into thrombin and thus preventing the transformation of fibrinogen into fibrin. * **Sodium and potassium oxalate:** It inhibits blood coagulation by forming insoluble complex with calcium ions which are necessary for coagulation. * **Ammonium oxalate** (double oxalate): 3 part of ammonium oxalate + 2 part of potassium oxalate to form double oxalate. It is used for certain hematological investigation (2mg/ml) of blood is used to prevent clotting. It should not be used when NH3 or urea is wanted for estimation because it rises the ratio of nitrogen. * **Sodium citrate:** This salt does not precipitate calcium but change it into non-ionized salt. 3mg/ml blood is used. * **Sodium fluoride:** Although usually considered as a preservative for blood glucose determination, also act as a weak anticoagulant such as potassium oxalate. 2mg/ml blood is used to prevent clotting. It exerts its action by inhibiting the enzyme system involved in glycolysis (Enolase). * **Ethylene diamine tetra acetic acid** (EDTA): It is useful for hematological examination since it preserves the cellular constituents of the blood. It is used as disodium or dipotassium salt, the later being more soluble. 2mg/ml of blood is used to prevent clotting, EDTA is a chelating agent derives its anticoagulant activity by chelating the calcium which is needed for clotting mechanism. ## Protein precipitants: In clinical chemistry analysis the first stage is to remove protein from serum in order to prepare protein free filtrate (PFF) coz protein is colloidal substance which will produce turbidity of the solution which will affect the reading of absorbance in spectrophotometer. The common protein precipitant used in clinical chemistry laboratory are: 1. Tri chloroacetic acid 2. Alkaline zinc salt 3. Organic solvent. ## 2. Urine: It is an ultra filtrate fluid from the blood, the filtration occurs in the kidney and excreted outside the body through urinary tract. ### Terms used to describe urine volume: * **Polyuria:** It is an increase in the daily output of urine as in cases of D.M. * **Oligouria:** Decrease in urine volume output/day as in fever, acute nephritis, diarrhea and nephritis. * **Anuria:** It is the total suppression of urine excretion found in shock, mercury poissoning, and sulphadrug treatment. ## Changing of urine constituents on keeping: 1. **Bacteriological factor**: Bacterial growth may take place which convert urea to ammonia as in the following reaction: The liberation of ammonia will change the pH of urine from acidic to basic side. 2. **Some yeast**: It may act on urine sample changing the glucose content to pyruvic acid, their fore such sample is not suitable for glucose determination. 3. **Phosphate**: It may be precipitated when the pH of urine is alkaline. 4. **Uric acid and other urates:** They are precipitated in cold urine; their fore such sample should be warmed before starting analysis. ## Urine sample processing includes: 1. **Chemical examination**: Sugar, protein, acetone, bilirubin,...etc. 2. **Macroscopically examination**: Color, odor, specific gravity. transparency.....etc. 3. **Microscopically examination**: Cast, pus, RBC, crystals, etc ## Urine Samples: * **Random sample**: This sample may differ from person to another depending on physical, pathological and dietary condition taken at any time and kept in covered vessel. **Morning sample:** Usually the most concentrated of any other time of collection and is the most uniform from day to day, that is why it is preferred for analysis than any other sample. * **Timed sample**: Are needed when quantitative estimation is to be carried out, used for measuring the kidney function. The sample may be 24hr, 3hr, ...etc collected in suitable container containing preservative like benzoic acid, toluene,.... can be used for measuring the total urinary excretion per day. * **Catheterized sample**: Collected in cases of anuria (total suppression of urine excretion) using a catheter. * **Bacteriological sample**: Collected in sterilized, clean test tube, mid stream is preferred for such analysis, the tube should be closed directly after urine TEMP has been put in. ## 1. Normal constituents of urine: * **Urea**: Constitute about half of total constituents of urine, it is the principal end product of protein metabolism. **Ammonia:** Normal urine contain little amount of ammonia, its formation increased in cases of acidosis due to D.M. * **Creatine and creatinine:** Creatinine is a product of creatine dehydration, excreted normally in constant amount in urine from day to day. Creatine is synthesized in the liver, this compound transmitted by the blood to the muscle, stored as creatinephosohate which serves as energy source, then liberate energy and metabolized to creatinine which is excreted in urine. * **Uric acid:** It is the end product of purine metabolism. Derived from nucleic acids DNA & RNA Amino acids: All naturally occurring. As are found in normal urine, their excretion is increased in urine of adults when compared with urine of infants * **Salt & minerals:** As chloride, sulfate, phosphate, oxalate, sodium, potassium, calcium, magnesium, etc. ## 2. Abnormal constituents of urine: * **Albumin:** It is a protein. The presence of albumin in urine is referred to as albumin uria. **Physiologic proteinuria:** Found after severe exercise, emotional stress. **Pathologic proteinuria:** Pre renal, renal & post renal. * **Glucose**: Glycosuria is the appearance of glucose in urine in high concentration as in the case of D.M. **Ketone bodies:** Acetone, acetoacetic acid and P-hydroxy butyric acid are present in urine of patient of severe D.M due to an increase in fatty acid catabolism. * **Bilirubin**: It is a pigment found in urine of patient with jaundice. * **Blood:** Heamaturia means the presence of RBCs in urine due to the lesion of any part of urinary tract. ## 3. Electrolytes: Electrolytes are distributed in solution through all the body fluids, the cations (positively charged) of these electrolytes are Na, K, Ca & Mg. The chief cation of plasma and other extracellular fluid is Na and in the intracellular fluid is K. Among the anions of body fluids are Cl, HCO3, H2PO4, HPO4, SO₄ and protein. The chief anion of plasma and other extracellular fluids is Cl and in the intracellular fluids is HPO4 and protein. ### A. Sodium ion (Na): Sodium ion has the following functions in the body: 1. It is the major cation of extracellular fluid, combined with HCO, or Cl for acid-base regulation. 2. Sodium maintains osmotic pressure of extracellular fluid (ECF) balance. 3. Keep normal irritability of muscles and the permeability of the cells. Daily requirement is 4-5 gm as NaCl, absorbed directly from intestine and excreted with urine. It is present as (1.09) gm of (Na*) per Kg of body weight. **Regulation of sodium reabsorption:** The maintenance of Na balance is under the control of the hormone aldosterone from the adrenal cortex. Aldosterone increases the reabsorption of Na by the kidney tubules. **Clinical significance:** NR. for Na* in serum = 135-148 m.mole/L. NR. for Nat in urine = 143-217 m.mole/L.. NR. for Nat in CSF = 138-155m.mole/L. * **Hyponaturemia Dehydration**: as in vomiting, diarrhea, in acidosis due to poor renal reabsorption, polyuria, excessive sweating. When there is a decreased secretion of aldosterone hormone as in Addison disease, Hypotension. * **Hypernaturemia:** Increased in serum sodium is rare, it may occur in the following conditions: 1. When there is an increased secretion of aldosterone hormone as in Cushing disease. 2. When large amount of fluid given to patient intravenously. 3. In high cortisone intake. ### B. Potassium ion (K): Potassium is mainly an intracellular cation as only 2% of total body potassium is located extracellular. **Functions:** 1. In extracellular fluid, it affects the normal function of muscle especially heart muscle. 2. In intracellular fluid, it has the same function of Na, by keeping normal osmotic pressure, normal acid-base balance, it activates some enzymes and affects ribosome for protein synthesis. **Absorption:** Absorbed directly from intestine and excreted with urine, the capacity of the kidney to excrete K is so great so hyperkalemia will not occur. Aldosterone hormone increases excretion of K. **Clinical significance:** * N.V. of serum K = (3.5-5) m. mole/ L. * N.V. of Urine K = (3.5-5) m. mole/L. ### Hyperkalemia: 1. Renal failure 2. Advanced dehydration. 3. Addison disease 4. Myocardial infarction. 5. Metabolic acidosis. 6. Urinary obstruction. 7. Insulin deficiency ### Hyporkalemia: 1. Malnutrition 2. Severe diarrhea & Vomiting 3. Alkalosis will lead to hypokalemia. 4. Cushing disease In most of these cases, intracellular K is transferred to the extracellular fluid, and this K is quickly removed by the kidney coz aldosterone hormone increases excretion of K. ## 4. Phosphorus Phosphorus is found in every cell of the body, but most of it about 80% is combined with calcium to form calcium fluorophosphate in the bone, the remainder 20% is found in the following forms: 1. Free inorganic phosphate (in serum) 2. Phospholipid (in serum) 3. Organic phosphate (nucleic acids, nucleotides, organic phosphate esters) in erythrocytes. **Functions:** 1. Mineralization of bone along with Ca. 2. TEMPHas a major as a component of vital molecule such as nucleic acid nucleotides (NAD, NADP) phospholipids and some protein. 3. Integral part in buffering system of blood HPO4, H2PO4 in ratio 4:1 at PH=7.4. 4. Essential components of ATP they form of energy transport and storage in living cell. **Absorption:** 1. Free phosphate absorbed in jejunum by active and passive diffusion. 2. Unlike calcium phosphate absorption is proportional to dietary intake. 3. Phosphate absorption increased by present (Vit. D) and the synthesis of (Vit.D) increased when (PO4) decreased. 4. Deposition of phosphate as hydroxyl apatite in regulated by (PTH). 5. 85-90% of plasma phosphate is filtered by glomerular filtration. 6. Reabsorption of PO, is inhibited by PTH and increase by Vit.D. **Clinical significance:** * N.V. of S. PO, 1st year of live - 4-7 mg/dl. * Children - 4.5-6.5 mg/dl. * Adult-2.4-4.5 mg/dl. * PO, decrease during menstrual period. * PO, decrease after Meal. **Hyperphosphatemia:** 1. Hypoparathroidism 2. Hypervitaminosis 3. Renal failure 4. In normal children due to secretion growth hormone 5. Diabetic ketosis. **Hypophosphatemia:** 1. hyperparathyroidism 2. Rickets(decrease Vit D) 3. Acute alcoholism 4. Fanconi syndrome (disorder in PO, reabsorption). 5. Osteomalicia ## 5. Iron (Fe): The total amount of iron in an adult is approximately (4-5)gm distributed as follows: * Hemoglobin (60-70)%. * Myoglobin (3-5)%. * In enzymes such as cytochrome, cytochrome oxidase, peroxidase, catalase < (1)%. * Storage form mainly ferritin (25)% **Functions:** 1. In humans, iron is an essential component of proteins involved in oxygen transport, as it is a part of Hb molecule. 2. It is also essential for the regulation of cell growth and differentiation. 3. It serves in electron transport system in living tissue depending on its ability to undergo valence changes, such as its function in cytochrome, cytochrome oxidase and catalase enzyme. **Clinical significance:** * **Increased** 1. RBCs destruction as in hemolytic anemia. 2. Decreased utilization of iron as in lead poisoning. 3. Hepatitis, due to increased release of iron from body stores. 4. Homochromatosis, due to increased absorption of iron from stomach and intestine. 5. Lead poisoning.