Carbohydrates - Clinical Chemistry 1 Lab PDF

CARBOHYDRATES Clinical Chemistry 1 Laboratory Introduction  Hydrates if aldehydes or ketone derivatives based on the location of the CO functional group  Primary energy source stored primarily as glycogen  Disease states involved hyperglycemia and hypoglycemia  Contain C, H, and O (Cx(H20)y...

CARBOHYDRATES Clinical Chemistry 1 Laboratory Introduction  Hydrates if aldehydes or ketone derivatives based on the location of the CO functional group  Primary energy source stored primarily as glycogen  Disease states involved hyperglycemia and hypoglycemia  Contain C, H, and O (Cx(H20)y with C=O and –OH functional groups Introduction  Functions of Carbohydrates:  Major component of the human diet, an important source of body energy  Found as part of the cell membrane  Found as antigens in RBCs to determine the blood type(ABO Blood groups)  Glucose is the principal and almost exclusive carbohydrate circulating in the blood.  The brain is completely dependent on blood glucose for energy production – 2-3 of glucose utilization in resting adults that occurs in the CNS. Introduction Introduction Introduction Cellulose Glucose Metabolism  Glucose can be derived from  Diet  Body stores like glycogen  Endogenous synthesis from proteins or glycerol of triglycerides. Glucose metabolism Glucose Metabolism  Glycolysis (EMP Pathway)  Metabolism of Glucose to Lactate or pyruvate for the production of energy  Glucose – Insulin -> ATP + Lactate or Pyruvate Glucose Metabolism  Gluconeogenesis  Formation of G-6-P from non- carbohydrate source  Fats(Ketone bodies), Protein(Urea Nitrogen) Glucose Metabolism  Glycogenesis  Conversion of Glucose to Glycogen  Glucose –Insulin -> Glycogen Glucose Metabolism  Glycogenolysis  Breakdown of glycogen to G-6-P  Glycogen –Glucagon -> Glucose Glucose Metabolism  Glycolysis (EMP Pathway)  Metabolism of Glucose to Lactate or pyruvate for the production of energy  Glucose – Insulin -> ATP + Lactate or Pyruvate  Gluconeogenesis  Formation of G-6-P from non-carbohydrate source  Fats(Ketone bodies), Protein(Urea Nitrogen)  Glycogenesis  Conversion of Glucose to Glycogen  Glucose –Insulin -> Glycogen  Glycogenolysis  Breakdown of glycogen to G-6-P  Glycogen –Glucagon -> Glucose Glucose Metabolism  Lipogenesis  Conversion of carbohydrates to Fatty acids  Lipolysis  Decomposition of fat Regulation of Glucose Metabolism  Insulin  Glucagon  Epinephrine  Cortisol  Growth Hormone  Thyroxine  Somatostatin Regulation of Glucose Metabolism  Insulin (Hypoglycemic agent)  Primary hormone responsible for decreasing ↓ glucose  Synthesized by the β cells of the islets of Langerhans  ↑ glycogenesis, glycolysis, lipogenesis; and ↓ glycogenolysis.  Regulation of Glucose Metabolism  Glucagon (hyperglycemic agent)  Primary hormone responsible increasing blood glucose  Synthesized by the α cells of the islets of Langerhans (pancreas)  Released during stress and fasting states  ↑ glycogenolysis and gluconeogenesis. Regulation of Glucose Metabolism  Epinephrine  Produced by the adrenal medulla, ↑ blood glucose  Released during times of physical and emotional stress  Inhibits insulin secretion, ↑ glycogenolysis and lipolysis Regulation of Glucose Metabolism  Cortisol (Glucocorticoids)  Produced by the adrenal cortex, ↑ plasma glucose  ↓ intestinal entry of glucose into the cell, ↑ gluconeogenesis, glycogenolysis and lipolysis Regulation of Glucose Metabolism  Growth hormone  Produced by the anterior pituitary gland; ↑ plasma glucose  ↓ glucose entry to cells, ↑ glycolysis Regulation of Glucose Metabolism  Thyroxine  Produced by the thyroid gland; ↑ plasma glucose  ↑ glycogenolysis, gluconeogenesis and glucose intestinal absorption Regulation of Glucose Metabolism  Somatostatin  Produced by the Delta cells of the islet of Langerhans in the pancreas and hypothalamus  ↑ plasma glucose by inhibition of insulin, glucagon, GH, etc. Disease states in Glucose Metabolism Clinical Chemistry 1 - Carbohydrates Diabetes Mellitus  Metabolic disease characterized by hyperglycemia resulting from defects in insulin secretion or insulin action  Type 1 (IDDM)  Type 2 (NIDDM)  Other  Gestational Type 1 DM  Insulin Dependent Diabetes Mellitus  β-Cell Destruction  Absolute insulin deficiency  Autoantibodies  5-10% of all cases of diabetes  Occurs in childhood and adolescence  Absence of Insulin with excess in glucagon Type 2 DM  90% of all cases of diabetes  Adult onset  Insulin resistance w/ secretory defect  Relative insulin deficiency  ↑ with age, obesity and lack of exercise  Insulin is present (hyperinsulinemia)  Glucagon secretion is attenuated  Non Insulin dependent  Ketosis tendency is seldom  Greater tendency to develop hyperosmolar states  Polydipsia, Polyphagia, Polyuria Diabetes Mellitus Diabetes Mellitus  Other  Associated with secondary conditions  Gestational  Glucose intolerance during pregnancy  Due to metabolic and hormonal changes Diabetes Insipidus  rare but treatable condition in which your body produces too much urine and isn’t able to properly retain water.  happens when your body doesn’t make enough antidiuretic hormone (ADH) or your kidneys don’t use it properly. Your body needs ADH to retain appropriate amounts of water. Without ADH, your body loses water through urine.  Summary Characteristics Type 1 DM Type 2 DM Frequency 5% - 10% 90% - 95% Age of Onset Any, but most common in children and Most common with advancing age, but young adults can occur in children and adolescents Risk Factors Genetics, Autoimmune, environmental Genetic, obesity sedentary lifestyle,, race/ethnicity/hypertension, dyslipedemia, PCOS Pathogenesis Destruction of pancreatic beta cells, No autoimmunity insulin resistance and usually autoimmune progressive insulin deficiency C-peptide Levels Very low or undetectable Detectable Prediabetes Autoantibodies (GAD65, IA-2, IAA, ZnT8) Autoantibodies absent  Summary Characteristics Type 1 DM Type 2 DM Medication Therapy Insulin absolutely necessary; multiple Oral agents and/or noninsulin injectable daily injections or insulin pump hypoglycemic drugs Insulin commonly needed Therapy to prevent None known Lifestyle and oral medications or delay onset of diabetes Previous names Insulin dependent DM Non-insulin-dependent DM Juvenile onset DM Adult-onset DM Hyperglycemia  Increased glucose in plasma and urine  Increase urine specific gravity  Increased serum and urine osmolality  Ketones in serum and urine (ketonemia and ketonuria)  Decreased blood and urine pH  Electrolyte imbalance Hypoglycemia  Causes of Hypoglycemia  Patients Appears Healthy  No coexisting disease  Insulinoma, Islet hyperplasia  Factitial hypoglycemia (insulin/sulfonylurea)  Severe exercise, Ketotic hypoglycemia  Compensated coexistent  Drugs/disease  Patients appears ill  Drugs, Predisposing illness, Hospitalized patient Glycogen Storage Disorders Clinical Chemistry 1 Laboratory - Carbohydrates Glycogen Storage Disorders  Results from deficiency of specific enzyme that causes an alteration in the glycogen metabolism  Patients exhibit hepatomegaly and hypoglycemia Glycogen Storage Disorders Hypoglycemia  Genetic Defects in Carbohydrate Metabolism  Von Gierke Disease (glucose-6-phosphatase deficiency type 1)  Glycogen build up in the liver due to inhibition of hepatic glycogenolysis  Galactosemia (galactose-1-phosphate uridyl transferase deficiency)  Inhibition of glycogenolysis Laboratory Considerations  WB glucose concentration is 11% lower than plasma  Serum / plasma must be refrigerated and separated from the cells within 1 hr  Sodium flouride (gray-top) can be used to inhibit glycolytic enzymes  FBG should be obtained in the morning after 8 to 10 hours fasting (not longer than 16 hours) – Bishop  Arterial Blood>Venous  Capillary>Venous Determination Methods  Fasting Blood Glucose  Point of care  2-Hrs Post Prandial Sugar  Oral Glucose Tolerance Test  HbA1C – Glycated Hemoglobin  Fructosamine – Glycated Albumin* Measurement  Non-Enzymatic Methods of Glucose Measurement  Nelson Somogyi  Hagedorn Jensen  Ortho-toluidine (Dubowski)  Enzymatic Methods of Glucose Measurement  Glucose oxidase (Saifer Gernstenfield)  Hexokinase (Reference method)  Clinitest Non-Enzymatic Methods of Glucose Measurement  Nelson Somogyi  Copper reduction method (uses BaSO4 to remove saccharoids)  Glucose + arsenomolybdic acid -> arsenomolybdenum blue  Hagedorn Jensen  Ferric reduction method (inverse colorimetry)  Glucose + Ferricyanide (yellow) -> Ferrocyanide (colorless)  Ortho-toluidine (Dubowski)  Condensation of carbohydrates with aromatic amines  producing Schiff bases (green) Enzymatic Methods of Glucose Measurement  Glucose oxidase (Saifer Gernstenfield)  Couple reaction is known as Trinder’s reaction  False Low results due to ↑ uric acid, bilirubin and ascorbic acid  O2 Consumption electrode (polarographic glucose analyzers) measure oxygen depletion Enzymatic Methods of Glucose Measurement  Hexokinase (Reference method)  ↑ in absorbance is measured at 340 nm  False low results due to gross hemolysis and ↑↑↑ bilirubin Measurement  Point of Care test Measurements  2-Hour Postprandial Tests  A solution (75g of glucose) is administered and a specimen  Oral Glucose Tolerance Test  FBS is taken. Glucose load is administered. Blood glucose is determined in 30 min, 1st, 2nd and 3rd hrs.  HbA1C  Index for long term plasma glucose control (2-3 month period)  Based on charged differences between HbA1C and Non-HbA1C  Specimen requirement is EDTA WB sample. N.V: 4.5 to 8.0% Measurements  HbA1C  Immunoassays  Affinity Chromatography  Cation-Exchange Chromatography  Electrophoresis  Isoelectric Focusing  High performance Liquid Chromatography Ketones  Produced by the liver through metabolism of stored lipids  Acetone (2%)  Acetoacetic acid(20%)  3-β-hydroxybutyric acid (78%)  Ketonemia  accumulation of ketones in blood  Ketonuria  accumulation of ketones in urine Ketones: Measurement  Gerhardt’s Test  Acetoacetic acid + Ferric chloride -> Red color  Nitroprusside  Acetoacetic acid + nitroprusside –alkaline pH-> Purple color  Enzymatic  NADH + H + acetoacetic acid –BHBD -> NAD + B-Hydroxybuturic acid