CC 1 LEC-MIDTERMS - Carbohydrates PDF

## CC LEC-MIDTERMS - Google Drive ### Carbohydrates - **These are compounds containing C, H, and O.** - **General formula for a carbohydrate is Cx(H2O)y.** - **All carbohydrates contain C=O and -OH functional groups.** - **Derivatives with addition of other chemical groups, such as phosphates, sulfa...

## CC LEC-MIDTERMS - Google Drive ### Carbohydrates - **These are compounds containing C, H, and O.** - **General formula for a carbohydrate is Cx(H2O)y.** - **All carbohydrates contain C=O and -OH functional groups.** - **Derivatives with addition of other chemical groups, such as phosphates, sulfates, and amines.** #### Models: - Fischer - Haworth ### Classification of carbohydrates is based: 1. **The size of the base carbon chain** - **Trioses:** with three (3) carbons - **Tetroses:** with four (4) carbons - **Pentoses:** with five (5) carbons - **Hexoses:** with six (6) carbons 2. **The location of the CO functional group** - **Aldose:** has a terminal carbonyl group (O=CH-) called an aldehyde group - **Ketose:** has group carbonyl group (O=CH-) in the middle linked to two other carbon atoms called a ketone group 3. **Stereochemistry of the compound** - **Stereoisomers:** have the same order and types of bonds but different spatial arrangements and different properties - **Enantiomers:** images that cannot be overlapped and are non-superimposable - L-isomer - D-isomer 4. **Number of sugar units** - **Monosaccharides** - Simple sugars that cannot be hydrolyzed to simpler form - Examples: glucose, fructose, galactose - **Disaccharides** - Formed by two monosaccharides joined by glycosidic linkage - Hydrolyzed by disaccharide enzymes (i.e., lactase) produced by the microvilli of the intestine - Examples: - Maltose = glucose + glucose - Lactose = glucose + galactose - Sucrose = glucose + fructose - **Oligosaccharides** - Chaining of 2 to 10 sugar units - **Polysaccharides** - Linkage of many monosaccharide units - Yield more than 10 monosaccharides upon hydrolysis - Examples: starch, glycogen ### Chemical properties 1. **Reducing substances** - Contain a ketone or aldehyde group - **With free anomeric carbon** - Can reduce other compounds - Examples: glucose, maltose, fructose, lactose, galactose 2. **Non-reducing Substances** - Do not have an active ketone or aldehyde group - **No free anomeric carbon** - Will not reduce other compounds - Example: sucrose (table sugar) ### Glucose and its metabolism - End product of carbohydrate digestion in the intestine - Enzymes involved: - **Amylase** (salivary & pancreatic) - digests nonabsorbable polymers to dextrins and disaccharides - **Maltase** (from the intestine) - digests disaccharides to monosaccharides - **Sucrase & Lactase** - hydrolyze sucrose & lactose respectively - Functions - Provides energy for life processes - The only CHO that can be directly used for energy or stored as glycogen - Forms: ~35% alpha & 65% beta ### Major metabolic pathways 1. **Embden-Meyerhoff or glycolysis** - Substrate: D-glucose - **End-products:** 2 moles of PYRUVIC ACID, 2 moles NADH and 2 moles of ATP - Can occur aerobically or anaerobically. If aerobic, pyruvate is formed. If anaerobic, lactate is formed. - Other substrates can enter this pathway at various points - Glycerol (from TAG) enters at 3-phosphoglycerate - Fatty acids, ketones and some amino acids are converted to acetyl-CoA - Other amino acids enter as pyruvates or as deaminated a-ketoacids and a-oxoacids ### Inborn errors of carbohydrates metabolism 1. **Galactosemia** - a cause of failure to thrive syndrome in infants; - congenital deficiency of one of three enzymes involved in galactose metabolism, resulting in increased plasma galactose levels. - Galactose-1-phosphate uridyl transferase - MOST COMMON enzyme deficiency - Fructose-1-phosphate aldolase deficiency 2. **Essential fructosuria** - An autosomal recessive disorder characterized by fructokinase deficiency. - Fructokinase catalyzes the conversion of fructose to fructose-1-phosphate. - **Diagnostic indicator:** the presence of fructose in urine (Fructosuria) 3. **Hereditary fructose intolerance** - A defect of fructose 1-6-biphosphate aldolase activity in the liver, kidney and intestine. - Clinical features: irritability, lethargy, seizures and hepatomegaly. 4. **Fructose-1,6-biphosphate deficiency** - A defect in fructose-1,6-biphosphate results in failure of hepatic glucose generation by gluconeogenic precursors such as lactate and glycerol. - Clinical features: hypoglycemia, lactic acidosis, convulsions and coma. 5. **Glycogen storage diseases** - deficiency of a specific enzyme that causes alteration of glycogen metabolism - **von Gierke's dse (Type I)** - Enzyme Deficient: Glucose-6-phosphatase - Clinical Features: Severe fasting hypoglycemia, Lactic acidosis, Accumulation of amount of glycogen on all organs, Presence of abnormally LARGE LYSOSOMES - **Pompe's dse (Type II)** - Enzyme Deficient: a-1,4-glucosidase - Clinical Features: Hypoglycemia, hepatomegaly, seizures and mental retardation - **Forbe's dse (Type III)** - Enzyme Deficient: Debrancher enzyme - Clinical Features: Progressive liver enlargement or cirrhosis and muscular weakness by age 2 - **Andersen's dse (Type IV)** - Enzyme Deficient: Brancher enzyme - Clinical Features: Absence of storage glycogen. Unbranched AMYLOPECTIN ### Specimen collection and handling - Glucose levels in whole blood are about 15% lower than in serum/plasma. - Whole blood glucose decreases by roughly 10 mg/dL per hour. - Separate serum/plasma within 1 hour (or 30 minutes ideally) to prevent glucose loss. - Glucose is metabolized at 7 mg/dL/h at room temperature and 2 mg/dL/h at 4°C. - Refrigerated serum/plasma is stable for up to 48 hours. - Sodium fluoride preserves glucose for up to 48 hours. - Glycolosis reduces serum glucose by 5-7% per hour in uncentrifuged blood. - Fasting blood glucose requires a 10-16 hour fast. ### Hypoglycemia - Decrease in plasma glucose levels: 65-70 mg/dl (3.6-3.9 mmol/L) - plasma glucose concentration at which glucagon and other glycemic factors are released - It results from an imbalance between glucose utilization and production - 50-55 mg/dl (2.8-3.0 mmol/L) - symptoms of hypoglycemia appear - Warning S/S are all related to CNS #### Symptoms of hypoglycemia - **Neurogenic/adrenergic** - tremors, palpitations, anxiety, diaphoresis - **Neuroglycopenic** - dizziness, tingling, blurred vision, confusion, behavioral changes. ### Diagnostic tests for hypoglycemia - 72 hours fast which requires the analysis of glucose insulin, C-Peptide and proinsulin at 6-hour intervals - **Positive result:** >45mg/dL; hypoglycemic - Symptoms appear alter 72 hours had elapsed. ### Diabetes mellitus #### Secondary diabetes mellitus - associated with secondary conditions - Genetic defects of ẞ-cell function. - Pancreatic disease - Endocrine disease - Cushing syndrome - excessive cortisol - Pheochromocytoma epinephrine excess - Acromegaly - growth hormone excess - Drug or chemical induced - Insulin receptor abnormalities - Other genetic syndromes - Maturity onset diabetes of youth (MODY) - rare; autosomal dominant #### Gestational diabetes mellitus (GDM) - Any degree of glucose intolerance with onset or first recognition during pregnancy. - Due to metabolic or hormonal changes - Infants born to mothers with this kind of diabetes are at increased risk to respiratory distress syndrome, hypocalcemia & hyperbilirubinemia #### Laboratory findings in hyperglycemia - Increased glucose (plasma & urine), urine specific gravity, serum and urine osmolality - Ketonemia and ketonuria - Decreased blood and urine pH (acidosis) - Electrolyte imbalance (Na+, Cl-, HCO3-, and K+) #### Diagnostic criteria for diabetes mellitus - RPG ≥200 mg/dl (11.1 mmol/L) + symptoms of diabetes - Fasting PG ≥126 mg/dL (7.0 mmol/L) - 2-h PG ≥200 mg/dl (11.1 mmol/L) during OGTT #### Categories of fasting plasma glucose - Normal Fasting glucose FPG <110mg/dL - Impaired fasting glucose FPG ≥110mg/dL but <126 mg/dL - Provisional diabetes dx FPG ≥ 126 mg/dL ### Tests for carbohydrate disorders #### Diagnostic tests - **Fasting blood sugar** - Normal-70-110 mg/dl - Diabetes ->126 mg/dl - **2hr Post prandial blood sugar (PPBS)** - Normal - <126 mg/dl - Diabetes ->200 mg/dl - **Post-loading glucose** - Similar to PPBS - *Glucose load is standardized - *Diabetics ≥200 mg/dl #### Tests for monitoring - **Glycosylated hemoglobin** - Assessment of long-term control - Average glucose level over 60 days (2-3 months) - **Microalbuin** - Detects small amounts of protein in urine of diabetic patients to assess renal damage - **C peptide of insulin (reflects pancreatic insulin secretion)** - Normal 1:1 (insulin: C-peptide) - Diabetes > 1:1 - C-Peptide after insulin injection ### Methods of HbA1c measurement #### Methods based on structural differences - **Immunoassays** - Polyclonal or monoclonal antibodies toward the glycated n-terminal group of the ẞ chain of Hgb - **Affinity chromatography** - Separates based on chemical structure using borate to bind glycosylated proteins - Not affected by temperature and other hemoglobins ### Methods for analysis #### Chemical - **Cupric ion Reduction** - **Folin-Wu** - measure of ALL REDUCING SUBSTANCES in the blood - Reagent that binds with Cu+: phosphomolybdic acid - End product: phosphomolybdenum blue - End color: blue - **Nelson Somogyi - MEASURE OF TRUE GLUCOSE** - Reagent that binds with Cu+: arsenomolybdic acid - End product: arsenomolybdenum blue - End color: blue - **Neocuproine** - Reagent that binds with Cu+: neocuproine - End product: cuprous-neocuproine complex - End color: yellow/yellow orange - **Ferric ion Reduction - Inverse Colorimetry** - reduction of yellow ferricyanide to a colorless ferrocyanide by glucose - **Hagedorn Jensen** #### Condensation - **Orthotoluidine (Dubowski method)**: can be used for urine and CSF without protein precipitation - Absorbance: 630 nm - Reagent: aromatic amine, glacial acetic acid - End color: green - Interfering substances: galactose and mannose - **Enzymatic**: Acts on glucose but not on other sugars and not on other reducing substances. - **Glucose Oxidase Method**: It also measures CSF glucose - Disadvantages: This reaction is inhibited by high concentrations of uric acid, vitamin C, bilirubin, glutathione, creatinine, dopamine, methyldopa, and citric acid - **Colorimetric glucose Oxidase Method** - **Polarographic Glucose Oxidase** - measures oxygen consumption with PO2 electrode (Clark) - Molybdate catalyzes the oxidation of iodide to iodine by H2O2 - Catalase-catalyzes oxidation of ethanol by H2O2 forming acetaldehyde and H₂O #### Hexokinase - Generally accepted as the REFERENCE METHOD - MORE ACCURATE THAN OXIDASE - Coupling reaction using G6PD is highly specific - Measured by quantitating reduced NADPH formation - NADPH is measured directly at 340 nm or coupled to chromogen and measured in visible range - Interfering substances: gross hemolysis & extremely elevated bilirubin (cause ↑values) - May be performed using serum or plasma (heparin, EDTA, fluoride, oxalate & citrate) - Excellent for glucose determination in urine, CSF and serous fluids #### Other methods: - **Glucose Dehydrogenase Method:** The amount of NADH generated is proportional to the glucose concentration - **Dextrostics (cellular strip):** Important in establishing correct insulin amount for next dose. Effective in reducing the rate of development of diabetic complications ### Other important tests #### Ketones - Produced by the liver through metabolism of fatty acids to provide ready energy source from stored lipids at times of low carbohydrate availability - Three ketone bodies: - Acetone (2%) - Acetoacetic acid (20%) - β-hydroxybutyric acid (78%) - Causes of increased ketone levels: - Diabetes Mellitus - Starvation/fasting - High-fat diets - Prolonged vomiting - Glycogen storage diseases - **Ketonemia:** accumulation of ketones in the blood - **Ketonuria:** accumulation of ketones in the urine ### Measurement of ketones - For patients with Type 1 Diabetes, it is recommended during acute illness, stress, pregnancy, or elevated blood glucose levels above 300 mg/dL or when patients have signs of ketoacidosis - Specimen: FRESH **SERUM** or **URINE** tightly stoppered and analyzed immediately. ### Methods for ketone analysis #### Chemical test - **Gerhardt's test** - historical test: - Used FERRIC CHLORIDE reacted with ACETOACETIC ACID to produce a RED color - Reacts only with acetoacetate - **Sodium Nitroprusside** - more common method - Uses **SODIUM NITROPRUSSIDE** which reacts with **ACETOACETIC ACID** in an **ALKALINE pH** to form a **PURPLE COLOR** - If **GLYCERIN** is also added, **ACETONE** will be detected - Used in urine reagent strips and Acetest tablets - 10x more sensitive to acetoacetate than to acetone