Medical Technology Board Examination Review Notes Must Know (Rodriguez) 4

Enzymes
Serum á Enzyme concentration = á reaction rate
Reagent If enzyme > substrate, á substrate = á reaction rate
When substrate concentration reaches a maximal value, higher concentration of substrate no longer results in increased rate of reaction
Nonprotein entities
Organic compound Ex. NADP á Coenzyme = á Velocity
Inorganic ions Alters spatial configuration of the enzyme for proper substrate binding Ex. Ca2+ (#1 activator), Zn2+ (LDH), Cl- (AMS), Mg2+ (CK, ALP)
Inorganic ion attached to a molecule Ex. Catalase, cytochrome oxidase
Interferes with the enzymatic reactions
Binds to the active site of an enzyme Reversible (Substrate > Inhibitor)
Binds to the allosteric site (cofactor site) Irreversible
Binds to the enzyme-substrate complex á Substrate = áES = áInhibition
Same catalytic reactions but slightly different molecular structures Fractionation of isoenzymes
37’C = optimum temperature for enzyme activity áTemperature = áReaction rate (ámovement of molecules)
Denaturation of enzymes
Inactivation of enzymes
For every 10OC increase in temperature, there will be a two-fold increase in enzyme activity
Most physiologic reactions occur in the pH range of 7-8
Enzymes: -20’C = for longer period of time Substrate and Coenzymes: 2-8’C LDH (LD4 & 5): Room temperature
Mostly increases enzyme concentration
Decreases enzyme concentration
1st digit: classification 2nd and 3rd digits: subclass 4th digit(s): serial number
“OTHLIL” Oxidoreductases Transferases Hydrolases Lyases Isomerases Ligases
Redox reaction Dehydrogenases: -Cytochrome oxidase -LDH -MDH -Isocitrate dehydrogenase -G-6-PD
Transfer of a chemical group other than hydrogen from 1 substrate to another Kinases, Transaminases, Aminotransferases: -CK -GGT -AST -ALT -OCT
Hydrolysis/splitting by addition of water Esterases: -ACP -ALP -CHS -LPS Peptidases: -Trypsin -Pepsin -LAP Glycosidases: -AMS -Galactosidases
Removal of groups w/o hydrolysis (product contains double bonds) Aldolase Decarboxylases: -Glutamate decarboxylase -Pyruvate decarboxylase -Tryptophan decarboxylase
Intramolecular arrangements Glucose phosphate isomerase Ribose phosphate isomerase
Joining of 2 substrate molecules Synthases
Water-free cavity Where the substrate interacts
Cavity other than the active site May bind regulatory molecules
Coenzyme that is bound tightly to the enzyme
Apoenzyme + Prosthetic group
Inactive form of enzyme
Shape of the key (substrate) must fit into the lock (enzyme)
Based on the substrate binding to the active site of the enzyme Acceptable theory
Enzymes catalyze reactions by lowering the activation energy level that the substrate must reach for the reaction to occur
Enzyme combines w/ only 1 substrate and catalyzes only 1 reaction
Enzymes combine w/ all the substrates in a chemical group
Enzymes reacting w/ specific chemical bonds
Reaction rate depends only on enzyme concentration Independent on substrate concentration
Reaction rate is directly proportional to substrate concentration Independent on enzyme concentration
Change in substrate concentration Change in product concentration Change in coenzyme concentration
1 micromole of substrate/minute
1 mole of substrate/second
Absorbance is made at 10-second intervals for 100 seconds
pH = 10.5 405nm Electrophoresis: (+) Liver ß Bone (Regan) ß Placenta ß Intestine (-) Heat fractionation: (Δ Stable) Regan ß Placenta ß Intestine ß Liver ß Bone (Δ Labile)
Inhibits Regan, placental and intestinal ALP
Inhibits Nagao ALP
Inhibits liver and bone ALP
Inhibits bone ALP
Low temperature = Increased ALP 1. Bowers and McComb (PNPP) – IFCC recommended 2. Bessy, Lowry and Brock (PNPP) 3. Bodansky, Shinowara, Jones, Reinhart = BGP (beta glycerophosphate) 4. King and Armstrong = PP (phenylphosphate) 5. Klein, Babson & Read = Buffered PPP (phenolphthalein phosphate) 6. Huggins and Talalay = PPDP (phenolphthalein diphosphate) 7. Moss = ANP (alpha naphthol phosphate)
Sprue Hyperparathyroidism Rickets (children) and osteomalacia (adults)
pH = 5.5 405nm Sources: Prostate (major), RBC, platelets, bone
Inhibited by L-tartrate ions
Inhibited by cupric and formaldehyde ions
Room temperature (1-2 hrs) = decreased ACP Thymolphthalein monophosphate = specific substrate, substrate of choice (endpoint) Alpha-naphthyl phosphate = preferred for continuous monitoring methods 1. Gutman and Gutman = PP 2. Shinowara = PNPP 3. Babsonm Read and Phillips = ANP (continuous monitoring) 4. Roy and Hillman = Thymolphthalein monophosphate (endpoint)
pH 7.5 340nm Sources: Cardiac tissue > Liver > Skeletal muscle > Kidney, pancreas, RBCs
pH 7.5 340nm Major Source: Liver
1. Karmen method = Kinetic 2. Reitman and Frankel = Endpoint -Color developer: DNPH -Color intensifier: 0.4N NaOH
DeRitis ratio (ALT:AST) >1.0 = Acute hepatitis (Highest) á20x = viral or toxic hepatitis Moderate elevation = chronic hepatitis, hepatic cancer, IM Slight elevation = Hepatic cirrhosis, alcoholic hepatitis, obstructive jaundice
Smallest enzyme (appears in urine) Earliest pancreatic marker P3: most predominant pancreatic AMS isoenzyme in AP Isoenzymes: S-type (ptyalin): anodal P-type (amylopsin): cathodal
Samples w/ high activity of AMS should be diluted w/ NaCl to prev. inactivation Salivary AMS = inhibited by wheat germ lectin Substrate: Starch
Reducing sugars produced Classic reference method (SU)
Degradation of starch
Increase in color intensity
Continuous-monitoring technique
Late marker (AP) Most specific pancreatic marker
Substrate: Olive oil/Triolein 1. Cherry Crandal (Reference method) 2. Tietz and Fiereck 3. Peroxidase coupling (most commonly used method)
Lacks specificity RBC: 150x LDH than in serum Sources: LD1 (α-HBD) and LD2 = Heart, RBC, Kidneys LD3 = pancreas, lungs, spleen LD4 an LD5 = liver and muscle LD6 = alcohol dehydrogenase
1. Wacker method (forward/direct) = pH 8.8, 340 nm, most commonly used 2. Wrobleuski LaDue (reverse/indirect) = pH 7.2, 2x faster 3. Wrobleuski Cabaud 4. Berger Broida
Hepatic carcinoma and toxic hepatitis
Viral hepatitis and cirrhosis
Isoenzymes: CK-BB = most anodal, brain CK-MB = myocardium (20%) CK-MM = least anodal, skeletal and smooth muscles (Major, 94-100%)
Total CK: 50x URL (highest)
Most specific indicator of myocardial damage (AMI) Not elevated in angina
1. Tanzer-Gilbarg (forward/direct) = pH 9.0, 340nm 2. Oliver-Rosalki/ Rosalki & Hess (reverse/indirect) = most commonly used method, faster reaction; pH 6.8, 340nm
Inside RBCs Interferes w/ CK assay Inhibited by adenosine monophosphate
Activate CK
Do not contain CK
Partially restore lost activity of CK
Reference method for CK
CKI (%) = CK-MB/Total CK x 100
Isoenzymes: Aldolase A = Skeletal muscles Aldolase B = WBC, liver, kidney Aldolase C = brain tissue
Marker for hepatobiliary diseases and infiltrative lesions of the liver Methods: 1. Dixon and Purdon 2. Campbell, Belfield and Goldberg
Located in the canaliculi of the hepatic cells Differentates the source of an elevated ALP level Sensitive indicator of occult alcoholism Increased: Obstructive jaundice Alcoholic hepatitis (most sensitive)
Substrate: gamma-glutamyl-p-nitroanilide 1. Szass 2. Rosalki and Tarrow 3. Orlowski
Monitor effects of relaxants (succinylcholine) after surgery Marker for organophosphate poisoning (Low CHS) Methods: 1. Ellman technic 2. Potentiometric
A.k.a. peptidyldipeptidase A or Kininase II Converts angiotensin I à angiotensin II (lungs) Indicator of neuronal dysfunction (Alzheimer’s disease – CSF)
Ferrooxidase enzyme
For hepatobiliary diseases
Drug induced hemolytic anemia (primaquine, antimalarial drug)
ALP = 30-90 U/L ACP: Total ACP (male) = 2.5-11.7 U/L Prostatic ACP = 0-3.5 ng/mL AST = 5-37 U/L ALT = 6-37 U/L AMS = 60-180 SU/dL (95-290 U/L) LPS = 0-1.0 U/mL LDH: Forward = 100-225 U/L Reverse = 80-280 U/L
Acute Myocardial Infarction Markers
Myoglobin	Troponin T	Troponin I	CK-MB	AST	LD
1-3 h	3-4 h	3-6 h	4-8 h	6-8 h	12-24 h
5-12 h	10-24 h	12-18 h	12-24 h	24 h	48-72 h
18-30 h	7 d (10-14 d)	5-10 d	48-72 h	5 d	10-14 d
Acute Pancreatitis Markers
Amylase			Lipase
2-12 h			6 h
24 h			24 h
3-5 d			7 d
Electrolytes
Equal no. of cations and anions Balance of charges
Average water content of the human body
1/3 of total body water
2/3 of total body water
93% water (Plasma: 13% > Whole blood) 7% solutes: (Increased in dehydration) -Proteins -Glucose -NPN -Lipids -Ions
Excretion of 10-20L H2O everyday
Sodium Potassium Chloride
EC = Na+ > Cl- > HCO3- > Ca2+(5th) > iPO4 IC = K+ > Mg2+(4th)
Potassium Calcium Magnesium
Calcium Magnesium (CK) Zinc Chloride (AMS) Potassium
Magnesium
Magnesium Phosphate
Bicarbonate
Magnesium
Major contributor of osmolality (92%, together w/ Chloride and Bicarbonate) á100 mg/dL glucose = â1.6 mmol/L sodium
áSodium âPotassium = â Magnesium
â Sodium
Excess water loss Decreased water intake Hyperaldosteronism (Conn’s disease) Hypothalamic disease (Chronic hypernatremia)
Renal failure SIADH (increased water retention) Marked hemolysis (dilutional effect) <125 mmol/L = severe neuropsychiatric symptoms
Major defense against hyperosmolality and hypernatremia 1-2% water deficit = severe thirst 150-160 mEq/L Na+ = Moderate deficit of water >165 mEq/L Na+ = Severe water deficit
Hyperlipidemia (turbidity) Hyperproteinemia
1. FEP 2. AAS 3. ISE = Glass aluminum silicate 4. Colorimetry = Albanese Lein
Concentration in RBC is 105 mmol/L Reciprocal relationship with H+
0.5% hemolysis = á 0.5 mmol/L Gross hemolysis = á 30% Serum K+ > Plasma K+ by 0.1-0.7 mmol/L  because of platelets (clot) á10-20% in muscle activity á0.3-1.2 mmol/L = mild to moderate exercise á2-3 mmol/L = vigorous exercise; fist clenching
Decreased resting membrane potential à incr. contractility à lack of muscle excitability Decreased renal excretion (Dehydration, renal failure, Addison’s disease) Acidosis (DM) Muscle injury Spironolactone
Increased resting membrane potential  à arrhythmia Leads to hypomagnesemia Vomiting Diuretics Cushing’s syndrome Alkalosis Insulin overdose
â pH by 0.1 = á K+ by 0.2-1.7 mmol/L
Lithium heparin plasma = preferred 1. FEP 2. AAS 3. ISE = Valinomycin gel 4. Colorimetry = Lockhead and Purcell
Chief counter ion of sodium in ECF
Chloride methods measure bromide and iodide âCl- = áHCO3-
1. Schales and Schales: -Mercurimetric titration -Diphenylcarbazone -Excess Hg++ -(+) Blue violet 2. Whiterhorn Titration method -Mercuric thiocyanate -Reddish complex 3. Ferric perchlorate 4. Cotlove chloridometer -Coulometric amperometric titration -Excess Ag++ 5. ISE -Ion exchange membrane -Tri-n-octylpropylammonium chloride decanol
Renal tubular acidosis Metabolic acidosis Diabetes insipidus (Dehydration) Prolonged diarrhea
Prolonged vomiting (âHCl) Aldosterone deficiency (âNa+ = âCl- = áK+) Metabolic alkalosis (áHCO3- = âCl-) Marked hemolysis (dilutional effect)
99% à Bones 1% à ECF Absorbed in the duodenum Absorption is favored at an acidic pH
50% = Free/Ionized/Unbound/Active Calcium 40% = Protein-bound (Albumin) 10% = Complexed with anions
á Ca2+ = á absorption (intestine) and reabsorption (kidney)
á Ca2+ = á resorption (bone) and reabsorption (kidney)
â Ca2+ = á urinary excretion (major net loss of calcium)
Serum = specimen of choice â Albumin (1g/dL) = â Ca2+ (0.8 mg/dL)
Acidosis (Ca2+: from Bones à Blood) Cancer Hyperthyroidism Milk-alkali syndrome
Tetany Alkalosis (Ca2+: from Blood à Bones) Acute pancreatitis (Ca2+: binds to damage pancreatic tissues)
Low PTH Parathyroid gland disease
High PTH Renal failure (á excretion)
1. Clark Collip precipitation method -(+) Oxalic acid -Renal calculi 2. Ferro Ham Chloranilic acid precipitation method -(+)Chloranilic acid 3. Colorimetric = Ortho-Cresolphthalein complexone dyes -Dye: Arzeno III -8-hydroxyquinoline = chelates (inhibits) Mg2+ 4. EDTA titration method (Bachra, Dawer and Sobel) 5. AAS = Reference method 6. ISE = Liquid membrane 7. FEP
85% à Bones 15% à ECF (iPO4) Maximally absorbed in the jejunum (Ca2+: duodenum) Trancellular shift: Once absorbed inside cells, it no longer comes out à used for energy production Dirunal variation: á late morning, â evening Organic phosphate = principal anion within cells Inorganic phosphate = part of the blood buffer (Measured in the clin.lab.)
55% = Free 35% = Complexed with ions 10% = Protein-bound
â PO4 = á Ca2+
á PO4 = â Ca2+
á PO4 (renal reabsorption)
Fasting is required (Nonfasting: â PO4)
Hypoparathyroidism Renal failure Hypervitaminosis D
Alcohol abuse = most common cause Primary hyperparathyroidism Avitaminosis D (Rickets, Osteomalacia)
Most accurate: unreduced phosphomolybdate formation (340nm) 1. Fiske Subbarow Method (Ammonium molybdate method) -Reducing agents: Pictol, Elon, Senidine, Ascorbic acid -(+) Phosphomolybdenum blue
53% à Bones 46% à Muscles and soft tissues 1% à Serum and RBC Vasodilator
55% = Free/Ionized/Physiologically active 30% = Protein-bound 10% = Complexed with ions
áMg2+ = á Ca2+ = â PO4
âMg2+ = â K+ = á Na+
Addison’s disease Chronic renal failure
Acute renal failure Chronic alcoholism
1. Calmagite -(+) Reddish-violet complex 2. Formazen dye method -(+) Colored complex 3. Magnesium Thymol blue method -(+) Colored complex 4. AAS = reference method 5. Dye-lake Method -Titan Yellow dye (Clayton Yellow or Thiazole yellow)
90% of the total CO2
HCO3- diffuses out of the cell in exchange for Cl- to maintain ionic charge neutrality w/in the cell
Difference between unmeasured anions and unmeasured cations QC for ISE
Uremia/renal failure Ketoacidosis Lactic acidosis Methanol poisoning Ethanol poisoning Ethylene glycol poisoning Salicylate poisoning
Hypoalbuminemia Hypercalcemia Hyperlipidemia Multiple myeloma
Defective gene: Cystic fibrosis transmembranous conductance regulator (Chromosome 7) Miconeum ileus (Infants) Foul-smelling stool URT infection á Na+ and Cl-
Sweat inducer
Reference method (Sweat sodium and chloride)
Prooxidant 3-5g = Total body iron Ferrous = Hgb Ferric = Transferrin and Ferritin
1. Colorimetric = HCl and Ferrozine -(+) Blue color 2. Anodic stripping voltammetry
Hemochromatosis Viral hepatitis Non-IDA
IDA Malnutrition Chronic infection
UIBC + Serum Iron Increased: IDA, hepatitis, iron-supplemented pregnancy Decreased: Non-IDA, nephrosis
TIBC – Serum iron Measure of reserve iron binding capacity of transferrin
Index of iron storage Increased: Iron overdose, hemochromatosis, sideroblastic anemia Decreased: IDA (lowest), malignancy, chronic infection
TIBC (μg/dL) x 0.70 = mg/dL
Sodium 1/α Potassium Potassium 1/α Hydrogen ion Potassium α Magnesium Magnesium α Calcium Calcium 1/α Inorganic phosphate Chloride 1/α Bicarbonate
Sodium: Serum = 135-145 mmol/L [Critical: 160 mmol/L and 120 mmol/L] CSF = 136-150 mmol/L Potassium: Serum = 3.5-5.2 mmol/L [Critical: 6.5 mmol/L and 2.5 mmol/L] Chloride: Serum = 98-107 mmol/L Sweat = 5-40 mmol/L [Critical: >65 mmol/L] Calcium: Total = 8.6-10 mg/dL (adult) and 8.8-10.8 mg/dL (child) Ionized = 4.6-5.3 mg/dL (adult) and 4.8-5.5 mg/dL (child) [Critical: <7.5 mg/dL] Inorganic Phosphate: Adult = 2.7-4.5 mg/dL Child = 4.5-5.5 mg/dL Magnesium: Serum = 1.2-2.1 mEq/L Anion Gap: w/ K+ = 10-20 mmol/L w/o K+ = 7-16 mmol/L Iron: Male = 50-160 μg/dL Female = 45-150 μg/dL TIBC: Adult = 245-425 μg/dL >40 y.o. = 10-250 μg/dL NB and Child = 100-200 μg/dL % Transferrin Saturation = 20-50%