is an RBC enzyme - decreased levels are signigicant - deficiency in hemolytic anemia, anti-malarial drug promaquine - use blood

metabilized by the liver - useful in accessing liver disease in adults - distinct isoezymes found in liver and bone

- decrease levels are significant - used to monitor pesticide exposure - decreased in liver disease - prolonged apnea

- creatinine kinase - aldolase - LDH

- ALT - AST - ALP - GGT and 5' nucleotidease

ENZYMES Flashcards by Emily Treadwell

factors that influence and enzyme reaction

substrate concentration
enzyme concentration
pH
temperature
co-factors
inhibitors

How well did you know this?

Not at all

Perfectly

Two general methods used to measure an enzyme activity

fixed time

- continuous-monitoring or kinetic assay

How well did you know this?

Not at all

Perfectly

fixed time measurements

reactants ( and pt sample) are combined
RXN proceeds for a defined period of time
rxn is stopped(inactivate the enzyme)
measurement of some chromogen is made
RXN assumed to be linear over measurment time, so the greater the rxn, the more enzyme present
non linear may occur
- at high activities with substrate depletion
- during lag phase which may vary from sample

How well did you know this?

Not at all

Perfectly

Continuous-monitoring

multiple measurements during reaction

usually absorbance changes
specific intervals or continuously
advantages
- linearity of the reaction can be adequate verified
- background color has minimal effect since a change is abs is measured
- can recognize and compensate for lag phase

How well did you know this?

Not at all

Perfectly

How to measure macramylasemia

by precipitating with a high MW polymer and remeasuring or measure urine amylase. If serum is elevated but urine is normal, suggestive if macroamylase.

How well did you know this?

Not at all

Perfectly

specificity of Enzymes used to assess acute pancreatitis

increase amylase and Lipase

- lipase is more clinically specific

How well did you know this?

Not at all

Perfectly

G-6-PDH

is an RBC enzyme

decreased levels are signigicant
deficiency in hemolytic anemia, anti-malarial drug promaquine
use blood

How well did you know this?

Not at all

Perfectly

GGT in liver vs bone

metabilized by the liver

useful in accessing liver disease in adults
distinct isoezymes found in liver and bone

How well did you know this?

Not at all

Perfectly

Enzyme most sensitive alcohol intake

GGT

- used to monitor chronic alcohol abuse

How well did you know this?

Not at all

Perfectly

Applications for acid phosphatase

historically used as a prostate cancer marker

- used for an indication of late stage prostate cancer

How well did you know this?

Not at all

Perfectly

pseudocholinesterase

decrease levels are significant
used to monitor pesticide exposure
decreased in liver disease
prolonged apnea

How well did you know this?

Not at all

Perfectly

muscle enzymes

creatinine kinase
aldolase
LDH

How well did you know this?

Not at all

Perfectly

Liver Enzymes

ALT
AST
ALP
GGT and 5’ nucleotidease

How well did you know this?

Not at all

Perfectly

Pancreatic Enzymes

amylase

lipase

How well did you know this?

Not at all

Perfectly

Miscellaneous enzymes

acid phosphatase
cholinesterase
Gluc-6-Phose dehydrogenase

How well did you know this?

Not at all

Perfectly

Creatine Kinase

-skeletal/heart muscle and brain tissue
- normal less than 200 F, less than 300 M
- elevated AMI, Duchenne muscular dystrophy
- seen in muscle wasting disease
- hypothryroidism, polymyotisis
- flourescent detection of CK
-measures with a 3 step-coupled- reaction
- has lag phase
- NADPH is flourescent
- CK is unstable due to to disulfide bonds
-

CK isozymes

bb= brain- fast moving
mb= heart- intermediate
mm- skeletal muscle

Aldolase

muscle enzyme occas. with CK

not assayed in many labs
reference ranges vary (age dependent)
very sensitive to hemolysis due to high aldolase in RBCs
2-step reaction causing involving NADH using Fructose-1-6-phosphate

LD or LDH

increased in many disorders
- liver disease, muscle trauma, renal infart, hemolytic disease, pernicious anemia
-in MI “flipped isoenzyme”
- increased longer than CK in a MI
has 4 subunits
- assays are hemolysis sensitive
- shaking and partially filled tube problems
-assay has a forward reaction that results in NADH
- reverse reaction is 3x more sensitive to

LD isoenzyme patterns in disease states

MI: “ flipped” LD1>LD2
LD5 elevated skel. muscle or liver disease
central fractions elevated:
- lung sources or possibly some malignancy
- central fraction elevations usually not diagnositc
- all fractions elevated: shock, sepicemia

AST

liver, heart, skeletal muscle, RBC

primarily used for diagnosis of liver disorders
elevated in MI
increased in 24-48 hours
measured with an AST assay- measures absorbance decrease at 340 nm
pryridoxal-5-phosphate cofactor

Isoenzymes of LD

LD1-HHHH-heart,RBC, kidney
LD2-MHHH- heart
LD3- MMHH- lung
LD4-MMMH- many tissues
LD5- MMMM- skeletal muscle and liver

Diagnosis of an MI

MI first CK increases, then AST and then LD

ALT

found in the liver with lesser amount in the heart and skeletal muscle

used to diagnose liver disease
less sensitive to hemolysis than AST
measured with a couples reaction involving NADH
both ALT and AST are included with a standardized pane; (Comprehensive Metabolic Profile)

Alkaline Phosphate (ALP)

routinely included in chemistry panels - found bound to cell membranes - found in the liver, placenta and bone - has isoenzymes found in liver, bone placenta and intestine. 45% liver, 45% bone, 5% intestine

Elevated ALP

- cholestasis/obstructive liver disease - bone disease: osteoblastic increased in Pagets disease - intestinal infarction - placenta -like ALP- tumor marker, malignacy. third trimester in pregnancy

Fractionation of ALP isoenzymes

- electrophoresis in liver,bone,intestinal and placental fractions visualized - heat stablility: 56 deg for 10 mins - placenta: heat stable - bone heat liable - liver/intestinal: intermediate stability

Measuring of ALP

P-nitrophenol absorbs at 405- 415 - Mg is a cofactor - not hemolysis sensitive

GGT

- measure rate reaction using substrate which liberates p-nitrophenol - not hemolysis sensitive - elevated in billiary tract obstruction, pancreatic cancer - useful in liver disease - drugs metabolized in the liver may elevate GGT

5' nucleotidase

clinically significant in the same manner in GGT - elevated in liver and billiary tract - more difficult to assay - helpful in differentiating an elevated ALP whether due to liver or bone disease

Amylase

breaks down starches into simpler sugars and maltose - low MW enzyme- present in both serum and urine - found in Saliva and Panreas and give rise to S-type and P-type isozymes - used to diagnose acute pancreatitis -rises after attack, peaks at 24 hr - decreases gradually OTHER: - elevated in parotid gland disorders(mumps), renal insufficency, DM due to salivary isoenzyme - elevated in Macroamylsemia -

Amylase assys

amyloclastic: starch and iodine reaction saccharogenic: sugars produced upon action of amy-on starch - serum stable at room temp - requires chloride ion - urine enzyme liable at acidic pH of urine - ration urine amylase to urine creatinine using spot urine

Lipase

Hydrolyzes fats to produce fatty acids and glycerol - used to diagnose acute pancreatitis - greater clinical specificity to acute pancreatitis than amylase - amylase and lipase frequently ordered together

Acute pancreatitis

increased Amylase and lipase

Acid Phosphate

usually measures same reaction as ALP but assay at acidic - stabilize at acid pH - greatest concentrations in prostate, liver, spleen and bone - high levels used to detect semen in suspected rape cases(vaginal washings)

Cholinesterases

decreased levels are significant - RBC enzymes known as acetylcholinesterase make hemolysate and assay : used for monitoring pesticide exposure - Pseudocholinesterase(serum/plasma) - pesticide exposure