Lab 9 - Bilirubin Metabolism Flashcards
Br I
Indirect, bound to albumin, unconjugated, some get into blood
BrII
Direct, bound to glucuronic acid=conjugated, a little amount get into blood
Metabolism of bile acids
1️⃣Br+albumin in blood make brI and goes into liver
2️⃣br+glucuronic acid in liver make brII and is exreted into bile
3️⃣bacteria reduce br to urobilinofen which is either further red tot stercobilin or UBG is absorbed into blood(small molecule!) to the ileum
4️⃣from the ileum it will either be absorbed and go to the kidneys to be exreted, or to the liver for urea production
serum bilirubin measurement. Normal value
diazo method: Spectrophotometroc method. With SERUM sample
Tot br: caffeine/alcohol + diaso + NaCl
Direct br: diaso + NaCl
Tot Br= 8 μmol/l, horse: < 40 μmol/l (bc no gallbladder for storage - circ storage instead!)
Causes of incr Indirect br in the plasma
acute haemolysis
absorption of haemoglobin following massive internal haemorrhage, or after big
haematoma formation (so called ”resorption icterus”)
transfusion of stored blood, which contains many dyeing or dead RBCs
Decreased rate of conjugation of Br I by liver cells
i Decreased uptake of Br I from the blood by liver cells
Causes of incr Direct br in the plasma
(BrII) Ine Dykker Ikke
1️⃣Incr production after intravascular hemolysis
2️⃣decr excretion bc impaired hepatic function
3️⃣Impaired hepatic function due to obstruction of bile canal: swelling, inflammation, fibrosis, cells swell, blockage, compression
Measurement of bilirubin in the urine
Gmelin test ((br)biliverdin, UBG) Ehrlich test (UBG)
Diazo test strip (Br)
🍏Gmelin test
Nitric acid has to be carefully layered under urine in a test tube, and the width of the differently coloured layers at the meeting of the two fluid phases has to be evaluated.
condensed material on the surface of the glass tube - acidic urea, cats!
yellow - urine itself
white (opaque) – protein (ok in eq, otherwise kidney, cystitis(bladder), PLN
purple - indicane (indol-sulphate) ok in eq, otherwise show bacterial overgrowth - catabolism
🔺 green – biliverdin ok in dog as they have enzyme, path in cat, others
🔺 brown - urobilinogen (UBG) ok in all animals, missing in posthepatic icterus!
HNO3
Gmelin result for healthy animal: exam Q
condensed material on the surface of the glass tube - acidic urea (🐱) 🔺 yellow - urine itself white (opaque) – protein (🐴) purple - indicane (indol-sulphate)🐴 green – biliverdin 🐶 🔺 brown - urobilinogen (UBG) 🔺 HNO3
Indocane
Indicane is a sulphated derivate of indol which is produced by bacteria in the lumen of the large intestines (colon and caecum) and it is absorbed to the blood and filtrated by the kidneys and finally excreted in the urine. When it is oxidised (i.e. by HNO3) it becomes purple. Appearance of indicane in the urine is normal in horses but in carnivores it indicates pathological enteral catabolism (enteritis, constipation).
Ehrlich test
some drops of Ehrlich reagent in urine sample: checks UBG
Normal is mildly red from above and no color from the side
Abnormal is intensly red from above, mild/intense red from the side, and color change of the urine in general
Prehepatic jaundice, how it works, three causes
Haemolytic crisis, free hgb, all br are incr in blood.
Hemolysis, Br ➡️ Liver ➡️ bile to feces and urine(portal vein)
🔺 babesiosis
🔺 immune hemolytic anemia
🔺 viper venom (snake)
Hepatoc jaundice, disease, how it works
Liver damage - all br in blood are incr
Normal amount of hemolysis, Br ➡️ 20% taken up by liver, rest has to take extra rounds in blood(incr conc) ➡️ 60% is turned into BrII in liverm only 20% to bile duct, 40% back to blood(bile mix with blood!!)➡️ little BrII in feces, hypochromic. Since so much in blood ⬆️⬆️ but only some frome intestines ⬆️ UBG
Only 20% taken up - chirrosis! 80% dysfunction parenchyma
Posthepatic jaundice, disease, how it works
Bile duct obstruction
Normal amount of hemolysis, Br ➡️ liver will transform to BrII, but then into blood bc duct obstruction so ⬆️⬆️⬆️ BrII but no UBG in blood!
🔺Pancreatitis is the most typical reason as it will squeese the bile duct 🔺Cholestasis: small/large, intra/extra hepatic duct obstruction - diplomatic term!
🍏Lab result for prehepatic jaundice in blood
🔺Br I: ⬆️⬆️ hemolysis
🔺Br II: ⬆️ most stay in liver/bile
🔺UBG (not measured) ⬆️⬆️ intense colored feces
🔺Free haemoglobin (Hb) ↔️⬇️
🔺Haptoglobin cc. ⬇️
🔺Ht⬇️
🔺Reticulocyte count ⬆️⬆️⬆️ after some days, hemolysis!
🔺Coagulation parameters (PT, APTT, TT): ↔️⬆️ doesnt really change it, but if DIC present its bad!!
🔺Characteristic alterations in enzyme activities ⬆️ ALT/LDH: some from RBC
🔺Characteristic alterations in substrate concentrations: ⬆️ urea, liver catabolise globin to NH3 to urea
🍏Lab result for hepatic jaundice in blood
🔺Br I: ⬆️ normal amount hemolysis, but extra rounds
🔺Br II: ⬆️⬆️ most go to blood not bile
🔺UBG (not measured) ⬆️pale feces
👎🔺Free haemoglobin (Hb) ↔️⬇️
🔺Haptoglobin cc. ↔️
👎 🔺Ht⬇️↔️
(👎 complicated disease - bile acids in blood!)
👎 🔺Reticulocyte count ↔️⬇️
🔺Coagulation parameters (PT, APTT, TT): ⬆️⬆️⬆️ vit K give no change bc no coag factors bc liver no work
🔺Characteristic alterations in enzyme activities ⬆️ ALT/AST/GLDH liver parench enzymes
🔺Characteristic alterations in substrate concentrations: ⬆️ NH3, ⬇️urea, ⬆️ bile accids
🍏Lab result for posthepatic jaundice in blood
🔺Br I: ↔️ normal
🔺Br II: ⬆️⬆️⬆️ accumulation!
🔺UBG (not measured) - missing
🔺Free haemoglobin (Hb) ↔️
🔺Haptoglobin cc. ↔️
🔺Ht↔️
🔺Reticulocyte count ↔️
🔺Coagulation parameters (PT, APTT, TT): ⬆️⬆️ improves after vit K bc factors are present
🔺Characteristic alterations in enzyme activities ⬆️ ALKP, GGT bile obstruction enzymes
🔺Characteristic alterations in substrate concentrations: ⬆️ bile acids
🍏Bilirubin in feces
Incr direct(2) excretion incr ubg and then stercobilin which have an orange color
Hyperchromic: intense color due to hemolysis - prehepatic jaundice
Hypocholic:
light brown: liver failure, less direct(2) br (hepatic jaundice)
light gray, acholic: bile duct obstruction -(posthepatic jaundice)
🍏Urine parameters of Br, UBG and hgb in pre hepatic, hepatic, posthepatic jaundice
Prehep, hep, posthep:
Br: ⬆️, ⬆️⬆️, ⬆️⬆️⬆️ (brII)
UBG: ⬆️⬆️, ⬆️, ⬇️↔️ (missing)
Hgb: ⬆️⬆️⬆️, -, -
Tests for liver ability to conjugate and then excrete
1️⃣Brom-sulphalein- (sulphobromtalein-) (BSP) retention test
2️⃣ Measurement of BSP half life
3️⃣ Indocyane green (ICG) retention test
BSP
Brom-sulphalein retention test
- give function of UDP glucuronyl transferase activity:
- bromsulphalin IV admin - taken up by hepatocytes who splits it and conjugates with glucuronic acid - into bile
Sampling procedure: BSP
1️⃣ Blood sampling for basal value (blank)
2️⃣ Administration of BSP iv (5 mg/kg body weight in 5% solution)
3️⃣ Blood sampling after 3 minutes for 100% value
4️⃣Blood sampling after 27 minutes (30 minutes after BSP administration) for the %-age in question.
Result of BSP
Centrifuge and mix plasma with NaOH as BSP turns purple in alkaline environment! If its not conjugated it will be purple, spectrophotometry
- basal measurement is blank
- 3min sample as 100%
so liver dysfunction if more purple
- Retention of the BSP in plasma should be 5-10% in dog, 3-5% in cat (so around 95% is taken up by the liver)
BSP halflife
Used in large animals bc its expensive. Measure every 5 mins until 45. Do it in a healthy animal first to make standard curve to use in sick animal measurement.
