Biological Chemistry - Berezov, T. T., Korovkin, B. F. 1998

Metabolism of Complex Proteins
Chromoprotein Metabolism
Hemoglobin Breakdown in Tissues (Formation of Bile Pigments)

The lifespan of erythrocytes is 120 days, after which they break down and Hemoglobin is released. The primary Organs responsible for erythrocyte destruction and hemoglobin degradation are the Liver, Spleen, and Bone Marrow, although both processes can theoretically occur in Cells of other organs as well. Hemoglobin breakdown in the liver begins with the Cleavage of the α-methine bridge between the First and Second rings of the porphyrin ring system. This process is catalyzed by an NADPH-dependent oxidase, resulting in The formation of the green pigment verdoglobin (choleglobin):

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In the structural formulas presented here and hereafter for Bile pigments, M denotes a methyl group (CH₃), V denotes a vinyl group (—CH=CH₂), and P denotes a propionic acid residue (—CH₂—CH₂—COOH).

As can be seen from the formulas above, the iron atom and the protein component are still retained in the verdoglobin molecule. There is experimental evidence that Vitamin C, Fe²⁺ ions, and other Cofactors take part in this oxidative transformation of hemoglobin. Further degradation of verdoglobin most likely occurs spontaneously with the release of iron and the globin protein, yielding biliverdin, one of the bile pigments. This spontaneous breakdown is accompanied by the redistribution of double bonds and hydrogen atoms within the pyrrole rings and methine bridges. The resulting biliverdin is then enzymatically reduced in the liver to bilirubin, which is the principal bile pigment in humans and carnivorous animals:

The primary sites of bilirubin formation are the liver, spleen, and apparently erythrocytes (arising from The breakdown of red Blood Cells, which sometimes involves the cleavage of one of the methene bridges in protoporphyrin). Bilirubin produced in all these cells is transported to the liver, from where it enters the Gallbladder along with bile (see Chapter 16). Bilirubin formed in the Cells of the macrophage system is referred to as free or indirect bilirubin; because of its poor Water solubility, it readily adsorbs to Plasma Proteins, and its determination in blood requires the prior precipitation of proteins with alcohol. Following this, bilirubin reacts with Ehrlich's diazo reagent.

The blood of a healthy adult contains a relatively constant level of total bilirubin, ranging from 4 to 26 μmol/L, with an average of 15 μmol/L. Approximately 75% of this amount is accounted for by indirect bilirubin. An increase in its blood concentration to 35 μmol/L leads to jaundice. Higher blood levels of bilirubin cause severe toxic effects. Upon entering the liver via the bloodstream, indirect bilirubin undergoes detoxification through conjugation with glucuronic acid. This process involves a specific enzyme, UDP-glucuronosyltransferase, and UDP-glucuronic acid, which serves as the glucuronic acid donor. Two glucuronic acid residues attach to bilirubin to form a relatively inert complex, bilirubin diglucuronide, which is highly water-soluble and yields a direct reaction with the diazo reagent. Direct bilirubin is always present in bile. In the blood, the levels of Direct and Indirect bilirubin, as well as their ratio, change dramatically in Disorders of the liver, spleen, bone marrow, blood diseases, etc.; therefore, determining the levels of both forms of bilirubin is of crucial importance in the Cytology/practical/136.html">Differential Diagnosis OF Various Forms of jaundice. In cholelithiasis, indirect bilirubin is invariably detected within gallstones alongside their main component, Cholesterol. Due to its poor water solubility, it precipitates in the gallbladder as calcium bilirubinate, which contributes to stone formation.

The subsequent metabolic fate of bile pigments, specifically bilirubin, involves their transformation in the intestine by bacterial action. First, glucuronic acid is cleaved from the bilirubin complex, and the liberated bilirubin is reduced to stercobilinogen, which is excreted from the intestine. A normal human excretes about 300 mg of stercobilinogen per day. The latter is readily oxidized by light and air into stercobilin. The Mechanism of bacterial conversion of bilirubin to stercobilin has not yet been fully elucidated. Evidence indicates that the intermediate products of reduction are successively mesobilirubin and mesobilinogen (urobilinogen). Following absorption, a small fraction of mesobilinogen travels via the portal vein to the liver, where it is degraded to form mono- and dipyrrole compounds. In addition, a very small fraction of stercobilinogen, upon absorption through the hemorrhoidal Venous system, enters the systemic Circulation bypassing The Liver and is excreted in the urine in this form. However, calling it urobilinogen is not entirely accurate (see Chapter 18). The daily excretion of stercobilinogen in urine is about 4 mg, and stercobilinogen is indeed a normal organic constituent of urine. An elevated urinary excretion of urobilinogen (or more precisely, mesobilinogen) indicates impaired liver function, such as in hepatic or hemolytic jaundice, where the liver partially loses its ability to extract this pigment from portal blood. Chemically, urobilinogen (mesobilinogen) is not identical to urinary stercobilinogen (urobilinogen). The disappearance of stercobilinogen (urobilinogen) from urine in the presence of bilirubin and biliverdin indicates a complete cessation of bile flow into the intestine. This condition is frequently observed in the obstruction of the gallbladder duct (cholelithiasis) or the common bile duct (cholelithiasis, Pancreatic Cancer, etc.).

Thus, quantitative and qualitative analysis of bile pigments in urine can be of great clinical significance.