COBALAMINE (B12]

Gandham. RajeevDepartment of Biochemistry,Akash Institute of Medical Sciences & Research Centre,Devanahalli, Bangalore, Karnataka, India.

E-Mail: gandhamrajeev33@gmail.com

CobalaminExtrinsic factor (EF) of castleAntipernicious anemia factorChemistry:Vitamin B12 is water soluble, heat stable

and red in colorIt contains 4.35% cobalt by weightFour pyrrole rings co-ordinated with a

cobalt atom is called as a Corrin ring

Synonyms of Vitamin B12

The corrin ring has four pyrrol units, like porphyrin

Two of the pyrrole units (A&D) are directly bound to each other where as the other two (B & C) are held by methene bridges

The groups namely methyl, acetamide and propionamide are the substituents on the pyrrole rings

Cobalt present at the centre of the corrin ring is bonded to the four pyrrole nitrogens

Cobalt also holds dimethyl benzimidazole (DMB) containing ribose 5-Phosphate and amino isopropanol

A nitrogen atom dimethyl benzimidazole is linked to cobalt

The amide group of aminoisopropanol binds with D ring of corrin

The cobalt atom also possesses a sixth substituent group located above the plane of the corrin ring

A

D

B

C

Aminoisopropanol

Methyl, Adenosine, acetamide, propionamide

Aminoisopropanol

dimethyl benzimidazole

D

BA

C

All the forms of vitamin B12 are called cobalamins because of the presence of cobalt atom

Different forms of cobalamins include Cyanocabalamin:- It refers to the isolated form of vitamin B12 in which cyanide was added to promote crystallization of cobalamin during the isolation process

Hydroxycobalamin:- It was usually present in the tissues and in the naturally occurring forms of Vitamin B12

Methylcobalamin :- Methyl group replaces adenosyl group

Adenosyl cobalamin (Ado-B12):-When taken up by the cells, these groups

are removed and deoxy adenosyl cobalamin or Ado-B12 is formed

In the food, vitamin B12 is present as a complex with proteins

The free form of vitamin B12 is released by cooking, HCL present in gastric juice and proteolysis by pepsin in the stomach

Mechanism:-The absorbance of vitamin B12 requires

intrinsic factorIntrinsic factor is a glycoprotein secreted by

parietal cells of stomach

metabolism

Vitamin B12 combines with intrinsic factorThe vitamin B12 – intrinsic factor complex

reaches the ileum where it is absorbedIn the ileum, the complex attaches to a

specific receptor and is taken up by the mucosal cell.

In the mucosal cell, vitamin B12 is released from its complex and reaches the portal circulation

In the portal blood, it is transported in combination with transcobalamin II

Vitamin B12 is presented to cells where it is taken up by the cells through receptor mediated endocytosis

Storage:It is mainly stored in liver, leukocytes and

gastric mucosaIt is stored as complex with transcobalamin-

I&II

Transport

ProteinB12

B12

B12

B12

Protein

IF

IF

IFMucosal cell

B12

Methyl B12B12

B12

TC 1-B12(90%)

TC 1-B12(90%)

TCII -B12(10%)

TCII -B12(10%)

Tissues

B12

TCII

Methyl B12

Deoxyadenosyl

B12(LIVER)

GITPlasma Tissues

About 10 enzymes requiring vitamin B12 have been identified

Most of them are found in bacteria ( mutase, ribonucleotide reductase, etc.)

There are two reactions in mammals that dependent on vitamin B12

Biochemical functions

Methyl cobalamin is essential for the conversion of homocysteine to methionine and formation of FH4 from methyl FH4

The reaction is catalyzed by homocysteine methyl transferase

Synthesis of Methionine from homosysteine

HomocysteineHomocysteine methyl

transferase

Methioninemethylcobalamin

THFN5 methyl THF

The circulating methyl FH4 is converted to FH4 FH4 is either used for storage as

folylpolyglutamate form or it is utilized for other reactions such as formation of methylene FH4

Methyl folate trap:- In B12 deficiency, impaired conversion of methyl

FH4 to FH4 results in accumulation of methyl FH4 & is called as methyl folate trap

Methyl folate trap results in decreased availability of FH4 & FH4 derivatives that are

Significance of the reaction

Needed for purine nucleotide & thymidylate synthesis

Thus vitamin B12 deficiency, results in secondary folate deficiency

Role of methyl cobalamin & folate trap

Methyl FH4

FH4

N5,10 methylene FH4dUMP

dTMP

DNA

Homocysteine

Methionine

Methylfolate trap Methyl FH4Homocysteine methyl

transferase

FH4

SerineGlycine

Thymidylate synthase

B12

Isomerization of methyl CoA to succinyl CoA:-

The degradation of odd chain fatty acids and some amino acids (valine, leucine etc) and pyrimidines (thymine & Uracil) produce propionyl CoA, an imp. Compound methylmalonyl CoA

The methyl malonyl CoA mutase converts methyl malonyl CoA to succinyl CoA in the presence of Vitamin B12,deoxyadenosyl cobalamin

In B12 deficiency, methyl malonyl CoA accumulates and is excreted in urine as methyl malonic acid

This condition is called as methylmalonic aciduria, occurs in B12 deficiency.

Demyelination :- Myelination of nerves is impaired in B12 deficiency due to accumulation of methylmalonyl CoA

Demyelination is due excessive accumulation of methylmalonyl CoA

Odd chain FA

Propionyl CoA

Amino acids(Val, Ile, Thr,

Met)Thymine,

uracil

Methyl malonyl CoA

Succinyl CoA

Methyl malonyl CoA mutase

Methyl malonic acid

Excreted in urine

5-Deoxyadenosylcabalamin (of B12)

+ Methylmalonic aciduria

Rich sources of vitamin B12 are meats, egg, milk, sea foods

B12 is synthesized by microorganismsVitamin B12 is not present in Plant sources

Dietary sources

Adults -1 µg/dayPregnancy & lactation -2 µg/day

Causes:Inadequate intake-seen in pure vegetarians

and rarely in alcoholismImpaired absorptionThis is mainly caused by lack of intrinsic factorLack of intrinsic factor is called as pernicious

RDA

Deficiency

anemia & it is caused by destruction of gastric mucosa

Impaired absorption is also seen in small intestinal disorders

Impaired storage and transport:Inadequate utilization of vitamin occurs because

of liver diseases and abnormalities of transport proteins

Increased requirements are seen in hyperthyroidism, infancy & thalassemia

Increased excretion occurs in nephrotic syndrome

Vitamin B12 deficiency is characterized by Megaloblastic anemia:Feature of megaloblastic anemia include

pallor,fatigue, glossitis ( beefy red tongue) & slight yellow discoloration of the conjunctiva due to increased unconjugated bilirubin

Progression of anemia may result in angina & congestive cardiac failure

Clinical Features

Gastrointestinal dysfunction:GIT epithelial cells are undergoing rapid

turnover & dependent on vitamin B12B12 deficiency results in weight loss &

diarrheaDemyelination of nervous tissueSubacute combined degeneration:Damage to nervous system is seen in B12

deficiency

There is demyelination affecting cerebral cortex as well as dorsal column & pyramidal tract

Symmetrical paresthesia of extremities, alterations of tendon & deep senses & reflexes, loss of position sense, unsteadiness in gait, positive Romberg’s sign & positive Babinski’s sign are seen

Achlorhydria:Absence of acid in gastric juice is associated

with B12 deficiency

Serum B12: It is quantitated by radio-immunoassay or by ELISA

Methyl melonic acid is excreted in urineFIGLU excretion testPeripheral smear: Peripheral blood & bone

marrow morphology shows magaloblastic anemia

Homocysteinuria: Excretion of homocysteine in urine

Assessment of B12 deficiency

Schilling test:Radioactive labelled (Cobalt-60) vitamin B12

1μ g is given orally In gastric atrophy cases, there is no

absorption, hence the entire radioactivity is excreted in faeces & radioactivity is not observed in liver

If the cause is nutritional deficiency, there will be increased absorption

Then radioactivity is noted in the liver region, with very little excretion in feces

ReferencesHarper’s Biochemistry 25th Edition.Fundamentals of Clinical Chemistry by Tietz. Text Book of Medical Biochemistry-A R Aroor.Text Book of Biochemistry-DM VasudevanText Book of Biochemistry-MN ChatterjeaText Book of Biochemistry-Dr.U.Satyanarana