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BIOCHEMISTRY

Lec:4 : 2nd stage شيماء سبتي.د.م

B-COMPLEX VITAMINS

THIAMINE (VITAMIN B1)

Synonyms: Antiberiberi factor, antineuritic vitamin, aneurin.

►Biosynthesis: Synthesized by plants, yeasts and bacteria. Not synthesized by

human beings, hence should be supplied in diet. Intestinal bacterial flora can

synthesize the vitamin.

►Metabolism

Absorption: Free thiamine is absorbed readily from the small intestine, but the

pyrophosphate (ester-form) is not. Bulk of the dietary vegetable thiamine is in the

“free” form. In tissues, it is actively phosphorylated to form Thiamine

pyrophosphate (TPP) in Liver, and to a lesser extent in other tissues like muscle,

brain and nucleated RB Cells.

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►Plasma/blood level: Present in plasma and CS fluid in the “free” form, approx.

1 μg/100 ml. Blood cells contain 6 to 12 μg/100 ml where occurs as TPP.

►Storage: Capacity to store is limited. It is present in both free and combined

forms in heart (highest concentration), Liver and kidneys. In lower concentration in

skeletal muscle and brain. Total amount of Thiamine in body is approx. 25 mg.

►Excretion: If normal amount of thiamine is taken in the diet:

• About 10 per cent is excreted in the urine

• The remainder is (a) Partly phosphorylated and is used as coenzyme, and (b)

Partly degraded to neutral sulphur compounds and inorganic SO4 which are

excreted in urine.

►Occurrence and Food Sources

• Plant source: Widely distributed in plant kingdom. In cereal grains, it is

concentrated in outer germ/bran layers (e.g. rice polishings) (Richest source).

Other good sources are peas, beans, whole cereal grains, bran, nuts, prunes, etc.

Whole white bread is a good source.

• Animal source: Thiamine is present in most animal tissues. Liver, meat and eggs

supply considerable amounts. Ham/pork meats are particularly rich. Milk has low

concentration, but a good source as large quantities are consumed.

►METABOLIC ROLE

Biological active form is Thiamine pyrophosphate (TPP). Acts as a coenzyme in

several metabolic reactions.

• Acts as coenzyme to the enzyme pyruvate dehydrogenase complex (PDH) which

converts pyruvic acid to acetyl-CoA (oxidative decarboxylation) PDH Pyruvate

Acetyl-CoA TPP

Pyruvate PDH Acetyl-CoA

TPP

• TPP acts as the coenzyme (Co-carboxylase) of pyruvate carboxylase in yeasts for

the non-oxidative decarboxylation of pyruvate to acetaldehyde.

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►CLINICAL ASPECT

DEFICIENCY MANIFESTATIONS: Beriberi

The deficiency of thiamine produces a condition called beriberi. It is characterised

by the following manifestations.

1- CV manifestations: These include palpitation, dyspnoea, cardiac hypertrophy

and dilatation, which may progress to congestive cardiac failure.

2- Neurological manifestations: These are predominantly those of ascending,

symmetrical, peripheral polyneuritis. These neurological features may be

accompanied occasionally by an acute haemorrhagic polioencephalitis which is

then called as Wernicke’s encephalopathy.

3- GI symptoms: Amongst these, anorexia is an early symptom. There may be

gastric atony, with diminished gastric motility and nausea; fever and vomiting

occur in advanced stages.

Dry beriberi: When it is not associated with oedema.

Wet beriberi: Oedema is associated. It is probably in part to congestive cardiac

failure and in part to protein malnutrition (Low plasma albumin).

►Biochemical Features in Thiamine Deficiency

1- Decreased level of thiamine and TPP in blood and urine. Determination of

amount of thiamine excreted in 4 hours urine is used.

2- Accumulation of pentose sugars in RB cells due to retardation of transketolation

reaction.

3- Increased level of pyruvic acid and lactic acid in blood, due to retardation of

oxidative decarboxylation of pyruvic acid. – LA/PA ratio: Abnormal blood LA/PA

ratio is said to be more specific indicator of B1 deficiency.

4- Catatorulin effect: Decreased uptake of O2 by thiamine-deficient brain in vitro;

reversible by addition of thiamine.

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5- Saturation test (thiamine loading test): A lower urinary excretion of thiamine

and TPP after administration of a test-dose occurring in thiamine-deficient as

compared to normal subjects.

►Daily Requirements

• Adult: 0.5 mg for each 1000 calories; 1.0 to 1.5 mg for diets providing 2000 to

3000 C. Minimum requirement is 1.0 mg. Actual requirement is related more

directly to carbohydrates content of diet than to calorie value of diet.

• Children: Ranges from 0.4 mg for infants to 1.3 mg for preadolescents (10 to 12

years of age). Requirements Increases in Anoxia-shock and haemorrhage,

Serious illness and injury, During prolonged administration of broadspectrum oral

antibiotics, Increased calorie expenditure like fever, hyperthyroidism, Increased

carbohydrate intake, Increased alcohol intake, and pregnancy and in lactation.

RIBOFLAVIN (VITAMIN B2)

Synonyms: Lactoflavin, ovoflavin, hepatoflavin.

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►Biological Active Forms

The biological active forms, in which riboflavin serves as the prosthetic group (as

coenzyme) of a number of enzymes are the phosphorylated derivatives.

Two main derivatives are:

1- FMN (Flavin mononucleotide): In this the phosphoric acid is attached to

ribityl alcoholic group in position 5 ( Flavin-Ribityl-PO4).

2- FAD (Flavin adenine dinucleotide): It may be linked to an adenine nucleotide

through a pyrophosphate linkage to form FAD (Flavin-ribityl-P-P-ribose-Adenine)

Thus, FMN and FAD are two coenzymes of this vitamin.

The acidic properties given by phosphoric acid group influence their capacity for

combining with proteins apoenzyme-forming flavoproteins (Holoenzyme). Thus,

FP (holoenzyme) = FMN/FAD + Protein

(coenzyme) (Apoenzyme)

FP may also unite with metals like Fe and Mo thus forming Metalloflavoproteins.

►Biosynthesis

All higher plants can synthesize riboflavin. In nature, it occurs both as “free form”

and also as “nucleotide” form or as flavoproteins. Human beings and animals

cannot synthesise and hence solely dependent on dietary supply. In man,

considerable amounts can be synthesised by intestinal bacteria, but the quantity

absorbed is not adequate to maintain normal nutrition.

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Metabolism ►Absorption: Flavin nucleotides are readily absorbed in small intestine. Free

riboflavin undergoes phosphorylation, a prerequisite for absorption.

►Blood/Plasma level: Human blood plasma contains 2.5 to 4.0 μgm%, two-third

as FAD and bulk of remainder as FMN. Concentration in RB cells-15 to 30 μg/100

gm. Leucocytes and platelets-250 μg/100 gm. These values remain quite constant

even in severe riboflavin deficiency, hence determination of riboflavin in blood is

not useful. Riboflavin present in all tissues as nucleotides bound to proteins (FP),

highest concentration in liver and kidney.

►Excretion: Daily urinary excretion 0.1 to 0.4 mg (10 to 20% of intake).

• Milk: Riboflavin is secreted in milk, 40 to 80 per cent in ‘free’ form.

• Faeces: Free and nucleotides tend to remain quite constant, 500 to 750 μg daily,

largely from the unabsorbed bacterial synthesis.

►Occurrences and Food Sources

Widely distributed in nature, present in all plant and animal cells.

• Plant sources: High concentration occurs in yeasts. Appreciable amount present

in whole grains, dry beans and peas, nuts, green vegetable. Germinating seeds, e.g.

grams/Dals are very good source.

• Animal source: Liver (2–3 mg/100 gm), kidney, milk, eggs, Crab meat has high

content.

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►METABOLIC ROLE

FMN and FAD act as coenzymes in various H-transfer reactions in metabolism.

The hydrogen is transported by reversible reduction of the coenzyme by two

hydrogen atoms added to the ‘N’ at positions 1 and 10, thus forming dihydro or

leucoriboflavin. The principal enzyme reactions catalysed are as shown below:

FMN FAD

• Cytochrome-C-reductase • Xanthine oxidase (Xanthine → uric acid)

• D-amino acid oxidase • L-amino acid oxidase • Aldehyde oxidase

►CLINICAL ASPECT Deficiency Manifestations

There is no definite disease entity. Deficiency is usually associated with

deficiencies in other B-vitamins. In human beings lesions of the mouth, tongue,

nose, skin and eyes with weakness, and lassitude reported. They include:

• Lips: Redness and shiny appearance of lips.

• Cheilosis: Lesions at the mucocutaneous junction at the angles of the

mouth leading to painful fissures are characteristic.

• Tongue: Painful glossitis, the tongue assuming a red-purple (magenta)

colour.

• Seborrhoeic dermatitis: Scaly, greasy, desquamation chiefly about the

ears, nose and naso-labial folds.

• Eyes: May lead to corneal vascularisation and inflammation with

cloudiness of cornea, watering, burning of eyes, photophobia, scleral

congestion and cataract has also been reported.

• Protein synthesis: This is impaired in severe riboflavin deficiency; since

protein malnutrition interferes with utilisation and retention of riboflavin.

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►Daily Requirement

• Adults: 1.5 to 1.8 mg • Women in later half of pregnancy: 2.0 mg

• During lactation: 2.5 mg • Infants: 0.6 mg

• Children: 1.0 to 1.8 mg • Adolescence: 2.0 to 2.5 mg

Requirement increases After severe injury/burns, etc. during acute illness and

during convalescence, during increased protein utilization, in pregnancy and

actation, during oral broad spectrum antibiotic therapy.

NIACIN (VITAMIN B3)

Synonyms: Nicotinic acid, P-P factor, Pellagra-preventing factor of Goldberger.

In tissues: Occurs principally as the amide (nicotinamide, niacinamide). In this

form it enters into physiological active combination.

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►Biological “Active” Forms

In tissues, nicotinamide is present largely as a “dinucleotide”, the pyridine ‘N’

being linked to a D-ribose residue.

Two such neucleotide active forms are known:

• Nicotinamide adenine dinucleotide (NAD+) Other names are: DPN+, coenzyme-

I, cozymase, or codehydrogenase.

The compound contains:

– One molecule of nicotinamide,

– Two molecules of D-ribose,

– Two molecules of phosphoric acid, and

– One molecule of adenine. Structure may be shown schematically as follows

►Biosynthesis

• Amino acid tryptophan is a precursor of nicotinic acid in many plants, and

animal species including human beings. 60 mg of tryptophan can give rise 1 mg

of Niacin. Pyridoxal-P is required as a coenzyme in this synthesis (Refer

Tryptophan metabolism).

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• It can be synthesized also by intestinal bacteria. Bacteria in addition to

synthesis from tryptophan, can also synthesize from other amino acids, e.g.

glutamic acid, proline, ornithine and glycine.

• In human beings:

– In addition to dietary source,

– It is synthesised in tissues from amino acid tryptophan, and

– To a limited extent supplemented by bacterial synthesis in intestine.

Applied Aspect

In high corn diet, requirement of dietary niacin increases, as synthesis from

tryptophan cannot take place. The reason is the maize protein Zein lacks the amino

acid tryptophan. Hence pellagra is more common in persons whose staple diet is

maize.

Metabolism

►Absorption: Nicotinic acid and its amide are absorbed from the small intestine.

►Blood/plasma level

• Whole blood: 0.2 to 0.9 mg/100 ml (average 0.6 mg%)

• RB cells: 1.3 mg%

• Plasma-total activity: 0.025 to 0.15 mg% (average 0.075 mg%)

Note

1. Most of the nicotinic acid and its amide in the blood is in RB cells, presumably

as coenzyme.

2. Values in the blood are not altered significantly even in severe Niacin

deficiency. Hence its determination is of no value in the detection of clinical

deficiency states.

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►Excretion

In urine, it is excreted as follows:

• As nicotinic acid and nicotinamide: Normal adults on normal diet excretes both

nicotinic acid and its amide in urine.

Nicotinic acid: 0.25 to 1.25 mg daily. Nicotinamide: 0.5 to 4 mg daily.

• As N’-methyl nicotinamide: Major urinary metabolite is a methylated derivative-

N’-methyl nicotinamide. The methylation occurs in liver, by the enzyme

niacinamide methyl transferase.

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►Occurrence and Food Sources:

1. Both nicotinamide and coenzyme forms are distributed widely in plants and

animals.

2. Important food sources are:

• Animal source: Liver, kidney, meat, fish

• Vegetable source: Legumes (peas, beans, lentils), nuts, certain green vegetables,

coffee and tea. Nicotinamide is present in highest concentration in germ and

pericarp (bran) in cereal grains. Yeast also particularly rich. Poor sources are:

Fruits, milk and eggs.

Metabolic Role • The coenzymes NAD+ and NADP+ operate as hydrogen and electron transfer

agents by virtue of reversible oxidation and reduction.

• Function of NADP+ is similar to that of NAD+ in hydrogen and electron

transport.

• The two coenzymes are interconvertible.

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►CLINICAL ASPECT

Deficiency Manifestations

Pellagra: Nicotinic acid deficiency produces a disease called Cardinal features

described as “3 D’s” are • Dermatitis, • Diarrhoea, and • Dementia.

Precipitating factors are: (a) High-corn diet and (b) Alcoholism

Clinical Features

(a) Skin lesions: Typically involves areas of skin exposed to sunlight and

subjected to pressure, heat or other types of trauma or/irritation. The skin becomes

reddened, later brown, thickened and scaly.

(b) GI manifestations: Include

1- anorexia, nausea, vomiting, abdominal pain, with alternating constipation/

diarrhea, Diarrhoea becomes intractable later.

2- Gingivitis and stomatitis with reddening of the tip and margin of the tongue,

which become swollen and cracked.

3- Achlorhydria present in about 40% cases.

4- Thickening and inflammation of the colon, with cystic lesions of the

mucosa, which later becomes atrophic and ulcerated.

(c) Cerebral manifestations: These include headache, insomnia, depression and

other mental symptoms ranging from mild psychoneuroses to severe psychosis.

(d) General effects: These include:

1-Inadequate growth,

2-loss of weight and strength,

3-anaemia which may be due to associated deficiency of other vitamins.

4-dehydration and its consequences resulting from diarrhoea.

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Daily Requirement

• In adult: 17 to 21 mg daily

• Infants: 6 mg

• Pre-adolescence: 17 mg

Requirement Increases in:

Increased calorie intake or expenditure, acute illness or early convalescence, after

severe injury, infection and burns, high corn or Maize diet, pregnancy and

lactation.

• Effect on plasma Lipids: Nicotinic acid and NOT amide have been found to

reduce the plasma lipid concentration in certain cases of hyperlipidaemia. Large

doses of Nicotinic acid from 3 to 6 Grams per day have been found to reduce the

levels of cholesterol, β-lipoproteins and TG in

blood.

• Niacin toxicity: Excessive dosage can produce toxic effects:

– Dilatation of blood vessels and flushing.

– Skin irritation

– Can produce liver damage.