Water Soluble Vitamins - NEET PG Biochemistry

Vitamins are essential components in maintaining optimal body health and they play a vital role in many biochemical functions in the human body. There are two main groups of vitamins, fat-soluble (easily stored in fat after absorption) and water soluble vitamins (gets washed out easily and not easy to store).

In this blog post, we shall discuss everything about water-soluble vitamins for your Biochemistry Exam preparation. 

Water Soluble Vitamins

• Vitamin B Complex are involved in the core metabolic pathways.

Thiamine - Vitamin B1

• It is a sulphur containing vitamin.

• It is present in the aleurone layer of the grain.

Aleurone Layer

• Polishing these grains can cause the removal of aleurone layers and so does the thiamine.
• Parboiling of grains, attaches the aleurone layer to the grain, making it intact during polishing.

Biochemical Role

It is important in two biochemical reactions.

• Oxidative decarboxylation
  • PDH Complex
    • Significance of PDH Complex: It converts pyruvate (From Glucose) into acetyl CoA, which in turn is involved in the CAA cycle and releases it as carbon dioxide. It is necessary for oxidative utilization of glucose.
    • If thiamine deficiency - Lactic acidosis.
      • The Pyruvates has two fates, either it gets converted into acetyl CoA or lactate upon action by PDH or Lactate dehydrogenase. In thiamine deficiency, PDH is blocked. Thus, pyruvate gets converted into lactic acid. Lactic acid accumulation causes lactic acidosis.
  • Alpha KGDH
  • Branched chain ketoacid dehydrogenase.
    • It is involved in branch chained amino acid metabolism. Thiamine Deficiency - MSUD.
  • Transketolase
    • They're involved in HMP Shunt, which utilises glucose. Thiamine Deficiency - Estimate RBC transketolase activity.

RDA

• It is dependent on calorie or carbohydrates intake. 0.5 mg/1000 calories.

Beri-Beri

• Dry Beri-Beri - CNS involvement.
  • Neurons use glucose for energy derivation. Neurons suffer due to the thiamine deficiency.
• Wet Beri-Beri - CVS involvement.

Alcoholism and Thiamine

• Alcohol is metabolised by Alcohol Dehydrogenase and forms one aldehyde. Aldehyde is converted into an acid via Aldehyde Dehydrogenase. Both the enzymes remove one H-Atom from their substrate and add it to NAD (Coenzyme) to form NADH.
• It signifies, alcohol can be demonstrated as ↑NADH/NAD ratio and ↑ATP/ADP ratio. This causes energy status to become higher, which decreases the appetite of the person. This causes that person to miss the mixed balanced diet. It can cause essential micronutrients deficiencies. Thus, alcoholism is the source of "Empty Calorie."

• Thiamine deficiency is the first seen micronutrient deficiency in chronic alcoholism, as alcohol interferes with thiamine absorption.
  • Reasons
    • High energy status.
    • Missed mixed balance diet.
    • Essential micronutrient deficiencies.
• Even thiamine supplements are taken without a stop on alcohol consumption there will be no change in the symptoms as the alcohol interferes with the thiamine absorption.
• Magnesium malabsorption - Hypomagnesemia.
  • Any thiamine in the system can't be transformed into thiamine pyrophosphate (Coenzyme Form of Thiamine).

Thiamine manifestations in chronic alcoholism

• Wenicke's encephalopathy - Acute thiamine deficiency. 
  • Manifestations:
    • G - Global Confusion. 
    • O - Opthalmoplegia.
    • A - Ataxia.

• Korsakoff syndrome - Chronic thiamine deficiency.
  • Manifestations:
    • Amnesia
      • Retrograde amnesia 
      • Anterograde amnesia.
      • Confabulation.
        • Fabrication of stories to fill up the memory gaps.
      • Sensory agnosia.

Riboflavin - Vitamin B2

• It has a carbohydrate, ribitol in its structure. 

Coenzyme Roles

• Coenzyme role of riboflavin is in the form of:
  • FMN
  • FAD

Biochemical Role

• ETC
  • Both FMN and FAD are used.
• Carbohydrate metabolism
  • PDH uses FAD as its coenzyme.
    • It is an enzyme complex and has 3 subunits and 5 coenzymes.
    • 5 coenzymes:
      • TPP
      • Lipoamide
      • CoA
      • FAD (Involved in aerobic form of glucose)
      • NAD
• Lipid metabolism
  • Acyl CoA dehydrogenase - 1st enzyme for fatty acid oxidation.
    • It uses FAD as its coenzyme. 
• Amino acid metabolism 
  • L - Amino acid oxidase - It uses FAD as its coenzyme.
• Citric acid cycle
  • Succinate Dehydrogenase - It uses the FAD.

Deficiency Manifestations

• Riboflavin is involved in many metabolic pathways and is necessary for rapidly dividing labile cells, thus epithelium gets affected.
• Example: Angular stomatitis.
• 1st sign of riboflavin deficiency - Circumcorneal congestion.
• For Vitamin B2 deficiency - RBC or erythrocyte glutathione reductase activity should be checked.
• Trick - Ribbon and Glue go hand in hand in artwork.
  • Ribbon - RBC.
  • Glue - Glutathione reductase activity.

Niacin or B3

• It can be synthesised via metabolic pathways.
• 60 mg of tryptophan = 1 mg of niacin.

Synthesis of Niacin

Step-01: Tryptophan → FormylKynurenine

• Catalysed by Tryptophan pyrrolase.

Step-02: FormylKynurenine → Kynurenine

• Catalysed by Formyltransferase
  • Converts Tetrahydrofolate to formyl Tetrahydrofolate.
  • It enters One carbon pool and is used to form C2 of purine rings.

Step-03: Kynurenine → 3-OH Kynurenine

• Catalysed by Hydroxylase.

Step-04: 3-OH Kynenurine → 3-OH Antitranilic acid

• It utilises pyridoxal phosphate.
• Catalysed by Kynenurinase and dependent on B6 or Pyridoxal phosphate.
• B6 is necessary for
  • All transaminases.
  • Most of the Decarboxylases.
  • Kynenurinase.

Step-05: 3-OH Antitranilic acid → Quinolinate 

• 3-OH Antitranilic acid undergoes a series of non-enzyme catalase reactions to form a ring (Quinolinate).

Step-06: Quinolinate → Niacin Mono Nucleotide

• Rings get attached to Ribose 5 Phosphate, and form nucleotides
  • Examples
    • Adenine + Ribose 5 Phosphate = AMP.
    • Guanine + Ribose 5 Phosphate = GMP.
• Quinolinate phosphoribosyl transferase (QPRTASE) is involved in the reaction.
  • Role: It converts the ring form (In step 6 - Quinolinate) to nucleotide by adding ribose 5 phosphate.

Step-07: Niacin Mono Nucleotide → Niacin Adenine Dinucleotide

• Another phosphoribosyl transferase that acts on NAD to form adds R5P.
• Niacin mono nucleotide is the first coenzyme form of niacin.

Step-08: NAD → NADP

• Kinase acts on the NAD to convert it into NADP.

Pellagra

• Deficiency of Niacin causes pellagra.
• Characterised by 3 D's;
  • Diarrhoea
  • Dementia
  • Dermatitis (Photosensitivity).
    • Castles necklace. 

Causes of Pellagra 

Diet with;

• Niacin deficiency.
• Tryptophan deficiency in the maize-based diet.
  • Farmers cultivate many crops like rice, wheat, and maize, but the economic value of rice and wheat is high.
  • They sell off the rice and wheat and consume only maize.
• Tryptophan malabsorption - Hartnup's disease.
  • It is caused by the malabsorption of tryptophan in the intestine.

Hartnup's Disease (Blue Diaper Syndrome)

• Tryptophan malabsorption syndrome.
• It is caused by the defect of neutral amino acid transporter specific for tryptophan.
• It is responsible for both.
  • Absorption along the intestine, if not:
    • Hartnup's disease.
    • Niacin deficiency.
  • Reabsorption from the renal tubule:
    • It causes amino aciduria.
    • As the tryptophan is present in urine and exposed to air, then the indole ring of tryptophan will get oxidised to form ketol compounds and produce blue colour.
    • Thus, causes blue discoloration of urine - Blue Diaper Syndrome.
    • produce blue colour.
• In carcinoid syndrome.
  • Tryptophan is predominantly used to form Serotonin (5 hydroxytryptamine). Thus, tryptophan is not available for niacin synthesis. Niacin deficiency.
• Vitamin B6 deficiency:
  • It inactivates Kynenurinase. Thus, the precursor tryptophan cannot be used in niacin synthesis.
• Leucine pellagra:
  • Leucine inhibits Quinolinate phosphoribosyltransferase (QPRTASE). In a leucine rich diet, for example sorghum diet.

Causes of Pellagra

• Niacin deficiency in the diet. 
• Tryptophan deficiency in the diet - Maize diet.
• Tryptophan malabsorption - Hartnup's disease. 
• Carcinoid syndrome
• B6 deficiency
• Leucine pellagra - Sorghum based diet.

Fact - B6 Deficiency Detection

• In B6 deficiency, the kynenurinase enzyme becomes inactive. The precursor kynurenine gets converted into Xanthurenic acid. After tryptophan load, estimate the xanthurenic acid in urine.

Folate Deficiency Detection

• Histidine gets metabolised to form FIGLU (Formiminoglutamic acid).
• In general:
  • FIGLU will transfer its imino group to THFA, thus converting THFA into FITHFA. FITHFA enters 1 carbon pool. FIGLU becomes glutamate. Glutamate converts into alpha ketoglutarate and enters the CAA cycle. 
• In folate deficiency:
  • After histidine load, high FIGLU levels in urine is seen as it cannot transfer its imino group to THFA.

Vitamin B6 

• Its coenzyme form is pyridoxal phosphate.

Vitamin B6 - Coenzyme Roles

• It is necessary for the activity of:
  • Glycogen phosphorylase
    • Mcardle's disease - Type 5 GSD.
    • Muscle glycogen phosphorylase enzyme is defective.
    • It responds to B6 administration. 
  • Transaminases
  • Decarboxylases 
    • Examples:
      • Glutamate decarboxylases 
  • Kynenurinase
    • It involves the conversion of tryptophan to niacin.
  • Cystathionine beta synthase 
  • It helps in the conversion of homocysteine to cysteine.

Glutamate Decarboxylase

• It converts glutamate into GABA. The decarboxylation reaction is catalysed by glutamate decarboxylase which is dependent on PLP. Alpha carboxyl group is removed in the form of carbon dioxide. Thus, forming the gamma amino butyric acid, an inhibitory neurotransmitter.
• Structure:
  • Every amino acid has an alpha carbon atom which is attached to the amino group and carbohydrate group. Propionic acid (With gamma and beta carbon atoms) gets attached to form the glutamate.
• In B6 deficiency, glutamate gets accumulated making the person GABA deficient. Low levels of excretory neurotransmitters and high levels of inhibitory neurotransmitters, thus causes seizures.
• Seizures respond to B6 administration. 
• Case - If a neonate presents with seizures.
  • The most common causes:
    • Hypoglycemia
    • Hypocalcemia.
• To treat this condition calcium glutamate is given. If the seizures are not getting decreased, before any investigations on the causes of seizure, the seizure activity should be brought down. A B6 shot can be given in the IM route. Any glutamate present in the muscle gets converted into the GABA. Thus, reducing excitatory neurotransmitter levels and increasing inhibitory neurotransmitter levels.

ALA Synthase (heme synthesis)

• 1st step of heme synthesis - Decarboxylation reaction which needs pyridoxal phosphate. Glycine reacts with succinyl CoA in the presence of delta ALA synthase to form delta ALA.
• In pyridoxal phosphate deficiency;
  • Heme will not be synthesized, thus it causes anaemia.
  • Anaemia responds to B6 administration.

Vitamin B12

• It has got 2 coenzyme roles:
  • Methyl B12
  • Adenosyl B12

Adenosyl B12

• It acts as a coenzyme for methyl malonyl CoA mutase. 

In odd chain fatty acid oxidation:

• Propionyl CoA → Methyl malonyl CoA. 
• Methyl malonyl CoA → Succinyl CoA. 
  • It is catalysed by methyl malonyl CoA mutase, dependent on adenosyl B12.
• The succinyl CoA is involved in the CAA cycle and gets converted into carbon dioxide. 
  If any defect on the metabolism can cause the manifestations of accumulation of the respective substrates.

In B12 deficiency:

• Adenosyl B12 will not be available. Methyl malonyl CoA mutase becomes inactive. It causes methyl malonic aciduria. 

Investigation of B12 deficiency

• To rule out B12 deficiency, urine levels of methyl malonic acid are estimated after overnight fasting.
• Reason: Peripheral lipolysis occurs which pulls the odd chain fatty acids to the liver, which ultimately metabolises into the methyl malonyl CoA. The methyl malonic acid gets incorporated into myelin, which causes formation of abnormal myelin, in due course neurons undergo demyelination. Initially patients present with paraesthesia (Tingling sensation) in the periphery.
• In severe cases, it forms the SACD (Subacute combined degeneration).
  • Presents with:
    • Sensory neuropathy 
    • Motor neuropathy
      • Cranial nerve involvement.

Methyl B12

• It is a coenzyme for methionine synthase or homocysteine methyltransferase. It helps in the conversion of Homocysteine to Methionine.
• Homocysteine has two fates:
  • It can be converted into cysteine upon action by cystathionine beta synthase.
  • It can be converted into methionine, upon action by methionine synthase.
• Methyl THFA donates its methyl group to be methyl B12. The THFA form is recycled for other THFA purposes. 

In B12 deficiency:

• Methyl THFA cannot donate its methyl group and is trapped as such. This causes the functional deficiency of methyl THFA and is called Methyl Folate Trap. This condition occurs due to the low activity of methionine synthase or homocysteine methyltransferase. Indirectly B12 deficiency causes the folate deficiency, which in turn leads to macrocytic anaemia.
• The decreased activity of methionine synthase, homocysteine gets accumulated, homocystinuria.

Features of B12 Deficiency

• Adenosyl B12 deficient.
  • Methyl malonyl CoA mutase becomes inactive;
    • Demyelination - Neurological
      • It causes paresthesia and in severe terms SACD.
  • Methyl malonic aciduria
    • Urine levels of methyl malonic acid are estimated after overnight fasting.
• Methyl B12 deficient:
  • Low functional THFA.
  • Methyl Folate Trap, it causes Macrocytic anaemia.
  • Homocystinuria.

Factor Affecting Vitamin B12 absorption

• Non vegetarian sources: All the vitamin B12 sources are non-vegetarian.
• Pancreatic enzymes (Important): Vitamin B12 is attached to a protein (Blocks B12 absorption), which can be separated upon action by pancreatic enzymes.
• Cobalophilin receptors in terminal ileum: Once vitamin B12 is attached to the intrinsic factor, the cobalophilin receptors accept the vitamin B12.
• Intrinsic Factor bound Vitamin B12 (Important): IF is released by gastric mucosa.
• Intestinal Microorganism: Excessive microorganisms in the colon utilise vitamin B12.
• Terminal ileum (Important): It should be healthy for good vitamin B12 absorption, if not it won't get absorbed.

Causes of Vitamin B12 Malabsorption

They're;

• Pancreatic insufficiency. IF deficiency / Atrophic gastritis / Autoimmune gastritis / Type A gastritis. Blind loop syndrome - Excess colonic microorganisms.
• Terminal ileum - Crohn's disease.
  • Regional ileitis.

Schilling's Test

Two purposes:

• Detects vitamin B12 malabsorption. 
• Identify vitamin B12 malabsorption.

Steps Followed in Schilling's Test

Step 01: To treat Vitamin B12 and folate deficiency.

• Both vitamins are necessary for the health of rapidly dividing labile cells in the intestinal mucosa. If intestinal cells are not healthy it itself results in the vitamin B12 malabsorption. In this case malabsorption is the effect, not the cause.

Step 02: Oral radiolabelled Vitamin B12 is provided.

• They are in measured quantities.  Once given orally, vitamin B12 has two fates. It binds with the B12 receptors in the liver.
  • If overflow of receptors in the liver occurs, then the excess is released out of the body via urine.
  • In Schilling's Test, vitamin B12 levels in the urine are estimated.
    • Before the test, all the receptors for B12 binding should get saturated.

Step 03: IM unlabelled vitamin B12 is administered to saturate all vitamin B12 receptors present in the liver.

Step 04: 24 hours urine is collected and labelled vitamin B12 is estimated.

• Inference: If less than 10% of orally administered Vitamin B12 is found in urine - malabsorption is confirmed.
• Normal - >10% of orally administered vitamin B12 should be excreted in urine, if there is no malabsorption.

Step 05: Test is repeated after oral IF.

• Inference: If less than 10% of orally administered Vitamin B12 is found in urine- Autoimmune gastritis is excluded.

Step 06: Test is repeated after 3 weeks of antibiotics.

• Inference: If less than 10% of orally administered Vitamin B12 is found in urine- Blind loop syndrome is excluded.

Step 07: Test is repeated after 2 days of pancreatic enzymes.

• Inference 01: If less than 10% of orally administered Vitamin B12 is found in urine- pancreatic insufficiency is excluded.
• Inference 02: Probably, Crohn's disease or celiac sprue.

One Liners

1. Schilling's test is used to diagnose Vitamin B12 malabsorption.
2. Vitamin K form used to treat Vitamin K deficiency due to fat malabsorption is Menadione.
   • Explanation
     • Discussed in vitamin A, D, E, and K.
     • In menadione, the polyisoprenoid side chain is absent, thus it is water soluble and not dependent on factors which are necessary for fat absorption.
3. Vitamin E RDA is dependent on Fatty acid intake.
   • Explanation 
     • Vitamin E is an antioxidant. 
     • Major source of oxidative stress is fat intake.
4. RDA of Vitamin B1 is dependent on carbohydrate intake.
   • Explanation: Thiamine is predominantly involved in aerobic utilisation of glucose. 
5. Tryptophan load test is done to detect B6 deficiency.
6. Histidine load test is done to detect folate deficiency.

Q. Thiamine deficiency is diagnosed by measuring which of the following?

1. RBC Glutathione reductase activity
2. RBC Transketolase activity
3. RBC G6PD activity
4. RBC Glutathione peroxidase activity

Q. Riboflavin deficiency is diagnosed by measuring which of the following?

1. RBC Glutathione reductase activity
2. RBC Transketolase activity
3. RBC G6PD activity
4. RBC Glutathione peroxidase activity

Q. Methylfolate trap in Vitamin B12 deficiency is because of the inhibition of?

1. Methyl Malonyl CoA mutase
2. Methylmalonyl CoA isomerase
3. Methionine synthase
4. Cystathionine Beta synthase

Explanation: Methyl B12 acts as a coenzyme for methionine synthase or homocysteine methyltransferase.

Q. Is PLP necessary as a coenzyme for all except?

1. Transaminases
2. Glutamate decarboxylase
3. Methionine synthase
4. Cystathionine Beta synthase

Explanation

• PLP is necessary for.
  • Glycogen phosphorylase
  • Transaminases
  • Decarboxylases 
    • Examples:
      • Glutamate decarboxylases
      • ALA synthase
  • Kynenurinase
  • Cystathionine beta synthase.
• Methionine synthase is dependent on vitamin B12 and folate.

Q. All the following are PLP dependent states except?

1. Anaemia
2. Seizures
3. Pellagra
4. Albinism

Explanation

• Clinical conditions that respond to B6 administration.
  • McArdle’s disease
  • Seizures 
  • Anaemia
  • Pellagra
  • Homocystinuria
• Albinism - It is caused by the defect of tyrosinase which is dependent on copper.

Q. A chronic alcoholic present with an episode of hypoglycaemia. He is treated with dextrose infusion and thiamine injection. Before discharge, he is advised about effects of chronic alcoholism, and he wants to check his thiamine availability status. What is the investigation you would prescribe?

1. RBC transketolase activity
2. RBC Glutathione reductase activity
3. Urine methyl malonic acid level
4. Urine Xanthurenic acid level

Explanation

• RBC Glutathione reductase activity - For riboflavin deficiency. 
• Urine methyl malonic acid level - B12.
• Urine Xanthurenic acid level - B6.

Q. A known patient of Tuberculosis is on INH and he presents with microcytic hypochromic anaemia. His serum ferritin and transferrin saturation are normal. A vitamin deficiency is suspected. Which vitamin deficiency can cause microcytic hypochromic anaemia and what is the investigation of choice for diagnosing this condition?

1. B12, methylmalonic acid in urine
2. Folate, FIGLU level in urine
3. B2, Glutathione reductase activity in RBCs
4. PLP, Xanthurenic acid in urine

Explanation

• In a patient with microcytic hypochromic anaemia, the first suspect should be iron deficiency.
• If serum ferritin and transferrin are normal, then suspect for vitamin deficiency. 
• In heme synthesis, delta ALA synthase is dependent on B6. 
• If B6 deficiency is suspected.
  • Xanthurenic acid levels in urine are estimated.
• INH causes B6 deficiency, because the active form of vitamin B6 (Pyridoxal phosphate) cannot be formed.
  • Reason: Both B6 and isoniazid is an aldehyde.
  • B6 PLP in the presence of PLK.
  • INH is a competitive inhibitor of PLK.
  • Thus the active form of vitamin B6 has not formed.

Q. A person, who is on a mixed balanced diet, presents with tiredness, weakness, macrocytic anaemia and subacute combined degeneration. He was given radiolabelled B12 orally and then intramuscular unlabelled B12 was injected. 24 hours urinary radiolabelled 812 was less than 10% of oral administered dose. After oral IF, 24 hours urinary labelled B12 was more than 10% oral administered dose. Which of the following is the probable cause?

1. Dietary B12 deficiency
2. Methyl Folate trap
3. Pancreatic insufficiency
4. Type A gastritis

Explanation

• If given SACD, think of vitamin B12 deficiency.
• Schilling's test is being done.
• Vitamin B12 deficiency is most commonly caused by nutritional causes.
• In case, if a person is vegan or vegetarian.
  • In this case, the person is on a mixed balanced diet still presenting vitamin B12 deficiency, probably B12 malabsorption is the suspicion.
• If it is confirmed, supplement IF.
• The IF administration has treated the condition in this case.
• Thus, the diagnosis is Type A gastritis or autoimmune gastritis, where the parietal cells get damaged.

And that is everything you need to know about water-soluble vitamins for Biochemistry and ace your NEET PG preparation. For more informative and engaging posts like these, download the PrepLadder App and keep following our blog!