Which of the following enzyme activities can be estimated in red blood cells to diagnose vitamin B2 deficiency?

Correct Answer: C. Glutathione reductase

Vitamin B2 (riboflavin) is the precursor for flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN), which are essential cofactors for numerous oxidoreductase enzymes. Glutathione reductase is a FAD-dependent enzyme that catalyzes the reduction of oxidized glutathione (GSSG) to reduced glutathione (GSH), a critical antioxidant in red blood cells. In B2 deficiency, FAD levels drop, reducing glutathione reductase activity. This makes RBC glutathione reductase activity an ideal biomarker for B2 status—the enzyme is abundant in RBCs, easily accessible, and directly dependent on FAD. The erythrocyte glutathione reductase activity coefficient (EGRAC) is the standard functional test: EGRAC = (activity with added FAD) / (basal activity). A ratio >1.3 indicates B2 deficiency. This is the gold-standard functional assay recommended in Indian and international nutrition guidelines for detecting riboflavin deficiency, especially in populations at risk (pregnant women, lactating mothers, malnutrition). Unlike static plasma riboflavin measurement, EGRAC reflects tissue B2 status and is more sensitive in detecting marginal deficiency.

Why the other options are wrong

A. Pyruvate dehydrogenase — Pyruvate dehydrogenase is a thiamine (B1) dependent enzyme, not B2-dependent. It catalyzes the conversion of pyruvate to acetyl-CoA and requires thiamine pyrophosphate (TPP) as a cofactor. While B1 deficiency impairs this enzyme, it is not used to diagnose B2 deficiency. This is a common trap pairing different B vitamins with their respective enzymes. B. Transketolase — Transketolase is also a thiamine (B1) dependent enzyme, not B2-dependent. It requires TPP as a cofactor and is used to diagnose thiamine deficiency via the transketolase activity coefficient (TPAC) in RBCs. NBE pairs this with B2 to test whether students confuse the cofactor requirements of different B-complex vitamins. D. Kynureninase — Kynureninase is a pyridoxal phosphate (B6) dependent enzyme involved in tryptophan metabolism. It is used to assess B6 status, not B2 status. This option tests whether students know the specific cofactor dependencies of different B vitamins and their corresponding diagnostic enzymes.

High-Yield Facts

Mnemonics

B Vitamins & Their Diagnostic Enzymes B1 (Thiamine) → Transketolase & Pyruvate dehydrogenase (TPP cofactor). B2 (Riboflavin) → Glutathione reductase (FAD cofactor). B6 (Pyridoxine) → Kynureninase (PLP cofactor). B12 → Methylmalonic acid & homocysteine. EGRAC Memory Hook Erythrocyte Glutathione Reductase Activity Coefficient = (activity + FAD) / (activity − FAD). If ratio >1.3, B2 is deficient. Think: 'More FAD boost = more deficiency.'

NBE Trap

NBE pairs all four enzymes from different B-vitamin pathways to test whether students confuse cofactor dependencies. The trap is recognizing that pyruvate dehydrogenase and transketolase both require B1 (not B2), while kynureninase requires B6—only glutathione reductase is B2-dependent.

Clinical Pearl

In Indian clinical practice, EGRAC is the preferred functional test for B2 deficiency in antenatal clinics and nutrition programs because it detects marginal deficiency before clinical signs (angular cheilitis, glossitis) appear. A single RBC sample suffices, making it practical for resource-limited settings.

_Reference: KD Tripathi Pharmacology Ch. 48 (Vitamins); Robbins Pathology Ch. 8 (Nutritional Deficiencies); Harrison Principles of Internal Medicine Ch. 75 (Vitamin Deficiencies)_