# Usefulness of levels of 2-methylbutyrylglycine and 2-ethylhydracrylic acid in urine for diagnosing 2-methylbutyrylglycinuria

## TL;DR

This study compares two urine markers for diagnosing a rare metabolic disorder, finding both are highly sensitive but one has better specificity.

## Contribution

The study evaluates the diagnostic utility of 2-MBG and 2-EHA in urine for 2-methylbutyrylglycinuria, highlighting their sensitivity and specificity.

## Key findings

- Both 2-MBG and 2-EHA in urine showed 100% sensitivity for diagnosing 2-methylbutyrylglycinuria.
- 2-MBG had higher specificity (99.5%) compared to 2-EHA (97.8%).
- 2-MBG levels correlated with C5 acylcarnitine levels in blood, but 2-EHA did not.

## Abstract

The congenital disease 2-methylbutyrylglycinuria involves defective isoleucine metabolism and is currently diagnosed based on elevated levels of 2-methylbutyrylcarnitine/isovalerylcarnitine C5 in blood, followed by analysis of 2-methylbutyrylglycine (2-MBG) in urine at an early stage. However, 2-ethylhydracrylic acid (2-EHA) was reported as a more easily detected marker for 2-methylbutyrylglycinuria due to its larger volume in urine. Here we explored the usefulness of two urinary markers for diagnosing the condition.

We used electrospray-tandem mass spectrometry to assay levels of C5 and other acylcarnitines in blood and gas chromatography-mass spectrometry to assay levels of 2-MBG and 2-EHA in the urine of 12 individuals diagnosed with 2-methylbutyrylglycinuria based on ACADSB genotyping and in 166 controls. The diagnostic sensitivity and specificity of 2-MBG and 2-EHA were assessed based on receiver operating characteristic curves. Potential associations of levels of the two urinary markers with levels of C5 and other acylcarnitines in blood were explored using Pearson correlation analysis.

Urine of the 12 individuals with 2-methylbutyrylglycinuria contained a 2-MBG level of 1.78–11.89 (reference range, 0–0) and a 2-EHA level of 37.80-373.13 (reference range, 0-28.69), but levels of both analytes were undetectable or barely detectable in the urine of the 166 controls. Both markers showed 100% diagnostic sensitivity, and 2-MBG showed slightly higher specificity than 2-EHA (2-MBA, 99.5%, 95%CI 0.970-1.00; 2-EHA, 97.8%, 95%CI 0.943–0.994). Levels of 2-MBG, but not 2-EHA, in urine showed a reliable Pearson correlation with levels of C5 in blood (R2 = 0.71, P = 0.0006), but not with the ratios of C5 to other acylcarnitines in blood.

The urinary markers 2-MBG and 2-EHA show similar sensitivity for diagnosing 2-methylbutyrylglycinuria, but the lower specificity of 2-EHA and the fact that it can be elevated in several other congenital metabolic disorders means that it should be used as a diagnostic marker only in conjunction with other markers and clinical factors.