2-Methyl-3-hydroxybutyric acid, which is also known as 3-Hydroxy-2-methyl-butanoic acid (HMBA) is a normal urinary metabolite involved in the isoleucine catabolism, as well as presumably beta-oxidation of fatty acids and ketogenesis, excreted in abnormally high amounts in beta-ketothiolase deficiency.

Beta-ketothiolase deficiency is an inherited disorder in which the body cannot effectively process a protein building block (amino acid) called isoleucine. This disorder also impairs the body's ability to process ketones, which are molecules produced during the breakdown of fats.

2-Methylacetoacetic acid is a metabolite that has an increased excretion in patients with acetoacetyl-CoA thiolase deficiency. Thiolases are ubiquitous and important enzymes. Several isoenzymes are known, which can occur in the cytosol, the mitochondria, or the peroxisomes. Thiolases are CoA-dependent enzymes which catalyze the formation of a carbon-carbon bond in a Claisen condensation step and its reverse reaction via a thiolytic degradation mechanism. Mitochondrial acetoacetyl-coenzyme A (CoA) thiolase (T2) is important in the pathways for the synthesis and degradation of ketone bodies as well as for the degradation of 2-methylacetoacetyl-CoA. Moreover, 2-methylacetoacetic acid is found to be associated with beta-ketothiolase deficiency, which is also an inborn error of metabolism. 2-Methylacetoacetic acid is found in urine and can be used as a biomarker for the diagnosis of beta-ketothiolase deficiency.

2-Methylhippurate is a byproduct of detoxification of the common solvent xylene. Urinary excretion of 2-methylhippurate is a sensitive and specific marker for xylene exposure which increases oxidative stress.

2-Methylhippurate is a byproduct of detoxification of the common solvent xylene. Urinary excretion of 2-methylhippurate is a sensitive and specific marker for xylene exposure which increases oxidative stress.

Methylsuccinic acid is a normal metabolite found in human fluids. Increased urinary levels of methylsuccinic acid (together with ethylmalonic acid) are the main biochemical measurable features in ethylmalonic encephalopathy, a rare metabolic disorder with an autosomal recessive mode of inheritance that is clinically characterized by neuromotor delay, hyperlactic acidemia, recurrent petechiae, orthostatic acrocyanosis, and chronic diarrhea. The underlying biochemical defect involves isoleucine catabolism. 

Moreover, methylsuccinic acid is found to be associated with ethylmalonic encephalopathy, isovaleric acidemia, and medium-chain acyl-CoA dehydrogenase deficiency, which are also inborn errors of metabolism.

Note: These tests are used to check for rare metabolic disorders, usually in infants. There is no apparent reason nor benefit to checking ethylmalonic and methylsuccinic acid levels in adults who aren’t suspected to have rare genetic disorders.

2-hydroxyestrone and 16-hydroxyestrone are Phase I metabolites of Estrone (E1). Their ratio is of clinical significance in pre and peri-menopausal women. In post-menopausal women it does not have the same clinical significance. It is, however, hypothesized that the 2/16 ratio is important in menopausal women who are on hormone replacement therapy (HRT).

2-hydroxyestrone and 16-hydroxyestrone are Phase I metabolites of Estrone (E1). Their ratio is of clinical significance in pre and peri-menopausal women. In post-menopausal women it does not have the same clinical significance. It is, however, hypothesized that the 2/16 ratio is important in menopausal women who are on hormone replacement therapy (HRT).