of hmb-fa is 25–40% higher than for hmb-ca. the fraction of an ingested dose that is excreted in urine does not differ between the two forms. absorption of hmb-ca after ingestion, hmb-ca is converted to β-hydroxy β-methylbutyrate following dissociation of the calcium moiety in the gut. when the hmb-ca dosage form is ingested, the magnitude and time at which the peak plasma concentration of hmb occurs depends on the dose and concurrent food intake. higher hmb-ca doses increase the rate of absorption, resulting in a peak plasma hmb level (cmax) that is disproportionately greater than expected of a linear dose-response relationship and which occurs sooner relative to lower doses. consumption of hmb-ca with sugary substances slows the rate of hmb absorption, resulting in a lower peak plasma hmb level that occurs later. excretion of hmb-ca hmb is eliminated via the kidneys, with roughly 10–40% of an ingested dose being excreted unchanged in urine. the remaining 60–90% of the dose is retained in tissues or excreted as hmb metabolites. the fraction of a given dose of hmb that is excreted unchanged in urine increases with the dose. metabolism biosynthesis and metabolism of β-hydroxy β-methylbutyrate in humans l-leucine branched-chain aminoacid aminotransferase α-ketoglutarate glutamate glutamate alanine pyruvate muscle: α-ketoisocaproate (α-kic) liver: α-ketoisocaproate (α-kic) branched-chain α-ketoaciddehydrogenase (mitochondria) kic-dioxygenase(cytosol) isovaleryl-coa β-hydroxyβ-methylbutyrate(hmb) excretedin urine(10–40%) hmb-coa β-hydroxy β-methylglutaryl-coa(hmg-coa) β-methylcrotonyl-coa(mc-coa) β-methylglutaconyl-coa(mg-coa) co2 co2 o2 co2 h2o co2 h2o (liver)hmg-coalyase enoyl-coa hydratase isovaleryl-coadehydrogenase mc-coacarboxylase mg-coahydratase hmg-coareductase hmg-coa synthase β-hydroxybutyratedehydrogenase mevalonatepathway thiolase unknownenzyme β-hydroxybutyrate acetoacetyl-coa acetyl-coa acetoacetate mevalonate cholesterol human metabolic pathway for hmb and isovaleryl-coa relative to l-leucine. of the two major pathways, l-leucine is mostly metabolized into isovaleryl-coa, while only about 5% is metabolized into hmb. the metabolism of hmb is catalyzed by an uncharacterized enzyme which converts it to β-hydroxy β-methylbutyryl-coa (hmb-coa). hmb-coa is metabolized by either enoyl-coa hydratase or another uncharacterized enzyme, producing β-methylcrotonyl-coa (mc-coa) or hydroxymethylglutaryl-coa (hmg-coa) respectively. mc-coa is then converted by the enzyme methylcrotonyl-coa carboxylase to methylglutaconyl-coa (mg-coa), which is subsequently converted to hmg-coa by methylglutaconyl-coa hydratase. hmg-coa is then cleaved into acetyl-coa and acetoacetate by hmg-coa lyase or used in the production of cholesterol via the mevalonate pathway. biosynthesis hmb is synthesized in the human body through the metabolism of l-leucine, a branched-chain amino acid. in healthy individuals, approximately 60% of dietary l-leucine is metabolized after several hours, with roughly 5% (2–10% range) of dietary l-leucine being converted to hmb. the vast majority of l-leucine metabolism is initially catalyzed by the branched-chain amino acid aminotransferase enzyme, producing α-ketoisocaproate (α-kic). α-kic is mostly metabolized by the mitochondrial enzyme branched-chain α-ketoacid dehydrogenase, which converts it to isovaleryl-coa. isovaleryl-coa is subsequently metabolized by isovaleryl-coa dehydrogenase and converted to mc-coa, which is used in the synthesis of acetyl-coa and other compounds. during biotin deficiency, hmb can be synthesized from mc-coa via enoyl-coa hydratase and an unknown thioesterase enzyme, which convert mc-coa into hmb-coa and hmb-coa into hmb respectively. a relatively small amount of α-kic is metabolized in the liver by the cytosolic enzyme 4-hydroxyphenylpyruvate dioxygenase (kic dioxygenase), which converts α-kic to hmb. in healthy individuals, this minor pathway – which involves the conversion of l-leucine to α-kic and then hmb – is the predominant route of hmb synthesis. chemistry skeletal formula of butyric acid with the alpha, beta, and gamma carbons markedformula of β-hydroxy β-methylbutyric acid β-hydroxy β-methylbutyric acid is a monocarboxylic β-hydroxy acid and natural product with the molecular formula c5h10o3. at room temperature, pure β-hydroxy β-methylbutyric acid occurs as a transparent, colorless to light yellow liquid which is soluble in water. β-hydroxy β-methylbutyric acid is a weak acid with a pka of 4.4. its refractive index ( n 25°c λ = 589 n m _^ }} ) is 1.42. chemical structure β-hydroxy β-methylbutyric acid is a member of the carboxylic acid family of organic compounds. it is a structural analog of butyric acid with a hydroxyl functional group and a methyl substituent located on its beta carbon. by extension, other structural analogs include β-hydroxybutyric acid and β-methylbutyric acid. synthesis a variety of synthetic routes to β-hydroxy β-methylbutyric acid have been developed. the first reported chemical syntheses approached hmb by oxidation of alkene, vicinal diol, and alcohol precursors: in 1877, russian chemists michael and alexander zaytsev reported the preparation of hmb by oxidation of 2-methylpent-4-en-2-ol with chromic acid (h2cro4); in 1880 and 1889, schirokoff and reformatsky (respectively) reported that the oxidative cleavage of the vicinal diol 4-methylpentane-1,2,4-triol with acidified potassium permanganate (kmno4) yields hmb – this result is closest related to the first synthesis as cold dilute kmno4 oxidises alkenes to vicinal cis-diols which hot acid kmno4 further oxidises to carbonyl-containing compounds, and the diol intermediate is not obtained when hot acidic conditions are used for alkene oxidation. in other words, racemic 4-methylpentane-1,2,4-triol is a derivative of 2-methylpent-4-en-2-ol and β-hydroxy β-methylbutyric acid is a derivative of both; and, in 1892, kondakow reported the preparation of hmb by permanganate oxidation of 3-methylbutane-1,3-diol. first synthetic routes to β-hydroxy β-methylbutyric acid depending on the experimental conditions, cycloaddition of acetone and ketene produces either β-isovalerolactone or 4,4-dimethyloxetan-2-one, both of which hydrolyze under basic conditions to yield the conjugate base of hmb. the haloform reaction provides another pathway to hmb involving the exhaustive halogenation of the methyl-ketone region of diacetone alcohol with sodium hypobromite or sodium hypochlorite; diacetone alcohol is readily available from the aldol condensation of acetone. an organometallic approach to hmb involves the carboxylation of tert-butyl alcohol with carbon monoxide and fenton's reagent (hydrogen peroxide and ferrous iron). alternatively, hmb can be prepared through microbial oxidation of β-methylbutyric acid by the fungus galactomyces reessii. later synthetic routes to β-hydroxy β-methylbutyric acid detection in body fluids hmb concentrations measured in healthy individuals biofluid age group concentration sources mean range units blood plasmaadults (18+)4.00–10.0μm csfadults (18+)4.02.0–6.0μm sarcoplasmaadults (21–23)7.04.0–10.0μm breast milkadults (18+)–42–164μg/l urineadults (18+)–3.2–25.0μmol/mmol creatinine urinechildren (1–18)–0–68μmol/mmol creatinine the concentration of naturally produced hmb has been measured in several human body fluids using nuclear magnetic resonance spectroscopy, liquid chromatography–mass spectrometry, and gas chromatography–mass spectrometry methods. in the blood plasma and cerebrospinal fluid (csf) of healthy adults, the average molar concentration of hmb has been measured at 4.0 micromolar (μm). the average concentration of hmb in the intramuscular fluid of healthy men of ages 21–23 has been measured at 7.0 μm. in the urine of healthy individuals of any age, the excreted urinary concentration of hmb has been measured in a range of 0–68 micromoles per millimole (μmol/mmol) of creatinine. in the breast milk of healthy lactating women, hmb and l-leucine have been measured in ranges of 42–164 μg/l and 2.1–88.5 mg/l. in comparison, hmb has been detected and measured in the milk of healthy cows at a concentration of <20–29 μg/l. this concentration is far too low to be an adequate dietary source of hmb for obtaining pharmacologically active concentrations of the compound in blood plasma. in a study where participants consumed 2.42 grams of pure hmb-fa while fasting, the average plasma hmb concentration increased from a basal level of 5.1 μm to 408 μm after 30 minutes. at 150 minutes post-ingestion, the average plasma hmb concentration among participants was 275 μm. abnormal hmb concentrations in urine and blood plasma have been noted in several disease states where it may serve as a diagnostic biomarker, particularly in the case of metabolic disorders. the following table lists some of these disorders along with the associated hmb concentrations detected in urine or blood plasma. abnormal hmb concentrations measured in disease states medical condition biofluid age group concentration sources mean range units biotinidase deficiency†bloodadults (18+)9.50–19.0μm biotinidase deficiency†bloodchildren (1–13)88.010.0–166.0μm biotinidase deficiency†urinechildren (1–13)275.050.0–500.0μmol/mmol creatinine 3-methylglutaconic aciduria (type i)†urinechildren (1–13)200.0150.0–250.0μmol/mmol creatinine eosinophilic esophagitisurinechildren (1–13)247.40–699.4μmol/mmol creatinine gastroesophageal reflux diseaseurinechildren (1–13)119.85.5–234.0μmol/mmol creatinine hmg-coa lyase deficiency†urinechildren (1–13)2030.060.0–4000.0μmol/mmol creatinine mc-coa carboxylase deficiency†urinechildren (1–13)30350.01700.0–59000.0μmol/mmol creatinine a † indicates that the medical condition is a metabolic disorder. history the first reported chemical synthesis of hmb was published in 1877 by the russian chemists michael and alexander zaytsev. hmb was isolated from the bark of erythrophleum couminga (a madagascan tree) in 1941 by leopold ružička. the earliest reported isolation of hmb as a human metabolite was by tanaka and coworkers in 1968 from a patient with isovaleric acidemia. the effects of hmb on human skeletal muscle were first discovered by steven l. nissen at iowa state university in the mid-1990s. nissen founded a company called metabolic technologies, inc. (mti) around the time of his discovery, which later acquired six hmb-related patents that the company has used to license the right to manufacture and incorporate hmb into dietary supplements. when it first became available commercially in the late 1990s, hmb was marketed solely as an exercise supplement to help athletes and bodybuilders build muscle. mti subsequently developed two hmb-containing products, juven and revigor, to which abbott nutrition obtained the market rights in 2003 and 2008 respectively. since then, abbott has marketed juven as a medical food and the revigor brand of hmb as an active ingredient in food products for athletes (e.g., certain formulations of myoplex) and other medical foods (e.g., certain formulations of ensure).