Urine homogentisic acid and tyrosine: Simultaneous analysis by liquid chromatography tandem mass spectrometry

Hughes, Andrew T.; Milan, Anna M.; Christensen, Peter; Ross, Gordon; Davison, Andrew S.; Gallagher, James A.; Dutton, John; Ranganath, L.R.

doi:10.1016/j.jchromb.2014.06.002

Cited by 59 publications

(56 citation statements)

References 25 publications

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“…Reproducibility test was repeated after 2 week storage of DUS, with no appreciable loss compared to fresh spots (recovery 93.7-105.6%; CV ¼ 8.7%). These data and values expressed as mmol/mol creatinine (1,741 AE 541) or mmol/24 h (17.0 AE 7.4) were in agreement with those reported in the literature (Bory et al 1989;Mannoni et al 2004;Hughes et al 2014;Ranganath et al 2016). No appreciable HGA could be detected in DUS or urine samples from controls.…”

Section: Hplc Analysis Of Urines and Dus (Methods 2)supporting

confidence: 91%

“…Biochemical diagnosis is based on the detection of HGA in body fluids, mainly urine, by different analytical methods such as high performance liquid chromatography (Bory et al 1989), gas chromatography, capillary electrophoresis, mass spectrometry (Hughes et al 2014), enzymatic and spectrophotometric methods (Tokuhara et al 2014). Such sensitive methods can only be performed in specialized laboratories with appropriate equipments.…”

Section: Discussionmentioning

confidence: 99%

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Quick Diagnosis of Alkaptonuria by Homogentisic Acid Determination in Urine Paper Spots

et al. 2016

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Objectives: Two methods are described for homogentisic acid (HGA) determination in dried urine spots (DUS) on paper from Alkaptonuria (AKU) patients, devised for quick early diagnosis. AKU is a rare autosomal recessive disorder caused by deficiency of homogentisate 1,2-dioxygenase, yielding in accumulation of HGA. Its massive excretion causes urine darkening by exposure to air or alkalinization, and is a diagnostic marker. The deposition of polymers produced after HGA oxidation within the connective tissues causes ochronotic arthritis, a degenerative joint disease manifesting in adulthood and only rarely in childhood. No early diagnosis is usually accomplished, awareness following symptom development.Design and methods: Two methods were designed for HGA determination in DUS: (1) a rapid semi-quantitative reliable method based on colour development in alkali and quantification by comparison with dried paper spots from HGA solutions of known concentration and (2) a quantitative and sensitive HPLC-linked method, previously devised for purine and pyrimidine analysis in urine and plasma.Results: Colour intensity developed by DUS after alkali addition was proportional to HGA concentration, and calculated amounts were in good agreement with quantitative analysis performed by RP-HPLC on DUS and on urines as such.Conclusions: DUS, often used for different diagnostic purpose, are easily prepared and safely delivered. The simple and quick colour method proposed provides reliable HGA assessment and is fit for large screening. HGA concentration determined in 10 AKU patient DUS by both methods 1 and 2 was in agreement with direct urine assay and in the range reported by literature.A reliable HGA quantification based on colour development in paper urine spots is validated by HPLC-linked HGA quantification, and proposed as a quick diagnostic tool for AKU patients.

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Section: Hplc Analysis Of Urines and Dus (Methods 2)supporting

confidence: 91%

Section: Discussionmentioning

confidence: 99%

Quick Diagnosis of Alkaptonuria by Homogentisic Acid Determination in Urine Paper Spots

et al. 2016

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“…The urine and serum samples were analyzed according to the liquid chromatography-mass spectrometry methods described by Hughes et al (2014Hughes et al ( , 2015.…”

Section: Measurementsmentioning

confidence: 99%

Relationship Between Serum Concentrations of Nitisinone and Its Effect on Homogentisic Acid and Tyrosine in Patients with Alkaptonuria

et al. 2015

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Background: Alkaptonuria (AKU) is a serious genetic disease due to a defect in tyrosine metabolism, leading to increased serum levels of homogentisic acid (HGA). Nitisinone decreases HGA in AKU, but the concentration-response relationship has not been previously reported.Objectives: To determine the relationship between serum concentrations of nitisinone and the effect on both HGA and tyrosine; secondly to determine steady-state pharmacokinetics of nitisinone in AKU patients.Method: Thirty-two patients with AKU received either 1, 2, 4, or 8 mg nitisinone daily. Urine and serum HGA and serum tyrosine and nitisinone were measured during 24 h at baseline (before first dose) and after 4 weeks of treatment.Results: Nitisinone pharmacokinetics (area under the curve [AUC] and maximum concentrations [C max ]) were dose proportional. The median oral clearance determined in all patients, irrespective of dose, was 3.18 mL/hÁkg (range 1.6-6.7).Nitisinone decreased urinary excretion of HGA in a concentration-dependent manner, with a maximum effect seen at average nitisinone concentrations of 3 mmol/L. The association between nitisinone and tyrosine concentrations was less pronounced. Serum levels of HGA at Week 4 were below the limit of quantitation in 65% of samples, which prevented determination of the relationship with nitisinone concentrations.Conclusion: Nitisinone exhibits dose-proportional pharmacokinetics in the studied dosage interval. Urinary excretion of HGA decreases in a concentration-dependent manner, while the increase in tyrosine is less clearly related to nitisinone concentrations.

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“…Urine samples were extracted and analyzed within 1-3 months after sample collection. HGA is known to be unstable in alkaline conditions (Hughes et al 2014). We tested stability under our own storage times and conditions by comparing the HGA concentration from five different urine samples extracted shortly after collection, compared to aliquots of the same samples stored for 3 months at À80 C prior to extraction and analysis (Supplementary Table 1).…”

Section: Sample Storagementioning

confidence: 99%

Metabolic Effects of Increasing Doses of Nitisinone in the Treatment of Alkaptonuria

et al. 2015

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Alkaptonuria is an autosomal recessive disease involving a deficiency of the enzyme homogentisate dioxygenase, which is involved in the tyrosine degradation pathway. The enzymatic deficiency results in high concentrations of homogentisic acid (HGA), which results in orthopedic and cardiac complications, among other symptoms. Nitisinone (NTBC) has been shown to effectively treat alkaptonuria by blocking the conversion of 4-hydroxyphenylpyruvate to HGA, but there have been concerns that using doses higher than about 2 mg/day could cause excessively high levels of tyrosine, resulting in crystal deposition and corneal pathology. We have enrolled seven patients in a study to determine whether higher doses of NTBC were effective at further reducing HGA levels while maintaining tyrosine at acceptable levels. Patients were given varying doses of NTBC (ranging from 2 to 8 mg/day) over the course of between 0.5 and 3.5 years. Urine HGA, plasma tyrosine levels, and plasma NTBC were then measured longitudinally at various doses. We found that tyrosine concentrations plateaued and did not reach significantly higher levels as NTBC doses were increased above 2 mg/day, while a significant drop in HGA continued from 2 to 4 mg/day, with no significant changes at higher doses. We also demonstrated using untargeted metabolomics that elevations in tyrosine from treatment resulted in proportional elevations in alternative tyrosine metabolic products, that of N-acetyltyrosine and γ-glutamyltyrosine.

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Urine homogentisic acid and tyrosine: Simultaneous analysis by liquid chromatography tandem mass spectrometry

Cited by 59 publications

References 25 publications

Quick Diagnosis of Alkaptonuria by Homogentisic Acid Determination in Urine Paper Spots

Quick Diagnosis of Alkaptonuria by Homogentisic Acid Determination in Urine Paper Spots

Relationship Between Serum Concentrations of Nitisinone and Its Effect on Homogentisic Acid and Tyrosine in Patients with Alkaptonuria

Metabolic Effects of Increasing Doses of Nitisinone in the Treatment of Alkaptonuria

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