RESUMO
Mycotoxins in milk are a public health concern and have to be regularly monitored. A survey on the presence of aflatoxin M1 (AFM1) and ochratoxin A (OTA) in raw bulk milk was conducted in 2003 in the northwest of France, the main French milk-producing basin. Randomly selected farms (n = 132) were characterized by a diet based on corn silage and containing a large proportion of on-farm produced cereals, feeding sources that are frequently contaminated by mycotoxins. Farms were surveyed twice in winter and in summer. At each sampling time, a trained surveyor completed a questionnaire recording farm management procedures and production traits. The AFM1 was found in 3 out of 264 samples but at levels (26 ng/L or less) that are below the European legislation limit of 50 ng/L. Traces of AFM1 (less than 8 ng/L) were also found in 6 other samples. The OTA was detected in 3 samples also at low levels, 5 to 8 ng/L. Farms that tested positive to the presence of mycotoxins, 12 in total including 6 farms that had traces of AFM1, differed from negative farms by a more extensive use of total mixed rations, 58 vs. 27%. In addition, the positive farms tended to have lower milk yields. Although the incidence of milk contamination with AFM1 and OTA at the farm level was low during the period studied, production and management data from the surveyed farms suggest a link between feeding management practices and mycotoxin contamination.
Assuntos
Aflatoxina M1/análise , Indústria de Laticínios/métodos , Leite/química , Ocratoxinas/análise , Ração Animal , Animais , Bovinos , Dieta/veterinária , Grão Comestível , França , Estações do Ano , Silagem , Zea maysRESUMO
Hepatic microsomal glucuronoconjugation of the hypolipidemic drug clofibric acid was characterized in human liver and compared to the acylglucuronide formation of an endogenous substrate, bilirubin. The affinity of UDP-glucuronosyltransferase for bilirubin was 15-fold higher than for clofibric acid; the Vmax for the transformation of the two substrates were similar. The analysis of the specific activity in 32 liver biopsies showed that glucuronidation of clofibric acid or bilirubin were comparable in man and in rat. However, UDP-glucuronosyltransferase activity towards clofibric acid exhibited a large interindividual variation in man. Sex or age did not influence the glucuronidation of bilirubin and clofibric acid. Among the drugs given to the patients only clofibrate was able to increase the bilirubin conjugation. No effect of alcohol or smoking on the conjugation of the two substrates was observed. The absence of correlation between UDP-glucuronosyltransferase activities towards clofibric acid and bilirubin together with the specific induction of bilirubin glucuronidation by clofibrate suggested that these arylcarboxylic substrates were conjugated by separate forms of UDP-glucuronosyltransferase in human.
Assuntos
Bilirrubina/análogos & derivados , Clofibrato/análogos & derivados , Ácido Clofíbrico/farmacocinética , Microssomos Hepáticos/metabolismo , Adulto , Fatores Etários , Idoso , Consumo de Bebidas Alcoólicas , Animais , Bilirrubina/metabolismo , Feminino , Glucuronosiltransferase/metabolismo , Humanos , Cinética , Masculino , Pessoa de Meia-Idade , Ratos , Valores de Referência , Fatores Sexuais , FumarRESUMO
Ochratoxin A is often found in the sera of people exposed to this mycotoxin in their food (cereals such as barley, coffee, wines, fruit juices, spices, products of animal origin such as pig and poultry offal). Ochratoxin A is suspected of playing a role in the Balkan Endemic Nephropathy, a nephropathology described in Balkan areas where ochratoxin A is often found in cereals and in pork-derived products. In North Africa like Tunisia where high incidence of chronic interstitial nephropathies of unknown aetiology are pointed out, the involvement of ochratoxin A was suspected but contradictory studies on the degree of human exposure did not succeed in evidencing the role of ochratoxin A. In the present work, sera from 47 volunteers hospitalised for nephropathic damages including bladder tumours (21 people), and from 62 patients hospitalised for disorders other than nephropathic ones, were analysed for ochratoxin A contents. The determination of ochratoxin A in sera was done by a validated immunoaffinity-HPLC method. Sera from unaffected population exhibited percentages of 74.2%, 22.6% and 3.2% containing ranges of ochratoxin A as <0.10-0.5 microg/l, 0.51-1.0 microg/l and above 1.0 microg/l respectively. For patients affected with renal diseases, percentages were 59.5%, 25.5% and 14.9% on the same ranges of ochratoxin A levels respectively. The average ochratoxin A concentration for patients with urinary tract disease excluding cancer patients was 0.99+/-1.28 microg/l while that for the non-nephropathic patients was 0.53+/-1.00 microg/l. However the average levels in the cancer patients was only 0.26+/-0.20 microg/l. Those results are in line with most of previously published works and did not confirm very high ochratoxin A contents found in other reports from same regions.
Assuntos
Nefropatia dos Bálcãs/sangue , Exposição Ambiental/análise , Monitoramento Ambiental/métodos , Micotoxinas/sangue , Ocratoxinas/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Afinidade de Anticorpos , Nefropatia dos Bálcãs/epidemiologia , Nefropatia dos Bálcãs/etiologia , Cromatografia de Afinidade , Cromatografia Líquida de Alta Pressão , Monitoramento Epidemiológico , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Micotoxinas/isolamento & purificação , Ocratoxinas/isolamento & purificação , Tunísia/epidemiologiaRESUMO
A method is described for the determination of ochratoxin A (OTA) in red wine and vinegar using an acidic chloroform extraction, an immmunoaffinity clean-up step, and a high-performance liquid chromatographic determination with fluorescence detection. The detection limit was estimated at 0.002 microg/liter. The mean recovery factors were found at 91.3 and 96.6% for wine and vinegar, respectively. Thirty-one samples of red wine originating from Mediterranean sea countries and 15 samples of vinegar were examined for the presence of OTA. All red wine samples contained OTA. Seventy-two percent of these samples were found to be contaminated over 0.1 microg/liter. Among them, nine samples contained ochratoxin A in the range of 0.5 to 3.4 microg/liter, 12 samples in the range of 0.10 to 0.50 microg/liter (median: 0.176 microg/liter), and 9 samples in the range of 0.010 to 0.100 microg/liter (median: 0.041 microg/liter). All 15 vinegar samples showed the presence of OTA. The most contaminated ones were three balsamic vinegar samples containing 0.156 microg/liter, 0.102 microg/liter, and 0.252 microg/liter of OTA. In the remaining 12 samples, ochratoxin A levels ranged from 0.008 microg/liter to 0.046 microg/liter (median: 0.012 microg/liter). These data are in good agreement with the hypothesis that wine originating from Southern countries might contain significant OTA concentration and showed the possible occurrence of traces of OTA in vinegar.
Assuntos
Ácido Acético/análise , Cromatografia Líquida de Alta Pressão/métodos , Ocratoxinas/análise , Vinho/análise , Fluorescência , Microbiologia de Alimentos , Sensibilidade e EspecificidadeRESUMO
A collaborative study was conducted to evaluate the effectiveness of an immunoaffinity column cleanup liquid chromatographic method for determination of aflatoxin M1 in milk at proposed European regulatory limits. The test portion of liquid milk was centrifuged, filtered, and applied to an immunoaffinity column. The column was washed with water, and aflatoxin was eluted with pure acetonitrile. Aflatoxin M1 was separated by reversed-phase liquid chromatography (LC) with fluorescence detection. Frozen liquid milk samples both naturally contaminated with aflatoxin M1 and blank samples for spiking, were sent to 12 collaborators in 12 different European countries. Test portions of samples were spiked at 0.05 ng aflatoxin M1 per mL. After removal of 2 noncompliant sets of results, the mean recovery of aflatoxin M1 was 74%. Based on results for spiked samples (blind pairs at 1 level) and naturally contaminated samples (blind pairs at 3 levels) the relative standard deviation for repeatability (RSDr) ranged from 8 to 18%. The relative standard deviation for reproducibility (RSDR) ranged from 21 to 31%. The method showed acceptable within- and between-laboratory precision data for liquid milk, as evidenced by HORRAT values at the low level of aflatoxin M1 contamination.
Assuntos
Aflatoxina M1/análise , Leite/química , Animais , Cromatografia de Afinidade , Imunoquímica , Indicadores e Reagentes , Padrões de Referência , Soluções , Espectrometria de FluorescênciaRESUMO
Two different immunoaffinity columns (IACs) were prepared for detection of staphylococcal enterotoxins (SETs) from dairy products. First, a specific IAC for staphylococcal enterotoxin A (SEA), IAC-1, was prepared by coupling monoclonal antibody (mAb) directed against SEA; second, a polyspecific IAC for SEA, staphylococcal enterotoxin B (SEB), staphylococcal enterotoxin C (SECs), and staphylococcal enterotoxin D (SED), IAC-2, was prepared by coupling a mixture of mAbs against SEA, SECs, and SED, and rabbit IgG against SEB. These columns were applied for detection of SETs in dairy products, after extraction, immunoaffinity chromatography, and enzyme immunosorbent assay (EIA). Overall recoveries from dairy products spiked with 1 ng SEA/25 g averaged 81.2% (range, 76-85%) on IAC-1. The repeated use of IAC-1 was then determined with good efficiency of 91.5%, in more than 10 runs. On the other hand, a recovery yield of 77% of SETs (SEA, SEB, SEC, and SED) from dairy products spiked with 2.5 ng of each enterotoxin per 25 g, was obtained with IAC-2. IAC-2 was also successfully subjected to the chromatography of naturally contaminated foods implicated in staphylococcal food poisoning outbreaks. This new extraction-concentration-immunoaffinity-chromatography method (ECIC) is very useful for improving staphylococcal enterotoxin detection and eliminating matrix effect in EIA of dairy products.
Assuntos
Laticínios/análise , Enterotoxinas/análise , Staphylococcus aureus/química , Anticorpos/química , Queijo/análise , Cromatografia de Afinidade , Diálise , Reutilização de Equipamento , Contaminação de Alimentos/análise , Técnicas Imunoenzimáticas , Indicadores e ReagentesRESUMO
AIM: Immunological tools used to detect staphylococcal enterotoxins (SEs) in foods are numerous. The aim of this study was to evaluate, on naturally contaminated milk product samples, the performance of the Vidas SET2, in comparison to the Transia plate SET. METHODS AND RESULTS: The Vidas SET2 was compared with the Transia plate SET on supernatants of Staphylococcus aureus isolates and on naturally contaminated milk products. It is noteworthy that when using IgG rabbit treatment, both kits can be considered as equivalent to detect enterotoxins in naturally contaminated milk products. CONCLUSIONS: This study demonstrated that the Vidas SET2 performance is similar to that of Transia plate SET kit, when a rabbit IgG treatment step is used before detection step. This additional treatment significantly decreased, from 42% to 8%, the rate of positive deviations observed using the Transia plate SET detection kit. SIGNIFICANCE AND IMPACT OF THE STUDY: The Vidas SET2 was clearly found as more specific, when no preliminary rabbit IgG treatment was used, and which results in a better workflow when a large number of samples have to be analysed within a few days. Considering the results obtained, the Vidas SET2 detection kit can be used to assess the safety of milk products for SEs.
Assuntos
Laticínios/microbiologia , Enterotoxinas/análise , Análise de Alimentos/normas , Leite/microbiologia , Staphylococcus aureus/isolamento & purificação , Animais , Anticorpos Monoclonais , Técnicas de Química Analítica/métodos , Análise de Alimentos/métodos , Microbiologia de Alimentos , Coelhos , Staphylococcus aureus/metabolismoRESUMO
Milk products such as cheeses may be contaminated by aflatoxin M1 when manufactured with milk from dairy cattle that have consumed aflatoxin B1-contaminated feeds. The usefulness of immunoaffinity columns to determine aflatoxin M1 content in many kinds of cheeses with very good recoveries is demonstrated. The analysis of aflatoxin M1 in a 1990 to 1995 limited survey in France shows that the occurrence of this mycotoxin in cheeses is rather infrequent. With the exception of samples from 1989 to 1990 when aflatoxin B1-contaminated maize meals were incidentally imported to supplement dairy cattle feed, very few samples were found with above 0.200 µg of aflatoxin M1 per kg of cheese, the maximum acceptable level.
RESUMO
A collaborative study was conducted under the auspices of the International Dairy Federation (IDF), the International Union of Pure and Applied Chemistry (IUPAC) and the International Atomic Energy Agency (IAEA), a collaborative Food and Agricultural Organization (FAO) body fully to validate a method combining immunoaffinity clean-up to thin-layer chromatography for the determination of aflatoxin M(1) in milk. Work was done in order to afford those laboratories not equipped with high-performance liquid chromatography, mainly from developing countries, with a simplified but fully validated method as an alternative to the European validated immunoaffinity-high performance liquid chromatography method published as an EN ISO Standard 14501, February 1999. The validation study was carried out on samples of aflatoxin M(1)-contaminated milk and milk powder at levels close to the tolerable level of 0.5 microg l(-1) as recommended by the Codex Alimentarius and to the regulatory level of 0.05 microg l(-1) as laid down by the European Commission. Fourteen laboratories representing 11 countries participated in the trial. The relative standard deviations for repeatability and reproducibility based on raw data were in the range 27-48 and 35-54%, respectively. The recovery rate varied from 32 to 120%. The mishandling of two crucial steps of the protocol such as matrix sample reconstitution and extract evaporation could explain the wide variation of the recovery rate. For this reason, data were then corrected for recovery. Consequently, the relative standard deviations for repeatability and reproducibility were recalculated after correction for recovery and were in the range 26-54 and 34-53%, respectively. The method will be published as a standard ISO/DIS 14674--IDF 190.
Assuntos
Aflatoxina M1/análise , Contaminação de Alimentos/análise , Leite/química , Animais , Cromatografia de Afinidade/métodos , Cromatografia em Camada Fina/métodos , Análise de Alimentos/métodos , Análise de Alimentos/normas , Cooperação Internacional , Laboratórios/normas , Reprodutibilidade dos TestesRESUMO
Diarrhetic Shellfish Poisoning (DSP) is a severe gastro-intestinal disease caused by consumption of seafood contaminated by microalgal toxins, mainly okadaic acid (OA) and structurally related toxins, dinophysistoxins (DTXs). Regulatory monitoring is generally based on rodent bioassays which, however, present some technical and ethical disadvantages. The most promising technique of analysis of these toxins involves an HPLC separation with spectrofluorimetric detection after derivatization of the toxins with a fluorescent reagent. The lack of specificity of the extraction procedure (liquid-liquid partition), and the presence of interfering compounds in the matrix, does not allow the determination and the quantification of low amounts of toxins in seafood. In this paper, the authors report the development and the characterization of immunoaffinity columns (IAC), which were elaborated using anti-okadaic acid monoclonal antibodies, for a specific retention of the OA group of toxins. The coupling yield and the stability of these columns were investigated as well as their capacity to remove interfering compounds. Cross-reactivity was observed between the antibodies and the DTX-1 and the DTX-2, allowing the detection of the different toxins in a single analysis. Different spiked (1 microgram OA/g) or naturally-contaminated (mussel digestive gland: 2 micrograms OA/g; algae: 165 micrograms OA/g) matrices were tested. The recovery for OA varied from 55 to 95% according to the matrices. The IAC purification was then included as a step of a global [IAC/HPLC/spectrofluorimetric detection] method and the performance of the method was evaluated. Estimations of the linearity and the accuracy (percentages of the presumptive response for OA in the range +101% to +114%) were satisfactory in accordance with the method validation criteria.
Assuntos
Dinoflagellida , Doenças Transmitidas por Alimentos/etiologia , Toxinas Marinhas/análise , Alimentos Marinhos , Animais , Cromatografia de Afinidade , Cromatografia Líquida de Alta Pressão , Fluorometria , Humanos , Ácido Okadáico/análise , Piranos/análiseRESUMO
A method using immunoaffinity as a purification step for the determination of aflatoxin M1 in cheese is described. A simple solvent extraction with dichloromethane followed by a washing step with N-hexane gives a prepurified extract. A comparison between two ways of aflatoxin M1 purification, by solid-phase extraction clean-up and by immunoaffinity, was carried out. The use of immunoaffinity columns containing monoclonal antibodies against aflatoxin M1 gives the best result, i.e. a very clean preparation containing purified aflatoxin M1. The quantification of aflatoxin M1 is then performed by high performance liquid chromatography using fluorometric detection. This method was successfully carried out on naturally-contaminated and spiked cheeses. Recoveries are about 75%. The limit of quantification is 0.020 microgram of aflatoxin M1 per kg of cheese. This method seems suitable for use in monitoring programmes for aflatoxin M1 contamination in dairy products such as cheese.
Assuntos
Aflatoxina M1/isolamento & purificação , Queijo/análise , Cromatografia de Afinidade , Contaminação de Alimentos , Inspeção de Alimentos/métodos , Cromatografia Líquida de Alta Pressão , Análise de AlimentosRESUMO
After induction by phenobarbital and 3-methylcholanthrene, UDP-glucuronosyltransferase involved mainly in the conjugation of planar substrates was purified. Compared to the microsomal enzyme, the purified protein exhibited less affinity towards the substrates, but the corresponding Vmaxs were increased. These results were attributed to a change in the lipid environment of the purified enzyme. The conjugation rate for 4-hydroxycoumarine was 15-45 times less than that measured for the 7-hydroxyisomer with the microsomal or the purified enzymes. Immunoprecipitation studies of the enzyme revealed that the two compounds were transformed by the same enzyme, or metabolized by two separate enzymes presenting the same antigenic site. The orientation of the hydroxyl group of planar aglycones in the active site is the determinant for the efficiency of catalysis.
Assuntos
Glucuronosiltransferase/biossíntese , Microssomos Hepáticos/enzimologia , Animais , Indução Enzimática/efeitos dos fármacos , Técnicas In Vitro , Cinética , Metabolismo dos Lipídeos , Masculino , Metilcolantreno/farmacologia , Fenobarbital/farmacologia , Ratos , Ratos EndogâmicosRESUMO
Glucuronidation of 4-nitrophenol, nopol (a monoterpenoid alcohol) and bilirubin, which in the rat, are catalyzed by three different enzymes, has been examined in liver biopsies from patients with various liver diseases, in particular cholestasis. These different activities were not correlated, which strongly suggests that at least three independently regulated forms of UDP-glucuronosyltransferases were present in the microsomes. Non ionic detergents (Triton X100, Emulgen 911) and deoxycholate produced similar activation (more than 2-fold) of the glucuronidation of 4-nitrophenol. Amphipathic substances, such as CHAPS (3-[3-cholamidopropyl-dimethylammonio]-1-propane sulfonate), and lysophosphatidylcholines maximally increased this UDP-glucuronosyltransferase activity, the most potent being oleoyl lysophosphatidylcholine (4-fold increase). Discriminant analysis of the data revealed no correlation between the three different UDP-glucuronosyltransferase activities and the age or sex of the patients. A good correlation was found on multidimensional analysis between form 1 of the enzyme (4-nitrophenol glucuronidation) and, in decreasing order of magnitude, epoxide hydrolase (measured with benzo(a)pyrene-4,5-oxide as substrate), cytochrome P-450, 7-ethoxycoumarin deethylase, aspartate aminotransferase and gamma-glutamyltransferase (r = 0.89); and between Form 3 of the enzyme (bilirubin glucuronidation) and NADPH cytochrome c reductase, alkaline phosphatase, (r = 0.81). These relationships may reflect the differential variation in enzymatic activities in various hepato-biliary diseases.
Assuntos
Colestase/enzimologia , Glucuronosiltransferase/metabolismo , Isoenzimas/metabolismo , Fígado/enzimologia , O-Dealquilase 7-Alcoxicumarina , Alanina Transaminase/metabolismo , Fosfatase Alcalina/metabolismo , Aspartato Aminotransferases/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Humanos , NADPH-Ferri-Hemoproteína Redutase/metabolismo , Oxigenases/metabolismo , gama-Glutamiltransferase/metabolismoRESUMO
In 1992, the European Union set up a network of National Reference Laboratories and charged the Community Reference Laboratory with the responsibility to design a proficiency testing scheme for assessing the analytical ability of laboratories involved in the official control of aflatoxin M1 in milk. Since 1996, two exercises of proficiency testing have been performed on samples of milk powder and liquid milk at various levels of aflatoxin M1 contents. The trials were conducted according to ISO Guide 43, in particular for the homogeneity testing of sample batches and for the calculation of laboratory z-scores. The National Reference Laboratories officially designated by their governments participated in this programme. Samples were naturally-contaminated milk obtained by feeding cows with aflatoxin B1-contaminated feed. The levels of aflatoxin M1 in the samples ranged from 0.2 to 0.7 microg/kg in milk powder and from 0.05 to 0.07 microg/l in liquid milk. These levels were chosen as being close to the European Union-regulated limit of 0.05 microg of aflatoxin M1 per litre. The results produced by laboratories were compiled and statistically analysed to detect any outlying results and to calculate the individual z-scores. Except for one laboratory in each exercise, all laboratories exhibited acceptable or questionable z-scores. The interlaboratory relative standard deviation for reproducibility (RSDR) obtained for both 1996 and 1998 exercises were in the range 15.7-30.3%. Compared with other published studies, this indicates a very good precision for the performance of this laboratory network in the analysis of traces of aflatoxin M1 in milk.
Assuntos
Aflatoxina M1/análise , Contaminação de Alimentos , Leite/química , Animais , União Europeia , Análise de Alimentos/legislação & jurisprudência , Análise de Alimentos/normas , Humanos , Laboratórios/normas , Controle de Qualidade , Valores de Referência , Reprodutibilidade dos TestesRESUMO
Ochratoxin A is a carcinogen and nephrotoxin which can enter the food chain resulting in human exposure. As pig herds are exposed to ochratoxin A through their feed, their kidneys, livers and pork meat are considered as a possible route of exposure for humans. France, an important producer of pork and pork products, set up a national monitoring programme which included the training of six routine public laboratories in the analysis of ochratoxin A using an immunoaffinity step followed by a HPLC-fluorimetric detection. The programme randomly sampled 300 healthy and 100 nephropathic pig kidneys in 1997 and 710 healthy pig kidneys in 1998. Less than 10% of samples were significantly contaminated by ochratoxin A : in the 1997 survey, 1% of samples contained 0.40-1.40 microg kg(-1) of ochratoxin A and in the 1998 survey 7.6 % exhibited ochratoxin A levels in the range 0.5-5 microg kg(-1). In the case of nephropathic kidneys, only traces of ochratoxin A (0.16 to 0.48 microg kg(-1)) were detected in six samples out of 100. Even if not a major route of exposure for humans, pigs are clearly exposed to this mycotoxin and monitoring of pork products and of feed for swine is necessary.
Assuntos
Carcinógenos/análise , Contaminação de Alimentos/análise , Contaminação de Alimentos/prevenção & controle , Rim/química , Micotoxinas/análise , Ocratoxinas/análise , Animais , Cromatografia Líquida de Alta Pressão , França , Nefropatias/metabolismo , Nefropatias/patologia , Laboratórios , Espectrometria de Fluorescência , SuínosRESUMO
Simple, sensitive and selective enzyme immunoassays (ELISA) for monitoring urinary dextromethorphan and its major metabolite, dextrorphan, were developed. Dextromethorphan and dextrorphan hemisuccinates were linked to bovine serum albumin and specific antisera against each immunogen were raised in rabbits. The sensitivity of the ELISA was high (limit of detection 740 and 600 pg.ml-1 for dextromethorphan and dextrorphan, respectively). Intra- and interassay variation was less than 10%, and cross-reactivity between the two compounds was less than 1%. The ELISA was employed to phenotype 216 French subjects. The frequency of poor metabolizers was 5.1%.
Assuntos
Dextrometorfano/metabolismo , Ensaio de Imunoadsorção Enzimática/métodos , Polimorfismo Genético , Cromatografia Líquida de Alta Pressão , Feminino , França , Humanos , Masculino , Oxirredução , FenótipoRESUMO
We describe a competitive enzyme immunoassay, the ExtrAvidin-biotin system, for determining osteocalcin in human serum or plasma. Antibodies were raised against bovine osteocalcin. Binding of the antibodies to osteocalcin was calcium-dependent. Limit of detection is 0.07 nmol/L (0.4 microgram/L). The standard curve for method is linear between 0.3 and 17.6 nmol/L (1.9 and 100 micrograms/L). Interassay CV over the range 0.9 to 14.8 nmol/L (5.3 to 84 micrograms/L) is 7.5% to 11.7%. Analytical recovery is 105% +/- 5% (mean +/- SD). The measurement, which is adapted to microtiter plates, requires only 20 microL of serum and 5 h. The coefficient of correlation between the concentrations measured by this method and by a commercially available radioimmunoassay kit (CIS Biointernational) is 0.91. Osteocalcin can be measured in serum or heparinized plasma. Hemolysis (174 mumol/L hemoglobin) reduces osteocalcin concentration by 54%. High concentrations of triglycerides (7 mmol/L) give an overestimation of 63%. Serum concentrations of osteocalcin measured in 130 healthy subjects (ages 15-64 years) and 86 children (ages 4-14 years) were 1.4 +/- 0.8 and 4.0 +/- 1.5 nmol/L (8.1 +/- 4.6 and 22.5 +/- 8.6 micrograms/L), respectively (mean +/- SD).