Determination Of Ketone Bodies Research Articles

The correlation between higher level of ketone bodies in the blood and the ketosis clinical signs of the Holstein highproductive cows

Ketosis of highly productive cows is one of the most common metabolic pathologies, the etiological basis of which is the negative energy balance of the animal's body. The consequences of this disease is a deterioration in the health and productive qualities of the animal. Therefore, it is necessary to carry out timely and comprehensive diagnostics. The aim of these studies was to analyze the level of ketone bodies in the blood of highly productive Holstein cows and their clinical condition in the first three weeks of milk production. The studies were carried out in one of the livestock farms of the Leningrad region on 35 highly productive cows (the annual milk yield is more than 10,000 liters of milk) of the Holstein breed щт 5, 10, 15 and 20 days after calving. At the same time, the clinical manifestation of ketosis and concomitant diseases, as well as changes in the dynamics of the level of ketone bodies in the blood, were taken into account using test strips for the determination of ketone bodies in the Optium Xceed glucometer. According to the results of the studies, the average level of ketone bodies in the blood of cows on the 5th day after calving was 1.59 ± 1.08 mmol / l, which corresponded to the norm. In cows, on the 10-15th day after calving, an increase in the level of ketone bodies was noted by 1.3-1.4 times and averaged 2.1 ± 1.14 and 2.25 ± 1.4 mmol / l, respectively. On the 20th day after calving, the level of ketone bodies stabilized. It was equal to 1.55 ± 1.22 mmol / l. During the clinical examination of the animals, no obvious signs of ketosis were found. Throughout the experiment, 31% of the cows have had postpartum endometritis of first degree (according to the grades adopted at the farm: I, II, and III degrees), 2.8% - second degree, and 2.8% - third degree endometritis. Mastitis was registered in 22.7% of cows, and purulent pododermatitis in 2.8%. In addition, the animals with an increased level of ketone bodies were not recorded clinical manifestations of ketosis and concomitant diseases. And the cows with high level of ketone bodies in the blood was noted the presence of clinical signs of concomitant diseases (mastitis, endometrium, etc.). We can suggested the absence of a characteristic relationship between the content of ketone bodies in the animal's body and its health. It could be necessary to continue research on a larger livestock of animals and it should be pay an attention on the farm’s management, cows’ production schedule (milking, feeding, rest) and their productivity as well.

Determination of somatic status of girls with chronic vulvovaginitis in the neutral period of life

The objective: a determination of the spread of somatic pathology and polymorphism of the MSM6 gene in the development of chronic recurrent vulvovaginitis in girls. Materials and methods. The study involved 54 girls 1–6 years old. Inclusion criteria: complaints of patients on itching or pain in the genital area, dysuria, the presence of synechiae of the vulva, clinical manifestations of vulvitis during the visual examination. General laboratory diagnostic methods were used (blood test, general urine analysis with determination of ketone bodies, bacterioscopic examination of vaginal smears, vaginal culture for flora and antibiotic sensitivity, determination of glucose levels in blood serum and urine). For diagnosis of lactose malabsorption by polymerase chain reaction, a mutation in the MCM6 gene was studied, the genetic marker of which is C (-13910). Results. The somatic status of patients with chronic vulvovaginitis was presented by pathology of the gastrointestinal digestive tract in 68.5 % cases, atopic dermatitis – 53.7 %, diseases of the respiratory system – 33.3 %, infectious processes of the kidneys and urinary tract – 24.1 %. 77.6 % of girls had nondiabetic acetone syndrome, 75.9 % – dysmetabolic nephropathy. The presence of lactose malabsorption was revealed by homozygosity for the MCM6 gene mutation (C/C) in 26.1% of patients, heterozygosity for the MCM6 gene mutation (C/T) - in 56.5 %, the absence of the MCM6 gene mutation (T/T) was in 8 (17.4 %). Conclusions. In girls in the neutral period of life with chronic vulvovaginitis, non-infectious forms of the disease were found, which were not independent pathology, but associated with other pathological processes of metabolic disorders. Taking into account the obtained data on lactose malabsorption, we consider to include in the examination the algorithm of the verification of the MCM6 gene mutation and examination of urine for lactosuria and blood with lactose load.

Determination of ketone bodies in biological samples via rapid UPLC-MS/MS

Efforts to enhance wellness and ameliorate disease via nutritional, chronobiological, and pharmacological interventions have markedly intensified interest in ketone body metabolism. The two ketone body redox partners, acetoacetate (AcAc) and D-β-hydroxybutyrate (D-βOHB) serve distinct metabolic and signaling roles in biological systems. A highly efficient, specific, and reliable approach to simultaneously quantify AcAc and D-βOHB in biological specimens is lacking, due to challenges of separating the structural isomers and enantiomers of βOHB, and to the chemical instability of AcAc. Here we present a single UPLC-MS/MS method that simultaneously quantifies both AcAc and βOHB using independent stable isotope internal standards for both ketones. This method incorporates one sample preparation step requiring only 7 min of analysis per sample. The output is linear over three orders of magnitude, shows very low limits of detection and quantification, is highly specific, and shows favorable recovery yields from mammalian serum and tissue samples. Tandem MS discriminates D-βOHB from structural isomers 2- or 4-hydroxybutyrate as well as 3-hydroxyisobutyrate (3-HIB). Finally, a simple derivatization distinguishes D- and L-enantiomers of βOHB, 3-HIB, and 2-OHB, using the same rapid chromatographic platform. Together, this simple, efficient, reproducible, scalable, and all-encompassing method will support basic and clinical research laboratories interrogating ketone metabolism and redox biochemistry.

Determination of Ketone Bodies in Blood by Headspace Gas Chromatography-Mass Spectrometry

A gas chromatography-mass spectrometry (GC-MS) method for determination of ketone bodies (β-hydroxybutyrate, acetone, and acetoacetate) in blood is presented. The method is based on enzymatic oxidation of D-β-hydroxybutyrate to acetoacetate, followed by decarboxylation to acetone, which was quantified by the use of headspace GC-MS using acetone-(13)C(3) as an internal standard. The developed method was found to have intra- and total interday relative standard deviations < 10% for acetone+acetoacetate levels (∼25 to 8300 μM) and D-β-hydroxybutyrate levels (∼30 to 16500 μM). Recovery values varied from 98 to 107%, demonstrating the suitability of the method for measuring ketone bodies over a wide concentration range. The method has been applied to cases in which ketoacidosis was suspected as the cause of death in diabetics or chronic alcoholics, as well as to cases in which another cause of death was identified.

High-performance liquid chromatography determination of ketone bodies in human plasma by precolumn derivatization with p-nitrobenzene diazonium fluoroborate

We developed and validated a sensitive and convenient high-performance liquid chromatography (HPLC) method for the specific determination of ketone bodies (acetoacetate and d-3-hydroxybutyrate) in human plasma. p-Nitrobenzene diazonium fluoroborate (diazo reagent) was used as a precolumn derivatization agent, and 3-(2-hydroxyphenyl) propionic acid was used as an internal standard. After the reaction, excess diazo reagent and plasma proteins were removed by passing through a solid-phase cartridge (C 18). The derivatives retained on the cartridge were eluted with methanol, introduced into the HPLC system, and then detected with UV at 380 nm. A calibration curve for acetoacetate standard solution with a 20-μl injection volume showed good linearity in the range of 1 to 400 μM with a 0.9997 correlation coefficient. For the determination of d-3-hydroxybutyrate, it was converted to acetoacetate before reaction with the diazo reagent by an enzymatic coupling method using d-3-hydroxybutyrate dehydrogenase and lactate dehydrogenase. A calibration curve for d-3-hydroxybutyrate standard solution also showed good linearity in the range of 1.5 to 2000 μM with a 0.9988 correlation coefficient. Analytical recoveries of acetoacetate and d-3-hydroxybutyrate in human plasma were satisfactory. The method was successfully applied to samples from diabetic patients, and results were consistent with those obtained using the thio-NAD enzymatic cycling method used in clinical laboratories.

Headspace SPME–GC Method for Acetone Analysis and its Biomedical Application

A new method for extraction and analysis of acetone in human urine based on headspace solid phase microextraction using a mixture of activated carbon and zeolite as sorbents in a PVC matrix coated on a silver wire and its application to the determination of ketone bodies is described. Unlike commercial fibers, which are coated on fused silica, the coating adheres strongly to the silver wire and is thermally stable up to 250 °C. After optimization of coating composition and microextraction conditions the fiber was used for the analysis of acetone in human urine.

DETERMINAÇÃO DE CORPOS CETÔNICOS NA URINA COMO FERRAMENTA PARA O DIAGNÓSTICO RÁPIDO DE CETOSE SUBCLÍNICA BOVINA E RELAÇÃO COM A COMPOSIÇÃO DO LEITE

Nas primeiras semanas de lactação as vacas de alta produção apresentam severas mudanças metabólicas, em virtude do déficit energético derivado do baixo consumo de matéria seca e da alta produção, constituindo fator predisponente para a cetose. As perdas econômicas ocasionadas pela cetose subclínica não têm sido adequadamente avaliadas. O presente trabalho teve por objetivo relacionar a presença de corpos cetônicos na urina com os níveis plasmáticos de beta-hidroxibutirato (BHB) e o conteúdo de sólidos totais no leite. Foram utilizadas 140 vacas multíparas da raça Holandesa de cinco rebanhos com produção intensiva nas condições do Sul do Brasil. Os animais foram amostrados nas semanas 2, 5, 8 e 11 de lactação. Em cada período foram coletadas amostras de urina, amostras de sangue para a quantificação de BHB e amostras de leite para a determinação de sólidos totais. Na urina foram aplicadas quatro tiras reativas para a determinação de corpos cetônicos pelo método do nitroprussiato de sódio e foi determinado o pH. A proporção de vacas positivas a cetonúria foi de 11,2%. Valores de BHB acima de 1,2 mmol/L (ponto de corte para cetose) estiveram presentes em 12,4% das vacas. Concluísse que a cetose subclínica pode ser monitorada através tiras reagentes de corpos cetônicos na urina. Não foi encontrada correlação entre os sólidos totais no leite e o estado de cetose do animal.

Relationship between concentration of citrate and ketone bodies in cow's milk.

The authors' hypothesis is that the members of the tricarboxylic acid cycle (TCA cycle) such as citrate decrease in association with increased ketone body formation. To prove this hypothesis the connection between ketone bodies and citrate formation of milk was studied. A fluorimetric method was used to determine citrate and a headspace sampling gas chromatographic (GC) method was developed for determination of ketone bodies. Under real conditions of milk sampling, transport and storage, preserved milk samples of 119 clinically healthy dairy cows obtained in the 48 hours after milking were investigated. A low level of acetoacetate (ACAC) was found in all samples. This fact can be explained by the spontaneous decarboxylation of acetoacetate during sample storage (previously decarboxylised acetoacetate = pdACAC) and, consequently, the majority of the amount of acetoacetate in the samples (AC + pdACAC) appeared in the measured acetone concentrations. Based on the measured acetone concentration of milk samples two groups were formed retrospectively: HA (high-acetone) group (n = 41) with an AC + pdACAC concentration of > 0.4 mmol/l and a LA (low-acetone) group (n = 78) with an AC + pdACAC level of < or = 0.4 mmol/l. In the milk of cows of Group HA a positive correlation (r = +0.623) and linear connection between acetone (AC + pdACAC) and beta-hydroxybutyrate (BOHB) levels was found [BOHB = 2.491 + 0.586 x (pdAC + ACAC)]. Furthermore, in this group a negative correlation between citrate and BOHB and AC + pdACAC was also established (r = -0.579). Focusing on the results of this group the authors found a significant drop of AC + pdACAC and citrate during the metabolically critical first 1-4 weeks of lactation. For this reason they suggest that simple, easy, automated methods (i.e. flow injection analysis, Fourier transformation infrared analysis) should be introduced for the simultaneous determination of acetone and citrate concentration in milk to make the evaluation of the energy status of high-producing dairy cows easier and more certain.

Determination of ketone bodies in postmortem blood by head-space gas chromatography

A method describing determination of the physiological ketone bodies: acetone, acetoacetate, and d-β-hydroxybutyrate in postmortem blood by head-space gas chromatography. The method involves enzymic dehydrogenation of d-β-hydroxybutyrate into acetoacetate and quantitative conversion of acetoacetate into acetone. The recoveries of acetone, acetoacetate and d-β-hydroxybutyrate were 94–100%, 88–104% and 83–91%, respectively.

Simultaneous determination of ketone bodies in biological samples by gas chromatographic headspace analysis.

A method is described on gas chromatographic headspace analysis for the determination of the physiological ketone bodies: acetone, acetoacetate and beta-hydroxybutyrate. The procedure proposed involves quantitative conversion of both acetoacetate by decarboxylation in a strong acidic medium, and beta-hydroxybutyrate by oxidation to acetoacetate and decarboxylation to acetone, which is then quantified chromatographically. The successful application of the method is illustrated by the values of ketone bodies in plasma, kidney, lung, and liver of fed and 48-hr-fasted rats.

The Stability and Automatic Determination of Ketone Bodies in Blood Samples Taken in Field Conditions

An automated, spectro-photometric determination of blood acetoacetate and β-hydroxybutyrate was developed with a Gilford 3500 autoanalyzer. The stability of ketone bodies was studied in different conditions. An immediate precipitation with 0.6 M perchloric acid and cooling the sample effectively prevent the loss of acetoacetate from samples during transport to the laboratory (at 4°C a 6 % loss of acetoacetate was noted during 24 h). Freezing the sample makes it practically stable (less than 2 % loss of acetoacetate per week during a study lasting 2 months). At room temperature (20°C) the sample’s acetoacetate was instable and disappeared with a rate of 6 % per h. β-hydroxybutyrate was stable in precipitated samples. Because the precipitation also retains the sample’s glucose, 3 main parameters for the indication of ketosis could be analyzed automatically from the same sample with a total capacity of 40 samples in 2½ h.

A micro-autoanalytic procedure developed for the determination of ketone bodies, gluconeogenic amino acids, pyruvate, lactate, and glucose in metabolic studies

A micro-autoanalytic procedure for the enzymic, fluorometric determinations of acetoacetate, β-hydroxybutyrate, pyruvate, lactate, alanine, glucose, glutamate, and glutamine has been described. As many as 60 to 80 duplicate samples can be analyzed simultaneously for either two or three compounds in 8 hr. The sensitivity and range of detection permit analyses of all eight compounds using only 50 μl of blood. The reproducibility, efficiency, and versatility of this autoanalytic system will warrant its wide application in metabolic studies.

Micro method for determination of ketone bodies by head-space gas chromatography

A method has been developed for the determination of ketone bodies. The principle is to transform acetoacetate and β-hydroxybutyrate to acetone, which is analyzed by means of head-space gas chromatography. Recoveries of acetoacetate and β-hydroxybutyrate were 101 and 86.1%, with standard deviations of 5–8 and 5–9%, respectively. With this method, concentrations from 1 μ M could be measured accurately, which is particularly convenient for biological analyses. If samples of acetoacetate and β-hydroxybutyrate are to be stored, the transformation of the respective substances to acetone should be carried out on the day of sampling. The acetone samples should be alkalized to prevent decarboxylation of acetoacetate.

A fluorimetric procedure for the determination of ketone bodies in very small quantities of blood

Determination of β-hydroxybutyrate (BHBA) in blood and milk samples is an important tool in the diagnosis of ketosis in dairy cattle. Apart from semiquantitative cow-side tests, well-established laboratory methods exist for measurements in blood serum or plasma. These spectrophotometric methods are, however, neither convenient nor reliable when transferred to analyses of milk. Due to its nontransparent nature, milk needs extensive pretreatment if traditional analyses are to be used. This paper describes a fluorometric determination of BHBA that is useful without pretreatment in opaque matrices such as milk and in blood plasma. The method is easy to automate, saves labor expenses, and is inexpensive. The analytical accuracy and precision are reliable for intensive as well as large-scale analysis; for example, in-line sampling from automatic milking systems. Analysis of 2500 random milk samples showed a BHBA content ranging from 10 to 631 μM (mean 49 μM). Furthermore, selected samples (n = 295) from diagnosed ketotic animals taken on d −35 to +35 from peak level ranged from 10 to 684 μM (median 79 μM, mean 141 μM). Using the same 1240 blood plasma samples, the fluorometric method was closely correlated with a traditional spectrophotometric method (r = 0.987). Hemolysis of samples does not appear to affect the fluorometric determination of BHBA.

Modification of the Spectrophotometric Determination of Ketone Bodies in Blood Enabling the Total Recovery of β-Hydroxybutyric Acid

Abstract A method is described for the determination of ketone bodies in blood, based on (1) their conversion to acetone by boiling in Greenberg and Lester's microrefluxing units, (2) the reaction of acetone with alkaline salicylaldehyde by a modification of Thin and Robertson's microdiffusion technic, and (3) the colorimetric determination of salicylacetone in water. A study of the effects of varying concentrations of dichromate and sulfuric acid on the recoveries of acetoacetic and β-hydroxybutyric acids is presented and the optimum reagent concentrations are established. Recoveries of 100 per cent are obtained for both acetoacetic and β-hydroxybutyric acids. The method is particularly sensitive to small changes of ketone bodies. In the range 0-8 mg. acetone per 100 ml. blood, differences of 0.06 mg./100 ml. are detected with ease.

The determination of ketone bodies in blood

Download options Please wait... Article information DOI https://doi.org/10.1039/AN9608500265 Article type Paper Download Citation Analyst, 1960,85, 265-271 BibTex EndNote MEDLINE ProCite ReferenceManager RefWorks RIS Permissions Request permissions The determination of ketone bodies in blood R. L. Reid, Analyst, 1960, 85, 265 DOI: 10.1039/AN9608500265 To request permission to reproduce material from this article, please go to the Copyright Clearance Center request page. If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given. If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page. Read more about how to correctly acknowledge RSC content. Social activity Tweet Share Search articles by author R. L. Reid

A specific, sensitive, and rapid micromethod for determination of ketone bodies in blood

A specific, sensitive, and rapid micromethod for determination of ketone bodies in blood

Determination of Ketone Bodies in Blood and Urine by Means of Vanillin in Alkaline Medium

Abstract Vanillin in an alkaline reaction mixture is a very sensitive reagent for acetone and diacetic acid. A qualitative and a quantitative method have been developed for the acetone bodies in blood and in urine by means of a vanillin-alkali reagent. Acetone and diacetic acid are distilled off and caught in a sodium bisulfite solution, which is allowed to react with vanillin in alkaline medium to form a color complex, which is measured spectrophotometrically. Vanillin has many advantages over the salicylaldehyde method of Behre. These have been enumerated in the text.

The determination of ketone bodies.

Research Article| September 01 1957 The determination of ketone bodies P. A. Mayes; P. A. Mayes 1Department of Biochemistry, King's College, Strand, London, W.C. 2 Search for other works by this author on: This Site PubMed Google Scholar W. Robson W. Robson 1Department of Biochemistry, King's College, Strand, London, W.C. 2 Search for other works by this author on: This Site PubMed Google Scholar Biochem J (1957) 67 (1): 11–15. https://doi.org/10.1042/bj0670011 Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Cite Icon Cite Get Permissions Citation P. A. Mayes, W. Robson; The determination of ketone bodies. Biochem J 1 September 1957; 67 (1): 11–15. doi: https://doi.org/10.1042/bj0670011 Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest filter your search All ContentAll JournalsBiochemical Journal Search Advanced Search This content is only available as a PDF. © 1957 CAMBRIDGE UNIVERSITY PRESS1957 Article PDF first page preview Close Modal You do not currently have access to this content.