Determination of serum arabinitol levels by mass spectrometry in patients with postoperative candidiasis

A combined gas chromatography-mass spectrometry (GC-MS) technique was used for serial determination of serum arabinitol levels in patients with postoperative candidiasis. Forty subjects were investigated, 18 patients with candidiasis, 7 patients with superficialCandida colonization and 15 postoperative control patients. The arabinitol levels were highly elevated, slightly elevated and normal in 13, 1 and 4 patients respectively with candidiasis; in 2, 2 and 3 colonized patients and in 1, 1 and 13 control patients (p<0.001,x ², between all the groups). The sensitivity of a single arabinitol determination for detection of postoperative candidiasis was 27.6 % and the specificity 89.2 %. Use of multiple samples improved sensitivity up to 72.2 % per patient (123 samples) with a specificity of 86.4 %. Highly elevated arabinitol concentrations were detected in only one patient before the onset of therapy. Determination of arabinitol levels by GC-MS is a specific test for diagnosing candidiasis, but multiple samples are required for adequate sensitivity, and the initiation of therapy must still be on an empirical basis.     

Comparison of serum mannan, arabinitol, and mannose in experimental disseminated candidiasis

Concentrations of arabinitol, mannose, and mannan in serum have independently been reported to be elevated in patients with invasive candidiasis. These three marker substances were compared in a rabbit model. Twelve rabbits, immunosuppressed with cortisone, were infected intravenously with Candida albicans 3181A. Six uninfected control animals also received cortisone, and four rabbits were neither infected nor immunosuppressed. Blood samples, drawn from 2 days before to 14 days after infection, were assayed for serum mannan by sandwich enzyme immunoassay, antibodies to mannan by indirect enzyme immunoassay, arabinitol and mannose by gas-liquid chromatography, and serum creatinine. Serum mannan, negative before infection, peaked in all infected animals 4 days after infection (mean, 18 ng/ml) and decreased thereafter. Significant increases (2 standard deviations greater than mean in normals) in arabinitol, the arabinitol/creatinine ratio, and mannose were found in 12, 8, and 12 of the infected rabbits, respectively, but also in all 6 uninfected animals receiving cortisone. Only serum mannan was specific in this immunosuppressed rabbit model.     

Gastrointestinal candidiasis in rats treated with antibiotics, cortisone, and azathioprine.

Conventional albino rats treated with peroral chloramphenicol, gentamicin, and/or parenteral cortisone were challenged with Candida albicans Antibiotics and cortisone were equally effective in predisposing the animals to colonization by the fungus. All animals treated with both antibiotics and cortisone developed defined, focal, superficial invasion of the cornified squamous epithelium of the stomach next to its junction with the glandular mucosa, as well as focal superficial invasion of the esophagus. Equivalent yeast cell and mycelial inocula of C. albicans were equally effective in producing colonization and invasion of the gut. Dissemination of the fungus from the gut was not found even after the addition of azathioprine to the treatment regimen; however, such addition did predispose to more extensive and severe lesions of the esophagus and stomach. Approximately 25% of infected cortisone- and antibiotic-treated rats developed agglutinins against C. albicans by 22 to 23 days after challenge, whereas 15% developed precipitins. The antibiotic-cortisone-treated rat may be a useful and consistent experimental model in the study of gastrointestinal candidiasis.     

Comparison of antibody, antigen, and metabolite assays in rat models of systemic and gastrointestinal candidiasis.

We compared serial measurements of antibodies to mannan and to a cytoplasmic antigen (enzyme-linked immunosorbent assays), detection of mannan and an unidentified candidal antigen (latex agglutination), and assays of mannose and arabinitol (gas chromatographic assay of per-O-acetylated aldonitrile derivatives). In a high-inoculum intravascular-infection model, antimannan assays were consistently positive beginning on day 2 postinoculation, anti-cytoplasmic antigen assays followed the same time course but were less sensitive, mannan was detected in all samples beginning on day 2 postinoculation, and serum mannose concentrations peaked on day 3 postinoculation and were less sensitive than mannan detection. Other assays were not useful. In a lower-inoculum intravascular-infection model, the antibody assays became positive after a similar interval and remained positive for 28 days, with antimannan again being the more sensitive. Mannan and mannose tests were positive in week 1 postinoculation only, with mannan detection being the more sensitive. In a gastrointestinal-colonization model, antimannan assays become positive after 2 weeks of colonization, whereas anti-cytoplasmic antigen and mannan tests remained negative. In a model of gastrointestinal colonization followed by invasive infection produced by induction of neutropenia, only mannan detection was diagnostically useful. These data, comparing this panel of modern serodiagnostic techniques in controlled models of clinically relevant syndromes of candidiasis, enhance understanding of previous efforts in serodiagnosis of candidiasis and provide a foundation for further prospective studies in patients.     

Levels of the Candida metabolite D-arabinitol in sera of steroid-treated and untreated patients with sarcoidosis.

We measured the Candida metabolite D-arabinitol and its enantiomer L-arabinitol in 42 serum samples from 33 patients with sarcoidosis and compared the results with those from 27 healthy adults and 4 patients with candidiasis. The D- and L-arabinitol concentrations and the D- and L-arabinitol/creatinine ratios did not differ significantly in the sarcoidosis patients and the controls; the D-arabinitol concentrations and the D-arabinitol/creatinine ratios were much higher in the patients with candidiasis. Among the patients with sarcoidosis, the D- and L-arabinitol levels in the steroid recipients did not differ significantly from those in patients not receiving steroids. Higher D-arabinitol/creatinine ratios were associated with roentgenographic evidence of pulmonary fibrosis and low forced vital capacities, but not with disease activity as determined by the proportion of lymphocytes to total nucleated cells in bronchoalveolar lavage fluid or the CD4/CD8 ratio in bronchoalveolar lymphocytes. We conclude that neither sarcoidosis nor corticosteroid treatment is associated with high levels of D-arabinitol in serum.     

An automated enzymatic method for measurement of D-arabinitol, a metabolite of pathogenic Candida species.

An automated enzymatic method was developed for the measurement of D-arabinitol in human serum. The assay is based on a novel, highly specific D-arabinitol dehydrogenase from Candida tropicalis. This enzyme catalyzes the oxidation of D-arabinitol to D-ribulose and the concomitant reduction of NAD+ to NADH. The NADH produced is used in a second reaction to reduce p-iodonitrotetrazolium violet (INT) to INT-formazan, which is measured spectrophotometrically. The entire reaction sequence can be performed automatically on a COBAS MIRA-S clinical chemistry analyzer (Roche Diagnostic Systems, Inc., Montclair, N.J.). Replicate analyses of human sera supplemented with D-arabinitol over a concentration range of 0 to 40 microM demonstrated that the pentitol could be measured with an accuracy of +/- 7% and a precision (standard deviation) of +/- 0.4 microM. Serum D-arabinitol measurements correlated with those determined by gas chromatography (r = 0.94). The enzymatic method is unaffected by L-arabinitol, D-mannitol, or other polyols commonly found in human serum. Any of 17 therapeutic drugs potentially present in serum did not significantly influence assay performance. Data illustrating the application of the assay in patients for possible diagnosis of invasive candidiasis and the monitoring of therapeutic intervention are presented. The automated assay described here was developed to facilitate the investigation of D-arabinitol as a serum marker for invasive Candida infections.     

Simultaneous determination of arabinitol and mannose by gas-liquid chromatography in experimental candidiasis.

A method is described for the simultaneous quantitation of D-arabinitol and D-mannose in serum by gas-liquid chromatography as an aid for the diagnosis of disseminated candidiasis. Both variables were observed as per-O-acetylated aldononitrile derivatives in each chromatographic run of sera from immunosuppressed rabbits experimentally infected with Candida albicans 3181A.     

Quantification of arabinitol in serum by selected ion monitoring as a diagnostic technique in invasive candidiasis.

D-Arabinitol was identified by mass spectrometry as a metabolite of Candida albicans and Candida tropicalis. For quantification, serum was deproteinized with acetone, the supernatant was evaporated to dryness, the silyl derivative was formed, and a portion was injected into a combined gas chromatograph-mass spectrometer system. Erythritol or 2-deoxy-galactitol was the internal standard. The protonated molecular ions, obtained in chemical ionization with isobutane, were monitored. For 39 normal subjects the mean endogenous arabinitol concentration was 0.52 microgram/ml (standard deviation, +/- 0.34). An increment of 0.2 microgram of arabinitol per ml could be quantified. Of 11 cases with diagnosed invasive candidiasis, 9 had arabinitol levels > 1.2 microgram/ml (8 microM) in the range of 1.2 to 25.0 micrograms/ml; the remaining 2 cases had levels in the normal range. Six cases of diagnosed colonized candidiasis showed normal arabinitol levels.     

Diagnosis of invasive candidiasis in neutropenic children with cancer by determination of D-arabinitol/L-arabinitol ratios in urine.

Determination of D-arabinitol/L-arabinitol ratios (referred to as D/L-arabinitol ratios) in urine as a tool for the diagnosis of invasive candidiasis was investigated in a prospective study comprising 100 children with cancer. The analyses were made by gas chromatography. Positive D/L-arabinitol ratios were found for 10 of 10 children with confirmed invasive candidiasis, 12 of 23 patients undergoing empiric antifungal chemotherapy, and 4 of 67 children not receiving antifungal treatment. D/L-Arabinitol ratios were positive 3 to 31 days (median, 12 days) before the first culture-positive blood sample was drawn or empiric therapy was initiated. The regular monitoring of D/L-arabinitol ratios in urine holds great promise as a sensitive method for diagnosing invasive candidiasis in immunocompromised children with cancer. Moreover, it may be possible to use an early rise in D/L-arabinitol ratios as a basis for the institution of antifungal chemotherapy and as a means of avoiding unnecessary treatment with potentially toxic antifungal agents.     

Gas chromatographic determination of D-arabinitol/L-arabinitol ratios in urine: a potential method for diagnosis of disseminated candidiasis.

A gas chromatographic procedure was developed to determine the relative amounts of D- and L-arabinitol in urine. Samples were filtered, diluted, purified through extractions, evaporated, and treated with trifluoroacetic anhydride; the arabinitol derivatives thus obtained were separated on a chiral stationary phase and registered by using an electron-capture detector. Urine samples from a patient with disseminated candidiasis had higher D-arabinitol/L-arabinitol ratios (referred to as D/L-arabinitol ratios)--up to 19.0--than samples from 96 study individuals with no signs of deep Candida infections (range, 1.1 to 4.5). D/L-Arabinitol ratios in urine samples from hospitalized patients without Candida infections were slightly higher than those in samples from healthy individuals; ratios in urine from children were slightly higher than those in adult urine samples. The D/L-arabinitol ratios in several urine samples culture positive for Candida albicans, but from patients without symptoms of disseminated candidiasis, did not differ from those in the urine of healthy individuals. The described gas chromatographic method is straightforward and can be implemented clinically to determine urine D/L-arabinitol ratios as a means of diagnosing disseminated candidiasis.     

Enantioselective measurement of fungal D-arabinitol in the sera of normal adults and patients with candidiasis.

A new method was used to measure D-arabinitol enantioselectively in the sera of 27 healthy adults and four patients with candidiasis. Arabinitol was measured by gas chromatography in serum that was treated with and without the Klebsiella pneumoniae enzyme D-arabinitol dehydrogenase, lactic dehydrogenase, NAD, and sodium pyruvate. Since enzyme treatment removed 98% of 0 to 20 micrograms of D-arabinitol per ml and none of 0 to 20 micrograms of L-arabinitol per ml from spiked sera, D-arabinitol could be determined from the difference in the treated and untreated samples. The concentrations of D- and L-arabinitol in serum from normal subjects were 0.22 +/- 0.052 and 0.16 +/- 0.055 micrograms/ml, respectively, and their D-arabinitol/creatinine and L-arabinitol/creatinine ratios were 0.024 +/- 0.0089 and 0.017 +/- 0.0053 (all means +/- standard deviations). The infected patients all had markedly elevated serum D-arabinitol levels, but their L-arabinitol levels were either normal or proportionately much lower. The excess arabinitol in the sera of individuals with candidiasis is D-arabinitol, and use of enantioselective analytical methods should result in improved ability to diagnose and estimate the severity of candidiasis.     

Effects of different forms of dietary hydrogenated fats on serum lipoprotein cholesterol levels.

BACKGROUND: Metabolic studies suggest that fatty acids containing at least one double bond in the trans configuration, which are found in hydrogenated fat, have a detrimental effect on serum lipoprotein cholesterol levels as compared with unsaturated fatty acids containing double bonds only in the cis configuration. We compared the effects of diets with a broad range of trans fatty acids on serum lipoprotein cholesterol levels. METHODS: Eighteen women and 18 men consumed each of six diets in random order for 35-day periods. The foods were identical in each diet, and each diet provided 30 percent of calories as fat, with two thirds of the fat contributed as soybean oil (<0.5 g of trans fatty acid per 100 g of fat), semiliquid margarine (<0.5 g per 100 g), soft margarine (7.4 g per 100 g), shortening (9.9 g per 100 g), or stick margarine (20.1 g per 100 g). The effects of those diets on serum lipoprotein cholesterol, triglyceride, and apolipoprotein levels were compared with those of a diet enriched with butter, which has a high content of saturated fat. RESULTS: The mean (+/-SD) serum low-density lipoprotein (LDL) cholesterol level was 177+/-32 mg per deciliter (4.58+/-0.85 mmol per liter) and the mean high-density lipoprotein (HDL) cholesterol level was 45+/-10 mg per deciliter (1.2+/-0.26 mmol per liter) after subjects consumed the butter-enriched diet. The LDL cholesterol level was reduced on average by 12 percent, 11 percent, 9 percent, 7 percent, and 5 percent, respectively, after subjects consumed the diets enriched with soybean oil, semiliquid margarine, soft margarine, shortening, and stick margarine; the HDL cholesterol level was reduced by 3 percent, 4 percent, 4 percent, 4 percent, and 6 percent, respectively. Ratios of total cholesterol to HDL cholesterol were lowest after the consumption of the soybean-oil diet and semiliquid-margarine diet and highest after the stick-margarine diet. CONCLUSIONS: Our findings indicate that the consumption of products that are low in trans fatty acids and saturated fat has beneficial effects on serum lipoprotein cholesterol levels.     

糖皮质激素对内分泌器官的广泛影响

目的：探讨大剂量糖皮质激素（Glucocorticoid,GC)对多种内分泌器官的影响,为临床减轻GC的副作用提供一些依据。方法：以大量醋酸可的松对大鼠进行注射,取其垂体、肾上腺、甲状腺、睾丸进行电镜样品制备及观察。结果：大量醋酸可的松可引起大鼠垂体、肾上腺、甲状腺、睾丸细胞内质网扩张,线粒体肿胀、空泡化,核固缩,细胞萎缩,脂滴分布的改变等超微结构的损伤。结论：大剂量GC对内分泌系统具有广泛的影响。