Separation of the enantiomers of ibuprofen and its major phase I metabolites in urine using capillary electrophoresis

This report describes a simple, rapid, automated microassay for measuring in vitro changes of oxidative burst of phagocytes following challenge with metals for orthopedic devices. The production of reactive oxygen species (ROS) by polymorphonuclear leukocytes (PMNs) was measured using 2',7'-dichlorofluorescin-diacetate (DCFH-DA) as fluorescent probe. DCFH-DA enters the cells and is oxidized by ROS to fluorescent DCF. The DCF generated was directly proportional to ROS produced intracellularly: The fluorescence intensity was read and converted to an index of ROS production by cells. In our experimental system, granulocytes (PMNs) were isolated from normal human blood and seeded in microplates. To verify if metals could influence ROS production, chromium, cobalt, nickel, molybdenum, titanium, aluminum, and vanadium prepared as aqueous extracts in phosphate-buffered saline were tested onto PMNs using phorbolmyristate acetate (PMA) as positive control. Molybdenum, aluminum, and vanadium increased ROS generation by PMNs, while signals not different from unstimulated PMNs were recorded for chromium, cobalt, nickel, and titanium. The DCFH-DA microplate-based assay provides an in vitro tool for the detection of oxygen-reactive species generated by PMNs as a response to metals.     

Greater concentration of dietary sucrose decreases dentin formation and increases the area of dentinal caries in growing rats

The effect of increasing dietary sucrose concentration on dentin formation and dentinal caries progression was studied. Weanling Wistar rats received 15, 30 or 43 g/100 g sucrose in a diet; for reference, another group was fed a nonpurified diet. At the onset, tetracycline was injected to mark the dentin formed during the experiment. After 6 wk, lower molars were sectioned sagittally; the areas and thicknesses of the dentin formation during the experiment and dentinal caries lesions were quantified separately in the first and second molars. Feeding the 43% sucrose diet resulted in a significantly lower dentin formation than in other diet groups (P < 0.05). The differences obtained from the area measurements were supported by thickness measurements. In the first molar, the 43% sucrose diet resulted in a significantly greater area of dentinal caries than in the other sucrose groups. The number and severity of caries lesions clearly increased as the concentration of sucrose in the diet increased (r = 0.5, P < 0.05 and r = 0.6, P < 0.05, respectively). This study suggests that the increase in the concentration of sucrose in the diet reduces dentin formation and increases the area of dentinal caries as well as the number and severity of caries lesions; the critical sucrose concentration appears to be between 30 and 43 g/100 g.     

On a class of nonstationary random processes

We consider the class of random processes having linear shift operators. This class is an extension of the class of wide-sense stationary processes (which have unitary shift operators). Conditions for a process to have linear shifts are formulated in terms of the covariance function of the process. Sufficient conditions for a purely nondeterministic process to have linear shifts are given. Theorems concerning superposition, products, and linear transformations of such processes are proved, and applications are indicated. A comparison with the class of locally stationary processes is made. The essential concepts involved are also extended to generalized random processes.     

Nonaqueous capillary electrophoresis--its applicability in the analysis of food, pharmaceuticals and biological fluids

The use of nonaqueous electrophoresis media for the application of capillary electrophoresis in the analysis of food, pharmaceuticals and biological fluids is reviewed. Some of the applications are discussed in detail and the benefits of using nonaqueous media in these cases are outlined. Three new applications within pharmaceutical analyses are presented. In these methods either a simple sample pretreatment by dilution with methanol (determination of chlorhexidine in a cream) or selective on-line capillary electrophoresis mass spectrometry (methods for identification of seizure drugs or opium alkaloids) are used. The choice of organic solvents and electrolytes for nonaqueous capillary electrophoresis are discussed. Furthermore, validation data obtained using capillary electrophoresis based on the nonaqueous principle are listed and discussed.     

No Association between HMOX1 and Risk of Colorectal Cancer and No Interaction with Diet and Lifestyle Factors in a Prospective Danish Case-Cohort Study

Red meat is a risk factor for colorectal cancer (CRC). We wanted to evaluate whether a functional polymorphism in the HMOX1 gene encoding heme oxygenase modifies risk of CRC or interacts with diet or lifestyle factors because this would identify heme or heme iron as a risk factor of CRC. The HMOX1 A-413T (rs2071746) was assessed in relation to risk of colorectal cancer (CRC) and interactions with diet (red meat, fish, fiber, cereals, fruit and vegetables) and lifestyle (use of non-steroidal anti-inflammatory drug and smoking status) were assessed in a case-cohort study of 928 CRC cases and a comparison group of 1726 randomly selected participants from a prospective study of 57,053 persons. No association between HMOX1 A-413T and CRC risk was found (TT vs. AA + TA; IRR = 1.15, 95% CI: 0.98–1.36, p = 0.10 for the adjusted estimate). No interactions were found between diet or lifestyle and HMOX1 A-413T. HMOX1 A-413T was not associated with CRC risk and no interactions with diet or lifestyle were identified in this large, prospective cohort with high meat intake. The results reproduced the previous findings from the same cohort and did not support a link between heme or heme iron and colorectal cancer. These results should be sought and replicated in other well-characterized cohorts with high meat intake.     

Predictors of polychlorinated biphenyl concentrations in adipose tissue in a general Danish population

Polychlorinated biphenyls (PCBs) are ubiquitously present in the environment and suspected of carcinogenic, neurological, and immunological effects. Our purpose was to identify predictors of adipose tissue levels of mono-, di-, and tri-ortho-substituted PCBs experienced by a general population and to establish whether predictors vary according to substitution group. In this study of 245 randomly selected persons from a prospective Danish cohort of 57,053 persons, we examined geographical area, age, lactation, BMI, and intake of eight major dietary groups as potential determinants of adipose concentrations of mono-, di-, and tri-ortho-substituted PCBs by linear regression analyses. Lactation, BMI, and intake of fruit, vegetables, and dairy products showed negative associations with PCB concentrations in adipose tissue in all models, and living in Copenhagen city, age, and consumption of fish (particularly fatty fish) were positively associated. The associations between several of the predictors and mono-ortho-substituted PCBs tended to differ from the associations found for di- and tri-ortho-substituted PCBs. In conclusion, geography, age, lactation, BMI, and consumption of fatty fish consistently predicted the concentration of PCBs in adipose tissue. Our results indicate that predictors of PCBs varied according to substitution group, suggesting that ortho-substituted groups should be analyzed separately.     

Separation of basic drug substances of very similar structure using micellar electrokinetic chromatography

Small molecules having very similar molecular structure--some even with the same mass over charge--are not trivial to separate by capillary electrophoresis in free solution. However, addition of surfactants to the electrophoretic buffer and thus using the principle of micellar electrokinetic chromatography may provide separation with high resolution. The use of zwitterionic and nonionic surfactants is demonstrated and an example of how a developed system may be used for drug purity testing is given.     

A novel organ culture method to study the function of human odontoblasts in vitro: gelatinase expression by odontoblasts is differentially regulated by TGF-beta1

Odontoblasts cannot be cultured by traditional cell culture methods, thus restricting in vitro studies. Here we present an organ culture method for human odonto-blasts that utilizes the pulp chamber as a culture crucible. Crowns of human third molars were dissected, pulp was gently removed, and the odontoblasts attached to and in the walls of the pulp chambers were cultured in serum-free OPTI-MEM medium, or DMEM/Ham's F12 medium containing 10% serum. Pulp tissues were cultured separately. Cell content and morphology were analyzed by SEM, and the removed pulps were examined by light microscopy. Proteins secreted into the medium with or without TGF-beta1 supplementation were metabolically labeled with [35S]methionine, and the total protein content was assessed by TCA precipitation and SDS-PAGE/fluorography. To assess the role of gelatinolytic enzymes on dentin matrix remodeling, we used enzymography to analyze the effect of TGF-beta1 on gelatinase A and B expression. SEM revealed odontoblasts in pulp chambers after 5 days of culture, with only few or no fibroblasts, and no alterations in the odontoblast cell morphology or differences between the cells cultured in serum-free and serum-containing media. Rarely were any odontoblasts present in pulp tissue. Radiolabeling revealed protein synthesis and secretion until day 6 in both the odontoblast and pulp cultures, with no marked differences between TGF-beta1-treated and control cultures. The level of gelatinase A remained constant up to 7 days, while gelatinase B expression was always low and decreased with time in culture. However, gelatinase B levels were markedly increased upon TGF-beta1 treatment of cells and remained high to day 7. The results suggest that this method provides a novel technique for the study of human odontoblasts in vitro and that odontoblasts can be cultured even in serum-free conditions.