Ultrastructural changes in the cemento-enamel junction caused by acidic beverages: An in vitro study.

The present in vitro study was aimed at evaluating the morphological changes in the cemento-enamel junction (CEJ) after exposure to acidic beverages using the scanning electron microscopy (SEM). The initial pH and titratable acidity (TA) was analyzed from follow groups: (I) Coca cola, (II) orange juice, (III) Cedevita, (IV) Red Bull, (V) Somersby cider, and (VI) white wine. The CEJ samples (n = 64), obtained from unerupted third molars, were allocated to one control (artificial saliva, n = 16) and six experimental groups (n = 8). The experimental samples were immersed in beverages (50 ml) for 15 min, three times daily, 10 days, and in artificial saliva between immersions. SEM analysis was performed in a blind manner, according to scoring scale. One-way ANOVA and Tukey's post hoc tests, as well as Kruskal-Wallis and Mann-Whitney U test used for statistical analysis. The pH values of the acidic beverages ranged from 2.65 (Coca cola) to 3.73 (orange juice), and TA ranged from 1.90 ml (Coca cola) to 5.70 ml (orange juice) of NaOH to reach pH 7.0. The SEM analysis indicated statistically significant differences between the control samples and those immersed in acidic beverages. The Groups IV, I, and II, showed the highest CEJ damage grade while those of the Group VI were the lowest. All the tested acidic beverages caused morphological changes in the CEJ with a smaller or larger exposure of dentine surface, and were not always related to the pH or TA of acidic beverages.

HPLC method development for determination of doxycycline in human seminal fluid.

The present paper reports the development and validation of an analytical method for doxycycline quantification in human seminal fluid by HPLC with UV detection. The separation of doxycycline was achieved at 40°C on a reversed-phase C18 column using isocratic elution. The mobile phase consisted of acetonitrile (A) and water buffered at pH 2.5 with a concentrated orthophosphoric acid (B) in the volume ratio of 20:80 (v/v), respectively. The detection was performed at 350nm. As an internal standard (IS), tetracycline was used. The proposed method involves the extraction of doxycycline from seminal fluid based on acidic precipitation of the proteins using perchloric acid. The method showed good intra- and inter-day precisions (RSD<7.0%), good accuracy (recovery for doxycycline>80%), and high correlation coefficient (r=0.998) for standards subjected to the entire procedure. The detection and quantification limits were 0.087µg/ml and 0.264µg/ml. The developed method was used to analyze doxycycline in the seminal fluids obtained from male subjects who were treated with doxycycline-hyclate. The mean doxycycline concentrations of 0.89±0.07µg/ml and 0.45±0.26µg/ml were detected in seminal fluid after 6h and 12h, respectively. This is the first study reporting extraction and HPLC determination of doxycycline in this complex sample and can be very useful in support of clinical and pharmacokinetic studies on this antibiotic.

RP-HPLC assay of doxycycline in human saliva and gingival crevicular fluid in patients with chronic periodontal disease.

A reversed-phased HPLC method with fluorescence detection was optimized and validated for determination of DOXY in human saliva and gingival crevicular fluid (GCF) with tetracycline as internal standard. Single step extraction with acetonitrile for both types of samples was performed. The separation was achieved at Zorbax Extend-C18 analytical column at 30°C. Mobile phase was consisted of an aqueous phase containing magnesium acetate, ammonium acetate, Na2EDTA, triethyl-ammonium acetate buffered to pH 7.5 with ammonium hydroxide solution and acetonitrile. The volume ratio of the buffered water mixture of salts and acetonitrile was 86:14. Fluorescence detector was set at λex=380 nm and λem=520 nm. Under the optimized experimental conditions, good linearity was found in the range of 5.0-250.0 ng/mL for GCF with LOD of 1.63 ng/mL and LOQ of 4.93 ng/mL and 20.0-500.0 ng/mL for saliva with LOD of 6.36 ng/mL and LOQ of 19.28 ng/mL. This method was successfully applied for determination of DOXY in saliva and GCF obtained from patients with chronic periodontal disease.

Determination of streptomycin in a pharmaceutical sample based on its degradation by hydrogen peroxide in the presence of copper(II).

The kinetics of streptomycin degradation by hydrogen peroxide at pH 7.4 was investigated. The reaction was catalyzed by traces of Cu(II) ions, and it was followed spectrophotometrically at 322 nm by applying the initial-rate method. The kinetic parameters of the reaction are reported, and a rate equation is suggested. From the dependence of the relative rate constants on the temperature, the activation energy was calculated to be 57.5 kJ/mol. Based on this reaction, a kinetic method for streptomycin determination is proposed. The calibration graph is linear in the ranges 1.94 - 15.48 microg/cm(3) and 0.15 - 1.94 microg/cm(3). The influence of foreign ions and molecules on the accuracy of the determinations was investigated. The proposed kinetic method has high selectivity and good sensitivity, and enabled to determine streptomycin in pharmaceutical samples.