Determination of atenolol in pharmaceutical formulations by diffuse reflectance spectroscopy

A simple analytical method for quantification of atenolol in pharmaceutical formulations by diffuse reflectance spectroscopy is described. The method is based on the reaction, on the filter paper surface, between the drug and p-chloranil producing a colored compound. The best reaction conditions were obtained with 20 µL of atenolol solution and 20 µL of p-chloranil. All reflectance measurements were carried out at 550 nm and the linear range was from 1.13x10-2 to 7.88x10-2 mol L-1 (r = 0.9992). The limit of detection was 2.80 x 10-3 mol L-1. The proposed method was successfully applied to analysis of different commercial brands of pharmaceutical formulations and the results obtained by the proposed method were in good agreement with those obtained using the British Pharmacopoeia method.

Potentiometric determination of Minoxidil in topical use pharmaceutical samples

A potentiometric titration method for the determination of minoxidil based on its redox reaction with K2Cr2O7 is described. The best results were observed using 1.00 x 10-3 mol L-1 K2Cr2O7 and 1.00 x 10-2 mol L-1 minoxidil solutions, and the minoxidil as titrant in 2.00 mol L-1 H2SO4 medium. The method was applied to commercial samples and compared with the results from a chromatographic procedure. Recoveries from 97.4 to 98.7 % were observed depending on the sample. Comparison with the chromatographic procedure reveled agreement within 90% confidence level.

Sensitive spectrophotometric determination of lamotrigine in bulk drug and pharmaceutical formulations using bromocresol green

Two new, simple, rapid and reproducible spectrophotometric methods have been developed for the determination of lamotrigine (LMT) both in pure form and in its tablets. The first method (method A) is based on the formation of a colored ion-pair complex (1:1 drug/dye) of LMT with bromocresol green (BCG) at pH 5.02±0.01 and extraction of the complex into dichloromethane followed by the measurement of the yellow ion-pair complex at 410 nm. In the second (method B), the drug-dye ion-pair complex was dissolved in ethanolic potassium hydroxide and the resulting base form of the dye was measured at 620 nm. Beer's law was obeyed in the concentration range of 1.5-15 µg mL-1 and 0.5-5.0 µg mL-1 for method A and method B, respectively, and the corresponding molar absorptivity values are 1.6932 x 10(4) and 3.748 x 10(4) L mol-1cm-1. The Sandell sensitivity values are 0.0151 and 0.0068 µg cm-2 for method A and method B, respectively. The stoichiometry of the ion-pair complex formed between the dug and dye (1:1) was determined by Job's continuous variations method and the stability constant of the complex was also calculated. The proposed methods were applied successfully for the determination of drug in commercial tablets.

Potentiometric and conductimetric studies of chemical equilibria for pyridoxine hydrochloride in aqueous solutions: simple experimental determination of pKa values and analytical applications to pharmaceutical analysis

The combination of two low-cost classical procedures based on titrimetric techniques is presented for the determination of pyridoxine hydrochloride in pharmaceuticals samples. Initially some experiments were carried out aiming to determine both pKa1 and pKa2 values, being those compared to values of literature and theoretical procedures. Commercial samples containing pyridoxine hydrochloride were electrochemically analysed by exploiting their acid-base and precipitation reactions. Potentiometric titrations accomplished the reaction between the ionizable hydrogens present in pyridoxine hydrochloride, being NaOH used as titrant; while the conductimetric method was based on the chemical precipitation between the chloride of pyridoxine hydrochloride molecule and Ag+ ions from de silver nitrate, changing the conductivity of the solution. Both methods were applied to the same commercial samples leading to concordant results when compared by statistical tests (95 and 98% confidence levels). Recoveries ranging from 99.0 to 108.1% were observed, showing no significant interference on the results.

A simple spectrophotometric method for the determination of tetracycline and doxycycline in pharmaceutical formulations using chloramine-T

A simple, rapid, accurate and inexpensive spectrophotometric method for the determination of tetracycline and doxycycline has been developed. The method is based on the reaction between these drugs and chloramine-T in alkaline medium producing red color products with absorbance maximum at the Λ = 535 and 525 nm for the tetracycline and doxycycline, respectively. The best conditions for the reactions have been found using multivariate method. Beer´s law is obeyed in a concentration ranges 1.03 x 10-5 to 3.61 x 10-4 mol L-1 and 1.75 x 10-5 to 3.48 x 10-4 mol L-1 for the tetracycline and doxycycline, respectively. The quantification limits were 5.63 x 10-6 mol L-1 and 7.12 x 10-7 mol L-1 for the tetracycline and doxycycline, respectively. The proposed method was successfully applied to the determination of these drugs in pharmaceutical formulations and the results obtained were in good agreement with those obtained by the comparative method at the 95% confidence level.

Recuperation and improvement of chromosome preparations stained with silver nitrate

Both periodic acid and photographic reagents have been used to remove the silver nitrate residues from cytological preparations. We used potassium ferricyanide to remove AgNO3 salts in cases of excessive chromosome impregnation. This method produced partial decolorization, with contrast enhancement. Counterstaining with Giemsa also promoted a better contrast between chromosome arms and NORs, which were preserved and retained a dark color. Application of this procedure to chromosome preparations treated sequentially for CBG banding/AgNO3 staining promoted complete decolorization of C+ regions, leading to "reverse" C-banding, generally with high contrast.

Assessment of the degree of contamination of rat germ cell preparations using specific cDNA probes

Recent reports showing a decrease in sperm count in men have brought new concerns about male infertility. Animal models have been widely used to provide some relevant information about the human male gamete, and extrapolations are made to men and to the clinical context. The present study assesses one of the methods used for separation of germ cells of the adult rat testis, namely centrifugal elutriation followed by density gradients (Percoll®). This method was chosen since it presents the best results for cell purity in separating germ cells from the rat testis. A comparison between continuous and discontinuous Percoll® gradients was performed in order to identify the best type of gradient to separate the cells. Maximal cell purity was obtained for spermatocytes (81 ± 8.2%, mean ± SEM) and spermatids (84 ± 2.6%) using centrifugal elutriation followed by continuous Percoll® gradients. A significant difference in purity was observed between elongating spermatids harvested from continuous Percoll® gradients and from discontinuous gradients. Molecular analysis was used to assess cell contamination by employing specific probes, namely transition protein 2 (TP2), mitochondrial cytochrome C oxidase II (COX II), and sulfated glycoprotein 1 (SGP1). Molecular analysis of the samples demonstrated that morphological criteria are efficient in characterizing the main composition of the cell suspension, but are not reliable for identifying minimal contamination from other cells. Reliable cell purity data should be established using molecular analysis

Developed pressure data may provide misinformation when used alone to evaluate systolic function in isovolumetric left ventricle preparations

We report data showing that developed pressure (DPmax) may lead to opposite conclusion with respect to maximal developed circumferential wall stress (smax) when used to assess contractile function in left ventricle isovolumic preparations. Isovolumetric left ventricle preparations of rats with cardiac hypertrophy (H; N = 10) induced by isoproterenol administration showed higher DPmax (174 ± 14 mmHg) than control (C; N = 8) animals (155 ± 12 mmHg) or rats with regression (R; N = 8) of hypertrophy (144 ± 11 mmHg). In contrast, the estimated smax for C (145 ± 26 kdynes/cm2) and R (133 ± 17 kdynes/cm2) was higher than for H (110 ± 13 kdynes/cm2). According to Laplace's law, the opposite results of DPmax and smax may depend on the increased mass/volume left ventricle ratio of the hypertrophied hearts, which favored pressure generation. These results clearly show that DPmax should be used with caution to analyze systolic function.

Efficacy, safety, quality control, marketing and regulatory guidelines for herbal medicines (phytotherapeutic agents)

This review highlights the current advances in knowledge about the safety, efficacy, quality control, marketing and regulatory aspects of botanical medicines. Phytotherapeutic agents are standardized herbal preparations consisting of complex mixtures of one or more plants which contain as active ingredients plant parts or plant material in the crude or processed state. A marked growth in the worldwide phytotherapeutic market has occurred over the last 15 years. For the European and USA markets alone, this will reach about $7 billion and $5 billion per annum, respectively, in 1999, and has thus attracted the interest of most large pharmaceutical companies. Insufficient data exist for most plants to guarantee their quality, efficacy and safety. The idea that herbal drugs are safe and free from side effects is false. Plants contain hundreds of constituents and some of them are very toxic, such as the most cytotoxic anti-cancer plant-derived drugs, digitalis and the pyrrolizidine alkaloids, etc. However, the adverse effects of phytotherapeutic agents are less frequent compared with synthetic drugs, but well-controlled clinical trials have now confirmed that such effects really exist. Several regulatory models for herbal medicines are currently available including prescription drugs, over-the-counter substances, traditional medicines and dietary supplements. Harmonization and improvement in the processes of regulation is needed, and the general tendency is to perpetuate the German Commission E experience, which combines scientific studies and traditional knowledge (monographs). Finally, the trend in the domestication, production and biotechnological studies and genetic improvement of medicinal plants, instead of the use of plants harvested in the wild, will offer great advantages, since it will be possible to obtain uniform and high quality raw materials which are fundamental to the efficacy and safety of herbal drugs.

Antagonism by hemoglobin of effects induced by L-arginine in neuromuscular preparations from rats

Nitric oxide (NO)-synthase is present in diaphragm, phrenic nerve and vascular smooth muscle. It has been shown that the NO precursor L-arginine (L-Arg) at the presynaptic level increases the amplitude of muscular contraction (AMC) and induces tetanic fade when the muscle is indirectly stimulated at low and high frequencies, respectively. However, the precursor in muscle reduces AMC and maximal tetanic fade when the preparations are stimulated directly. In the present study the importance of NO synthesized in different tissues for the L-Arg-induced neuromuscular effects was investigated. Hemoglobin (50 nM) did not produce any neuromuscular effect, but antagonized the increase in AMC and tetanic fade induced by L-Arg (9.4 mM) in rat phrenic nerve-diaphragm preparations. D-Arg (9.4 mM) did not produce any effect when preparations were stimulated indirectly at low or high frequency. Hemoglobin did not inhibit the decrease of AMC or the reduction in maximal tetanic tension induced by L-Arg in preparations previously paralyzed with d-tubocurarine and directly stimulated. Since only the presynaptic effects induced by L-Arg were antagonized by hemoglobin, the present results suggest that NO synthesized in muscle acts on nerve and skeletal muscle. Nevertheless, NO produced in nerve and vascular smooth muscle does not seem to act on skeletal muscle.

DNA extraction and quantification from touch and scrape preparations obtained from autopsy liver cells

The objective of the present study was to develop a simplified low cost method for the collection and fixation of pediatric autopsy cells and to determine the quantitative and qualitative adequacy of extracted DNA. Touch and scrape preparations of pediatric liver cells were obtained from 15 cadavers at autopsy and fixed in 95% ethanol or 3:1 methanol:acetic acid. Material prepared by each fixation procedure was submitted to DNA extraction with the Wizard® genomic DNA purification kit for DNA quantification and five of the preparations were amplified by multiplex PCR (azoospermia factor genes). The amount of DNA extracted varied from 20 to 8,640 µg, with significant differences between fixation methods. Scrape preparation fixed in 95% ethanol provided larger amount of extracted DNA. However, the mean for all groups was higher than the quantity needed for PCR (50 ng) or Southern blot (500 ng). There were no qualitative differences among the different material and fixatives. The same results were also obtained for glass slides stored at room temperature for 6, 12, 18 and 24 months. We conclude that touch and scrape preparations fixed in 95% ethanol are a good source of DNA and present fewer limitations than cell culture, tissue paraffin embedding or freezing that require sterile material, culture medium, laboratory equipment and trained technicians. In addition, they are more practical and less labor intensive and can be obtained and stored for a long time at low cost.

Lipid peroxidation, detoxification capacity, and genome damage in mice after transplacental exposure to pharmaceutical drugs

Data on genome damage, lipid peroxidation, and levels of glutathione peroxidase (GPX) in newborns after transplacental exposure to xenobiotics are rare and insufficient for risk assessment. The aim of the current study was to analyze, in an animal model, transplacental genotoxicity, lipid peroxidation, and detoxification disturbances caused by the following drugs commonly prescribed to pregnant women: paracetamol, fluconazole, 5-nitrofurantoin, and sodium valproate. Genome damage in dams and their newborn pups transplacentally exposed to these drugs was investigated using the in vivo micronucleus (MN) assay. The drugs were administered to dams intraperitoneally in three consecutive daily doses between days 12 and 14 of pregnancy. The results were correlated, with detoxification capacity of the newborn pups measured by the levels of GPX in blood and lipid peroxidation in liver measured by malondialdehyde (HPLC-MDA) levels. Sodium valproate and 5-nitrofurantoin significantly increased MN frequency in pregnant dams. A significant increase in the MN frequency of newborn pups was detected for all drugs tested. This paper also provides reference levels of MDA in newborn pups, according to which all drugs tested significantly lowered MDA levels of newborn pups, while blood GPX activity dropped significantly only after exposure to paracetamol. The GPX reduction reflected systemic oxidative stress, which is known to occur with paracetamol treatment. The reduction of MDA in the liver is suggested to be an unspecific metabolic reaction to the drugs that express cytotoxic, in particular hepatotoxic, effects associated with oxidative stress and lipid peroxidation.

Physicochemical study of CaCO3 from egg shells

Calcium carbonate, a pharmaceutical excipient, is widely used as diluent in solid dosage forms. It is also used as a base for medicinal and dental preparations, a buffering and dissolution aid for dispersible tablets, a food additive and a calcium supplement. Egg shells are a rich source of mineral salts, mainly calcium carbonate, which corresponds to about 94% of the shell. Layer farms produce large amounts of shells, whose final disposal poses a challenge from the environmental standpoint. This work was designed to evaluate the physicochemical and thermal properties of calcium carbonate obtained from egg shells. The findings indicated that calcium carbonate from egg shells can be used as an alternative pharmaceutical excipient.

Sensory analysis of hydrolysed meat preparations

The use of hydrolysed meat in diets contributes to the improvement of protein, vitamin and mineral supply. This work aims at checking the acceptance pattern in meat hydrolysates. Four preparations have been developed with three types of hydrolysates in domestic-like conditions. Acceptance was verified by means of sensory analysis using the nine-point hedonic scale. Sensory tests have been carried out in three sessions (according to the kind of hydrolysates). In the evaluation file, information on age groups has been included. The statistical analysis has been made by ANOVA and Tukey test. The best accepted preparation have been the turkey and chicken hydrolysed balls. Hydrolysates can be used in many different kinds of preparations, but it is necessary to know both the age group it will be used to and its sensory and chemical-physical features to ensure the taste and the original appearance of the final product.

Microbial profile of a kefir sample preparations: grains in natura and lyophilized and fermented suspension

Probiotics are supplementary foods developed by microbial strains that improve animal health beyond basic nutrition. Probiotics are consumed orally, regardless of being considered as normal inhabitants of the intestines, able to survive in enzimatic and biliary secretions. Kefir is a probiotic originated from the old continent, fermented by several bacteria and yeasts, encapsulated in a polyssacharide matrix, and resembles jelly grains. Kefir is also presented as its sourish product both in sugary or milky suspensions containing vitamins, aminoacids, peptides, carbohydrates, ethanol, and volatile compounds. Kefir is known to have a diverse microbial content depending on the country and fermentative substrates, which cause distinct probiotic effects. In this sense, the purpose of this work was to isolate, identify, and quantify the microbial content of a native sugary kefir sample (fermented suspension and lyophilized natural grains). Serial dilutions were plated on Rogosa agar (AR) and De Man, Rogosa and Sharpe (MRS), for Lactobacillus; Brain Heart Infusion (BHI), for total bacteria; Sabouraud-Dextrose-Agar (SDA), for yeasts and filamentous fungi; Thioglycolate Agar (TA), for Streptococcus, Acetobacteria and Leuconostoc; and Coconut Water Agar (CWA), and CWA supplemented with yeast extract (CWAY), for various genera. Genera and species for all strains were identified through biochemical reactions and specific API systems. The microbial profile of kefir was different from other sources of grains despite the presence of similar microorganisms and others which have not been reported yet. The data obtained with the CWA and CWAE media suggest that both substrates are alternative and salutary media for culture of kefir strains.