Research in Pharmaceutical Sciences
Volume:1 Issue: 1, Apr 2006

The SSCP technique is based on the appearance of new “refolding” conformations during electro-phoresis due to mutation. In order to develop a simple, non-radioactive SSCP analysis method so that it can reliably detect single nucleotide changes in PCR products up to 500 bp in length, extensive optimisation trials were performed. The best separation of SSCP bands of PCR products up to 500 bp in length was obtained with 14.5 to 15.5% polyacrylamide gels that had been run in the cold room at 15 V/cm for 65 to 70 hours. After the pre-run, 5 ml of PCR product was mixed with 10 ml of denaturing-loading dye and the mixture was heated to 94 °C for 10 min. Total mixture volume of the 15 ml was loaded into the well without quenching. After electrophoresis, the gels were stained with SYBR® Gold nucleic acid stain for 30-40 minutes and photographed under UV light using a SYBR® gel stain photographic filter. This method alongside with confirmatory sequencing has been utilised to identify three novel mutations in the 5¢-regulatory region of the human CYP3A4 gene. The final results were very satisfactory and the optimised method was able to reliably reveal new mutations in amplified DNA from blood samples. In fact the non-radioactive SSCP method developed here seems to be robust enough to analyse PCR products up to 500 bp long.

Gastric ulcer is one of the most prevalent gastrointestinal (GI) disorders, which affects approximately 5-10% of people during their life. In recent years, plentiful works have been carried out on herbal medicine to clarify their potential efficacy in gastric ulcer prevention or management. Tripleuro-spermum disciforme is one of the indigenous plants that is readily available and has been traditionally used to improve GI disorders. We decided to study its anti-ulcer effects in pylorus-ligated rats. Hydroalcoholic extract of flowers (125, 500, 2000 mg/kg), vehicle and ranitidine (50 mg/kg) were administered orally (p.o.) to separated groups of Wistar rats of either sex (n=8). Other groups received extract (500 mg/kg), ranitidine and vehicle intraperitoneally (i.p.). Volume of contents, pH, ulcer number, scoring, incidence, area and finally ulcer index were assessed and compared with control groups. Volume of gastric contents as well as pH (in reverse with acidity) increased in extract groups but the difference was not significant. In treatment groups, regardless of the changes in ulcer number and scores, the differences were not significant for both parameters compared to control groups. Both the extract and reference drug (ranitidine) resulted in significant reduction in ulcer area and ulcer index and for latter the range of reduction was 21.8-39.1%. The least dose of extract (125 mg/kg) was not effective. We conclude that hydroalcoholic extract of T. disciforme was effective to protect against ulcer formation in pylorus-ligated rats and the action is not likely to be mediated through acid reduction.

Hydroalcoholic extract of Pycnocycla spinosa has spasmolytic action in vitro. At oral dose of 250 mg/kg it inhibits castor oil induced diarrhea in mice. If P. spinosa extract has no serious adverse effect, it would be a good alternative medicine for treatment of diarrhoea and abdominal spasm. In this research, overall systemic effect of P. spinosa extract on mice behavior and reaction was investigated. Single acute dose of the extract (500 mg/kg and 1 mg/kg, i.p.) had negligible effect on animal behavior. However, some changes were seen with dose of 10 mg/kg of the extract. In a group of mice that were treated daily with the extract (10 mg/kg, i.p.) over 3 weeks, a reduction on alertness and marked decrease on defecation was observed on first and second weeks. No significant changes were seen in behavior of the animals that were received the extract in their drinking water over three weeks. Lethal dose causing 50% mortality indicates a large margin of safety.

Iron overload is a serious clinical condition which can be largely prevented by the use of iron-specific chelating agents. In this study، the synthesis and determination of partition coefficients (Kpart) of a range of N-alkyl hydroxypyridinones، as orally active iron chelators، are described. Synthesis of N-aryl hydroxypyridinones was achieved via a single step synthetic pathway. In this method، maltol (2-methyl-3-hydroxypyra-4-one) was reacted with an excess of suitable aryl primary amines under reflux condition in dilute hydrochloric acid at pH about 5. The progress of reactions was monitored by TLC. The reaction mixture was adjusted to pH 7 using sodium hydroxide and the product was collected by filtration. Purification was achieved by re-crystallization from hot methanol. In this work، 1-phenyl-2-methyl-3-hydroxypyridin-4-one، 1- (3-chlorophenyl) -2-methyl-3-hydroxypyridin-4-one، 1- (3-hydro-xyphenyl) -2-methyl-3-hydroxypyridin-4-one، 1- (3-carboxyphenyl) -2-methyl-3-hydroxy-pyridin-4-one and 1- (4-carboxyphenyl) -2-methyl-3-hydroxypyridin-4-one were synthesized. Identification and structural elucidation of compounds were achieved by 1HNMR، IR، elemental analysis، mass spectra and through physical constants. Kpart values of the compounds were also determined in an aqueous/octanol system using an automated continuous flow method (a filter probe method).

DNA amplification using Taq DNA polymerase is one of the most widely used techniques in molecular biology and biotechnology. The aim of this study was to amplify the gene of this enzyme from a thermophilic bacteria called Thermus aqauticus and clone it into a vector for future use. Using specific primers the cDNA of Taq DNA polymerase was amplified and ligated into the cloning vector pTZ57R using TA cloning technique. The recombinant plasmids were identified using restriction enzyme digestion. The presence of the Taq DNA polymerase gene was confirmed by DNA sequencing. In conclusion, Taq DNA polymerase gene has been cloned in our laboratory and can be used for the production of large quantities of this enzyme.

Plants produce a diverse array of secondary metabolites, many of which have antimicrobial activity. Otostegia persica Bioss. (Labiatae) grows in south east of Iran. No previous investigation on antimicrobial effects of this plant has been reported. Thus, the aim of this work was to examine the antimicrobial effect of O. persica extracts with different polarity on several microorganisms. Aerial parts of O. persica were collected from Sistan-Baluchestan province (Iran). Powdered aerial parts of the plant were extracted with ethanol. The extract was concentrated under reduced pressure. The dried extract was extracted with hexane, followed by chloroform and methanol, respectively. Antimicrobial activities of three extracts were tested on several microorganisms using well plate method, MIC (Minimum Inhibitory Concentration) and MBC (Minimum Bactericidal Concentration) methods. O. persica extracts showed antimicrobial activity against Gram positive strains including Listeria monocytogens, Enterococcus fecalis, Staphylococcus aureus, and Staphylococcus epidermidis with MIC values from 0.62 to 20 mg/ml. The MBC values were identical, two, four or eight times higher than MIC values for the corresponding MIC for extracts. The Gram negative strains; Escherichia coli, Pseudomonas aeruginosa, Salmonella spp., Klebsiella spp., and Proteus spp. were not inhibited by O. persica polar, semi-polar, and non-polar extracts. It can be concluded that the tested extracts of O. persica exhibit significant antibacterial activity. This investigation supports the idea of using O. persica extracts as a candidate for further antimicrobial and phytochemical researches.