A High-Performance Thin Layer Chromatography (HPTLC) Method for Simultaneous Determination of Diphenhydramine Hydrochloride and Naproxen Sodium in Tablets.

A rapid and simple high-performance thin layer chromatography (HPTLC) method with densitometry at 230 nm was developed and validated for simultaneous determination of diphenhydramine hydrochloride (DPH) and naproxen sodium (NPS) from pharmaceutical preparation. The separation was carried out on aluminum plates precoated with silica gel 60 F254 using mobile phase toluene:methanol:glacial acetic acid (7.5:1:0.2, v/v/v). The linearity range lies between 200 and 1200 ng/band for DPH and 1760 and 10,560 ng/band for NPS with correlation coefficients of 0.994 and 0.995, respectively. The R f value for DPH is 0.20 ± 0.05 and for NPS is 0.61 ± 0.06. % Recoveries of DPH and NPS was in the range of 99.70%-99.95% and 99.63%-99.95%, respectively. Limit of detection value for DPH was 13.21 ng/band and for NPS was 8.03 ng/band. Limit of quantitation value for DPH was 40.06 ng/band and for NPS was 24.34 ng/band. The developed method was validated as per ICH guidelines. In stability testing, DPH was found unstable to acid and alkaline hydrolysis, and DPH and NPS were found unstable to oxidation, whereas both the drugs were stable to neutral and photodegradation. The proposed method was successfully applied for the routine quantitative analysis of dosage form containing DPH and NPS.

Spectrophotometric and HPLC methods for simultaneous estimation of amlodipine besilate, losartan potassium and hydrochlorothiazide in tablets.

Two UV-spectrophotometric and one reverse phase high performance liquid chromatography methods have been developed for the simultaneous estimation of amlodipine besilate, losartan potassium and hydrochlorothiazide in tablet dosage form. The first UV spectrophotometric method was a determination using the simultaneous equation method at 236.5, 254 and 271 nm over the concentration range 5-25, 10-50 and 5-25 mug/ml for amlodipine besilate, losartan potassium and hydrochlorothiazide, respectively. The second UV method was a determination using the area under curve method at 231.5-241.5, 249-259 and 266-276 nm over the concentration range of 5-25, 5-25 and 10-50 mug/ml for amlodipine besilate, hydrochlorothiazide and losartan potassium, respectively. In reverse phase high performance liquid chromatography analysis is carried out using 0.025 M phosphate buffer (pH 3.7):acetonitrile (57:43 v/v) as the mobile phase and Kromasil C18 (4.6 mm i.dx250 mm) column as stationery phase with detection wavelength of 232 nm linearity was obtained in the concentration range of 2-14, 20-140 and 5-40 mug/ml for amlodipine besilate, losartan potassium and hydrochlorothiazide, respectively. Both UV-spectrophotometric and reverse phase high performance liquid chromatography methods were statistically validated and can be used for analysis of combined dose tablet formulation containing amlodipine besilate, losartan potassium and hydrochlorothiazide.

Simultaneous spectrophotometric estimation of norfloxacin and ornidazole in tablet dosage form.

Three simple, accurate and economical methods have been developed for the estimation of norfloxacin and ornidazole in tablet dosage form. First method is based on the simultaneous equations, wavelengths selected for analysis were 273.0 nm (lambda(max) of norfloxacin) and 318.5 nm (lambda(max) of ornidazole), respectively, in 0.1N NaOH. Second method is Q-analysis method, based on absorbance ratio at two selected wavelengths 297.0 nm (iso-absorptive point) and 318.5 nm (lambda(max) of ornidazole). Third method is first order derivative spectroscopy using 297.5 nm (zero cross for norfloxacin) and 264.0 nm (zero cross for ornidazole). The linearity was obtained in the concentration range of 4-20 mug/ml and 5-25 mug/ml for norfloxacin and ornidazole, respectively. The results of the analysis have been validated statistically and by recovery studies.

Simultaneous estimation of amlodipine besilate and olmesartan medoxomil in pharmaceutical dosage form.

Two UV Spectrophotometric and one reverse phase high performance liquid chromatography methods have been developed for the simultaneous estimation of amlodipine besilate and olmesartan medoxomil in tablet dosage form. First UV spectrophotometric method was a determination using the simultaneous equation method at 237.5 nm and 255.5 nm over the concentration range 10-50 mug/ml and 10-50 mug/ml, for amlodipine besilate and olmesartan medoxomil with accuracy 100.09%, and 100.22% respectively. Second UV spectrophotometric method was a determination using the area under curve method at 242.5-232.5 nm and 260.5-250.5 nm over the concentration range of 10-50 mug/ml and 10-50 mug/ml, for amlodipine besilate and olmesartan medoxomil with accuracy 100.10%, and 100.48%, respectively. In reverse phase high performance liquid chromatography analysis carried out using 0.05M potassuim dihydrogen phosphate buffer:acetonitrile (50:50 v/v) as the mobile phase and Kromasil C18 (4.6 mm i.d.x250 mm) column as the stationery phase with detection wavelength of 238 nm. Flow rate was 1.0 ml/min. Retention time for amlodipine besilate and olmesartan medoxomil were 3.69 and 5.36 min, respectively. Linearity was obtained in the concentration range of 4-20 mug/ml and 10-50 mug/ml for amlodipine besilate and olmesartan medoxomil, respectively. Proposed methods can be used for the estimation of amlodipine besilate and olmesartan medoxomil in tablet dosage form provided all the validation parameters are met.

High performance thin layer chromatographic method for simultaneous estimation of Ibuprofen and pseudoephedrine hydrochloride.

High performance thin layer chromatographic method is developed for simultaneous estimation of ibuprofen and pseudoephedrine hydrochloride in tablets. Silica gel 60F(254) plates were used as stationary phase and t.butanol: ethyl acetate: glacial acetic acid: water (7:4:2:2 v/v) as mobile phase. Wavelength selected for analysis was 254 nm. Percent estimation of ibuprofen and pseudoephedrine hydrochloride was found to be 99.56% and 98.77%, respectively. Percent recovery for both the drugs was found in the range of 98.27% to 100.91%, respectively.

Comparison of four nonculture diagnostic tests for Chlamydia trachomatis infection.

OBJECTIVE: Chlamydia trachomatis (CT) is one of the commonest sexually transmitted diseases leading to urethritis, epididymitis, prostatitis in men and urethritis, cervicitis, endometritis and pelvic inflammatory disease, sometimes complicated by infertility and ectopic gestation in women. Since culture of fastidious bacteria in a monocellular medium is not available in most laboratories we compared direct immunofluorescence antigen detecting test (DFA) with three other nonculture tests-antigen detecting enzyme immunoassay (EIA), Papanicolaou staining (Pap) and Geimsa stain for endocervical swabs from women in reproductive age group. METHODS: Three hundred and fifty seven women between 16 and 41 years of age and attending family welfare clinics of IRR were evaluated for the presence pap smears. In 100 cases DFA staining was compared with Geimsa staining. RESULTS: DFA test was positive in 60/357 (16.8%), EIA in 29 (8.1%) of cases and Pap smear in 37 (10%) cases. In the second group DFA was positive in 17 (17%) and Geimsa in 10 (10%) cases. CONCLUSION: Amongst the four tests DFA showed maximum sensitivity. ELISA is less expensive but has lower sensitivity. Pap stain also has less sensitivity and good specificity, the quality of smear is likely to affect the diagnosis. Though Geimsa stain is cheapest, for chlamydial cervicitis in our experience it was not as sensitive as DFA. Thus each laboratory must decide the method depending on its resources.

Chlamydia trachomatis detected in cervical smears from Copper-T users by DFA test.

OBJECTIVE: This study was carried out to determine the prevalence of Chlamydia trachomatis (CT) antigen in endocervical smears from women using the Copper-T200 (Cu-T) intrauterine device and comparing them with nonusers. METHODS: The direct fluorescent antibody (DFA) test was used to detect the CT antigen (CT Ag). RESULTS: A total of 422 women between 17 and 42 years of age was evaluated. Out of these 71 (16.8%) were positive for CT Ag. Among Cu-T users (n = 222), 14.0% were positive. The duration of Cu-T use varied from 6 to 80 months. Among nonusers (n = 200), the DFA test was positive in 20.0% (p = 0.11, NS). However, as a whole, symptoms related to genital tract infection were significantly more common in Copper-T users than in nonusers (p < 0.02) and signs were relatively more common in Cu-T users but not significantly so (p = 0.16). Similarly, as a group, symptoms as well as signs were significantly more common in all DFA-positive cases than in all negative cases (p < 0.02). Among Cu-T users, symptoms were relatively more common, and signs were significantly more common, in women positive for CT Ag than in the negative cases (p < 0.04). There were two cases of mild pelvic inflammatory disease (PID) diagnosed clinically, one in a Copper-T user, and the other a nonuser, both being positive by the DFA test. CONCLUSION: There was no correlation between the duration of Copper-T use and percent positivity for CT Ag. Although milder genital tract symptoms and signs were more common in women with chlamydial cervicitis, clinically PID appears to be uncommon (0.5%). PID in this study was not related to Copper-T use but related to chlamydial cervicitis in this group of women with low risk sexual behavior.

Ultrasonographic monitoring of ovarian follicles in women using norethisterone for contraception.

OBJECTIVE: The purpose of this study was to examine the effect of intranasal and oral norethisterone (NET) on ovarian folliculogenesis. METHODS: Sixteen healthy, sterilized women with regular menstrual cycles were recruited to the study. NET 300 micrograms per day was administered orally (n = 8) or intranasally (n = 8) for two consecutive menstrual cycles. Serial pelvic ultrasonography was performed to monitor ovarian follicular growth. RESULTS: Ultrasonographic evidence of normal follicular growth and ovulation was observed in 10 cycles whilst 22 cycles were anovulatory. Formation of follicular cysts was seen in 14 cycles, 13 of which were anovulatory and in one ovulation was observed in the opposite ovary. The size of the cysts varied between 27 and 44 mm. The cysts disappeared when NET treatment was discontinued. A positive correlation between cyst size and estradiol levels was observed with intranasal NET in 50% of cyst cycles. In three cycles, although normal follicular growth and endocrine profile were observed, the follicles failed to rupture. These were classified as luteinized unruptured follicles. Immature follicles < 10 mm were seen in six cycles. CONCLUSION: The study showed that NET administered either orally or intranasally evidently disturbs normal follicular growth and rupture.

The prevalence of Chlamydia trachomatis in young women.

BACKGROUND: Chlamydia trachomatis infection is common in women with pelvic inflammatory disease, infertility and ectopic gestation. In this study we examined the prevalence of this infection in young women undergoing routine gynaecological checks. METHODS: Three hundred and five women aged between 19 and 41 were included in our study. Endocervical smears were collected by speculum examination, air-dried, acetone-fixed and stained for chlamydial antigen using fluorescein-labelled monoclonal antibody. RESULTS: Ninety-five per cent of the women were asymptomatic. Only 5% had mild symptoms such as white discharge, abdominal pain or profuse periods; 34% gave a history of white discharge only. Chlamydial antigen was detected in cervical smears from 47 (15%) women. Of these, 9 (19%) had symptoms at examination and 20 (43%) had symptoms within 2 months. On gynaecological examination 25 women (53%) had minor signs such as erosion or cervicitis and only 1 (2%) had uterine tenderness suggestive of mild pelvic inflammatory disease. CONCLUSIONS: It is important to diagnose chlamydial infection early because in its later stages it is associated with an ascending infection and serious complications. Young, relatively asymptomatic women should be screened for the presence of this organism.

Early pregnancy factor (EPF) as a marker for the diagnosis of subclinical embryonic loss.

The validation of EPF as a possible correlate of early fertilization has made it possible to study and detect fertilization of the ovum in normal fertile women (during the luteal phase) and also in women with infertility, where the fertilization of the ovum may not be affected but there may be impairment in early embryonic development which results in early embryo loss or subclinical embryo loss. Our results have suggested that using EPF as a marker, we could detect subclinical embryonic loss in 57.8% of the infertile women where more than one menstrual cycle was studied and the blood was collected 4-7 days after ovulation. After the missed period, 80% of the patients who were negative for EPF but positive for hCG had spontaneous abortions. It would be interesting to study how EPF behaves as a marker, to detect subclinical embryonic loss in diverse pathological situations such as recurrent abortions, parental age and translocation carrier parents.

Effects of intranasal administration of norethisterone on folliculogenesis, cervical mucus, vaginal cytology, endometrial morphology and reproductive-endocrine profile in women.

The effects of intranasal administration of norethisterone (NET) on menstrual cycle length, folliculogenesis, serum levels of estradiol, FSH, LH and progesterone, vaginal cytology, cervical mucus and endometrial morphology were studied in 8 volunteers (age 28 to 39 years, weighing between 46 and 54 kg). The study period comprised 4 consecutive menstrual cycles. In the first cycle (pretreatment cycle), only the vehicle (alcohol, propylene glycol, water; 3:3:4) was sprayed intranasally (100 microliters in each nostril), using a metered nebulizer, once daily from day 3 to the last day of menstrual cycle. In the next two cycles (treatment cycles), NET (300 micrograms/day) was administered once daily, starting from day one of menstrual cycle, between 9 and 10 a.m. The fourth cycle was a post-treatment cycle in which the volunteers were monitored for recovery. Blood samples (about 5 ml each) were collected once daily from day 8 to 24 and thereafter on alternate days until the last day of cycle during all the 4 cycles. Levels of estradiol, FSH, LH and progesterone were measured in the serum samples by radioimmunoassay methods. Cervical mucus samples and vaginal smears were collected once daily starting from day 7 or 8 of each cycle until the mucus was very scanty. Serial pelvic ultrasonography was performed starting from day 7 or 8 until the growing follicle disappeared or throughout the cycle in case a growing follicular cyst was observed. Endometrial aspirates were collected once around day 22 in each cycle and processed for routine histological examination.

Ultrasonographically observed preterm grade III placenta and perinatal outcome.

Ultrasound studies of placenta were conducted in 270 singleton normal pregnancies. Women were enrolled between 31 and 34 weeks of gestation and were followed up for the outcome of pregnancy. Women with grade III placental maturity comprised the study group (n = 64) and those with grade I placenta were enrolled as control group (n = 206). Another 100 normal women were enrolled to note the prevalence of grade III placenta at term. There was an increased incidence of intrauterine growth retardation (6.20%) and fetal distress (7.8%) in the study group compared with the control group (nil), which was statistically significant. The incidence of low birth weight was also higher (34.37%) in the study group compared with the control group (22.33%). Three women in the study group developed preeclampsia at subsequent follow up visit but none in control group (P less than 0.01). Prevalence rate of grade III placenta at term was 28%. In view of these findings preterm grade III placenta is found to be a sensitive predictor of poor perinatal outcome.

Serum progesterone and norethisterone levels following injection of norethisterone enanthate in different sites and doses.

Serum progesterone (P) and norethisterone (NET) levels following injection of norethisterone enanthate (NETEN) were studied in 11 regularly menstruating women. In 6 subjects 200 mg NETEN was administered to the gluteal (IG) or deltoid (ID) region sequentially. The serum P levels remained anovulatory (less than 4 ng/mL) up to 12 weeks. Serum NET levels were similar whether injection was given IG or ID (paired t - test, NS). In 5 subjects given a 150-mg dose, serum P levels remained anovulatory up to 11 weeks. Serum NET levels declined faster than with the 200-mg dose, but the difference was not significant (Student's t - test, NS). Wide inter-subject variations were seen in both groups. NETEN therefore can effectively be administered in gluteal or deltoid regions. A lower dose of 150 mg may have contraceptive potential on an 8-week schedule.

Urinary estrone glucuronide, pregnanediol glucuronide and human chorionic gonadotrophin in threatened abortion.

Estimations of urinary estrone glucuronide, pregnanediol glucuronide and human chorionic gonadotrophin were carried out by ELISA to see their potential in predicting an abnormal outcome in cases with vaginal bleeding in early pregnancy. Reference values were set up with samples from women without bleeding in present or past pregnancies and with normal ultrasonic findings. None of the parameters were found to be sensitive enough to predict an abnormal outcome. However, predictability of an abnormal value was found to be 95% for estrone-3-glucuronide (E1G), 93% for pregnanediol glucuronide (PdG) and 87% for human chorionic gonadotrophin (hCG).