Utilizing experimental design and desirability function in optimizing RP-HPLC method for simultaneous determination of some skeletal muscle relaxants and analgesics.

An experimental design and response surface methodologies using Plackett-Burman and Box-Behnken designs were applied for selecting and optimizing the most appropriate parameters which significantly affect the separation and quantitative estimation of five skeletal muscle relaxants and four analgesic drugs (baclofen, methocarbamol, dantrolene sodium, orphenadrine citrate, cyclobenzaprine hydrochloride, ketoprofen, etoricoxib, ibuprofen, and mefenamic acid) with a relatively short duration of analysis in a single run. For the separation of the nine drugs, an INERTSIL ODS-V3-5 µm C18 column (250 × 4.6 mm I.D.) was used with the optimum mobile phase conditions (45.15 mM ammonium acetate buffer pH 5.56 adjusted with acetic acid, acetonitrile, and methanol in a ratio of 30.5:29.5:40, v/v/v with a flow rate of 1.5 mL/min) and UV-detection at 220 nm. The optimized method was successfully subjected to the validation steps as described in ICH guidelines for linearity, precision, accuracy, robustness, and sensitivity. The optimized and validated method was effectively applied to determine the content of the studied drugs in their pharmaceutical preparations and to expand its applicability to the counterfeit estimation of etoricoxib in different brands of tablet dosage forms.

Evaluation of anterolateral thigh flap dimensions with virtual flap models.

Free flaps are commonly used for head and neck reconstruction. However, flap dimensions are still evaluated by visual and tactile assessment. The aim of this study was to enable preoperative planning of flap dimensions for soft tissue reconstruction based on clinical parameters. Computed tomography records from 230 patients dated from 2009 to 2019 were analysed retrospectively. A virtual, three-dimensional anterolateral thigh flap model was standardized, aligned to segmented leg models in two positions, and flap thicknesses and volumes were determined. Associations of flap thickness and volume with clinical parameters were evaluated, and an approximative calculation method was derived. The laterally positioned anterolateral thigh flap showed an average (interquartile range) thickness of 15.6 mm (8.7 mm) and volume of 1.5 cm3 (0.9 cm3) per cm2. The medially positioned anterolateral thigh flap showed an average (interquartile range) thickness of 16.3 mm (8.7 mm) and volume of 1.6 cm3 (0.9 cm3) per cm2. For both flap positions, leg circumference was the strongest predictor of flap thickness (ß = 0.545, P < 0.001 and ß = 0.529, P < 0.001) and flap volume (ß = 0.523, P < 0.001 and ß = 0.480, P < 0.001). Flap dimensions can be calculated based on leg circumference, and this preoperative planning of flap dimensions can help the surgeon to select the appropriate flap.

Application of Box-Behnken experimental design and response surface methodology for selecting the optimum RP-HPLC conditions for the simultaneous determination of methocarbamol, indomethacin and betamethasone in their pharmaceutical dosage form.

An isocratic RP-HPLC method has been developed for the separation and determination of methocarbamol (MTL), indomethacin (IND), and betamethasone (BET) in combined dosage form using an Inertsil ODS-3v C18 (250 × 4.6 mm, 5 µm) column with UV- detection at 235 nm. Experimental design using Box-Behnken design (BBD) was applied to study the response surface during method optimization and to achieve a good separation with a minimum number of experimental runs. The three independent parameters were pH of buffer, % of acetonitrile and flow rate of the mobile phase while the peak resolution of IND from MTL and the peak resolution of BET from IND (R2) were taken as responses to obtain mathematical models. The composite desirability was employed to optimize a set of responses overall (peak resolutions). The predicted optimum assay conditions include a mobile phase composition of acetonitrile and phosphate buffer (pH 5.95) in a ratio of 79:21, v/v, pumped at a flow rate of 1.4 mL min-1. With this ideal condition, the optimized method was able to achieve baseline separation of the three drugs with good resolution and a total run time of less than 7 min. The linearity of MTL, IND, and BET was determined in the concentration ranges of 5-600 µg mL- 1, 5-300 µg mL- 1, and 5-300 µg mL- 1 and the regression coefficients were 0.9994, 0.9998, and 0.9998, respectively. The average percent recoveries for the accuracy were determined to be 100.41 ± 0.60%, 100.86 ± 0.86%, and 100.99 ± 0.65% for MTL, IND, and BET, respectively. The R.S.D.% of the intra-day precision was found to be less than 1%, while the R.S.D.% of the inter-day precision was found to be less than 2%. The RP-HPLC method was fully validated with regard to linearity, accuracy, precision, specificity, and robustness as per ICH recommendations. The proposed method has various applications in quality control and routine analysis of the investigated drugs in their pharmaceutical dosage forms and laboratory-prepared mixtures with the goal of reducing laboratory waste, analysis time, and effort.

The impact of tumor-infiltrating lymphocytes on tumor features and pathological characteristics in breast cancer patients: the role of cytotoxic T lymphocytes and regulatory T cells.

OBJECTIVE: Though tumor-infiltrating lymphocytes (TILs) have a predictive impact in cancer patients, their association with presentation and prognosis in breast cancer is less consistent. This study aimed to assess the level of infiltrating cytotoxic T lymphocytes (CTLs) and regulatory T lymphocytes (Tregs) and their association with the clinicopathological features of breast cancer. PATIENTS AND METHODS: Tissue samples from female patients (n=153) diagnosed with primary invasive breast cancer were stained with CD8 (a CTL marker) and Foxp3 (a Treg marker) using immunohistochemistry. RESULTS: CTLs were distributed between tumor bed and stroma whereas Treg cells were mainly located in the stroma. The level of intratumoral CTLs correlated positively with Tregs in both tumor and stroma (rho=0.312, p<0.001 and rho=0.176, p=0.031; respectively). Stromal CTLs correlated positively with stromal Tregs (rho=0.319, p=0.005). Tumor size correlated inversely with the number of Treg cells in the tumor bed (rho= - 0.179, p=0.028). Tregs were associated with lymphovascular invasion status in the tumor bed (p=0.042). The ratio of intratumoral CTLs to Tregs was associated with estrogen receptor positivity and luminal subtype (p=0.029 and p=0.045, respectively). The median number of CTLs was significantly lower in patients using aspirin or antihypertensive medications compared to nonusers (p=0.024 and p=0.03, respectively). CONCLUSIONS: TILs were distributed differently in tumor tissues of breast cancer patients. CTLs infiltrates were found in both tumor bed and stroma while Tregs were dominant in the stroma. TILs were also distinctly associated with tumor features. The impact of TILs on prognosis and treatment outcomes in Jordanian breast cancer patients needs further investigation.

Evaluation of the bactericidal effect of cold atmospheric pressure plasma on contaminated human bone: an in vitro study.

The use of cold atmospheric pressure plasma (CAPP) as a bacterial decontaminant for chronic wounds has shown good results. The purpose of this in vitro study was to evaluate the bactericidal effects of CAPP on the cancellous area of the bone. Sterile glass slides and processed sterile human bone allografts 1, 2, 3, and 4mm thick were used for initial contamination and further CAPP treatment. Each block was contaminated with Staphylococcus aureus suspension on one side. Each slide was turned 180° and treated on the reverse side. The bacterial count in colony-forming units (CFU) was then measured and compared with that of a control group, and the bactericidal effects of CAPP in relation to bone density evaluated. A significant reduction in count was measured between treated and untreated groups (groups A-D: p<0.01 and group E: p=0.04). A strong positive linear relation was found between bone density and the S aureus count (r=0.844, p=0.156). Treatment with CAPP had a bactericidal effect on bone structures with a penetration depth of up to 4mm. It might be used for all diseases involving infected bone, and so extends the existing range of treatments.

Correlation between extent of sinus surgery, radiographic disease, and postoperative outcomes.

BACKGROUND: The extent of endoscopic sinus surgery (ESS) required for optimal outcomes in chronic rhinosinusitis (CRS) is undefined. We evaluated whether concordance between the extent of surgery and degree of radiographic disease influences postoperative outcomes. METHODS: 247 CRS patients who underwent ESS were retrospectively assigned a concordance score reflecting the similarity between the extent of surgery and degree of radiographic disease. 0 points were assigned when sinusotomy was performed on a diseased sinus, or no sinusotomy was performed on a nondiseased sinus; plus 1 for sinusotomy on a nondiseased sinus; and -1 for a diseased sinus left unopened. The total possible score ranged from minus 10 to plus 10. Patients were divided into 5 subgroups according to variance from complete concordance. SNOT-22 scores and revision rates were compared at 6 and 24 months. RESULTS: All five subgroups had similar preoperative SNOT-22 scores and improved at 6 months postoperatively. At 6 months postoperatively, the most conservatively operated and most extensively operated subgroups each achieved equivalent improvements in SNOT-22 as the completely concordant subgroup. At 24 months, the most extensively operated subgroup had a 12.5-point smaller improvement in SNOT-22 scores compared to the completely concordant subgroup. Multivariate analysis showed no association between concordance score and revision rate. CONCLUSIONS: Symptom improvement and revision rates after ESS do not appear to correlate with the degree of concordance between extent of surgery and radiographic disease. More extensive surgery than indicated by CT confers neither greater symptomatic improvement nor long-term detriment.

Evaluation of the lingual fracture patterns after bilateral sagittal split osteotomy according to Hunsuck/Epker modified by an additional inferior border osteotomy using a burr or ultrasonic device.

This study was conducted to compare fracture patterns and operation times after sagittal split osteotomy (SSO) by Hunsuck/Epker approach, performed using a burr or ultrasonic device, with and without osteotomy modification. A total of 80 SSOs were performed in fresh human cadavers using a burr or ultrasonic device to investigate the influence of surgical instruments as well as an additional bone cut on the inferior border of the mandible in terms of lingual fracture patterns. The times required for osteotomy and sagittal split were measured, and postoperative cone beam computed tomography images of all splits were analyzed. Without an additional inferior osteotomy, preferred splits according to Hunsuck/Epker were achieved in 35% of cases (7/20) with the burr and 45% (9/20) with the ultrasonic instrument. The inferior modification resulted in a greater number of unwanted fracture patterns in both groups. There was no relationship between the split technique and the fracture pattern (P=0.7854). Statistically significant differences in osteotomy time were observed between burr osteotomy and modified burr osteotomy (P=0.006), as well as modified ultrasonic osteotomy (P<0.001), but not between burr and ultrasonic surgery both without the inferior cut (P=0.36). The bone cut on the inferior border did not improve split control, but rather increased the risk of unwanted fractures and extended the operation time.