Titration Method Validation of Live Vaccines against Infectious Bursal Disease

Validating a method of analysis goes through different steps, which aims at testing the normality of measurements distribution, estimating the uncertainty of the components of a measurement （i.e., accuracy and correctness）, and finally, define the control tests of non degradation of the method performances. This paper outlines the steps for validating a biological method of analysis. It involves the construction of an experimental design, a statistical model, and the preparation of an interne laboratory reference material （pilot vaccine）. The latter is used to study the impact of deviation and variation factors, in order to, optimize the analytical method, to evaluate the bias （random error）, and to calculate the uncertainty of measurement, and make the control charts. This method is applied in the titration of live viral vaccines of Gumboro disease on chicken＇s embryos fibroblasts. The experimental results show that potential influence factors related to the titration method had no significant influence on the obtained results. Taking into account these results, an operating mode has been elaborated. The finalized method proved to be faithful to standard deviation of repeatability and reproducibility of 0.21 and 0.22, respectively, with a confidence level of 95%. The calculated uncertainty of measurement is equal to 0.2, which represents the average error level of a titer. A homogeneous stock of interne laboratory reference vaccine （MRIL）, with an average titer of 5.9 log DIT 50, was produced and the control chart set in away to provide the laboratory with an important tool of control and monitoring of the viral titers evolution in time, as well as, the mastery of the validated titration method performances.

Robustness Study and Superior Method Development and Validation for Analytical Assay Method of Atropine Sulfate in Pharmaceutical Ophthalmic Solution

Background: The robustness is a measurement of an analytical chemical method and its ability to contain unaffected by little with deliberate variation of analytical chemical method parameters. The analytical chemical method variation parameters are based on pH variability of buffer solution of mobile phase, organic ratio composition changes, stationary phase (column) manufacture, brand name and lot number variation;flow rate variation and temperature variation of chromatographic system. The analytical chemical method for assay of Atropine Sulfate conducted for robustness evaluation. The typical variation considered for mobile phase organic ratio change, change of pH, change of temperature, change of flow rate, change of column etc. Purpose: The aim of this study is to develop a cost effective, short run time and robust analytical chemical method for the assay quantification of Atropine in Pharmaceutical Ophthalmic Solution. This will help to make analytical decisions quickly for research and development scientists as well as will help with quality control product release for patient consumption. This analytical method will help to meet the market demand through quick quality control test of Atropine Ophthalmic Solution and it is very easy for maintaining (GDP) good documentation practices within the shortest period of time. Method: HPLC method has been selected for developing superior method to Compendial method. Both the compendial HPLC method and developed HPLC method was run into the same HPLC system to prove the superiority of developed method. Sensitivity, precision, reproducibility, accuracy parameters were considered for superiority of method. Mobile phase ratio change, pH of buffer solution, change of stationary phase temperature, change of flow rate and change of column were taken into consideration for robustness study of the developed method. Results: The limit of quantitation (LOQ) of developed method was much low than the compendial method. The % RSD for the six sample assay of developed method was 0.4% where the % RSD of the compendial method was 1.2%. The reproducibility between two analysts was 100.4% for developed method on the contrary the compendial method was 98.4%.

Solriamfetol impurities:Synthesis,characterization,and analytical method(UPLC-UV)validation

Given that impurities may affect the quality and safety of drug products,impurity identification and profiling is an integral part of drug quality control and is particularly important for newly developed medications such as solriamfetol,which is used to treat excessive daytime sleepiness.Although the highperformance liquid chromatography analysis of commercial solriamfetol has revealed the presence of several impurities,their synthesis,structure elucidation,and chromatographic determination have not been reported yet.To bridge this gap,we herein identified,synthesized,and isolated eight processrelated solriamfetol impurities,characterized them using spectroscopic and chromatographic techniques,and proposed plausible mechanisms of their formation.Moreover,we developed and validated a prompt impurity analysis method based on ultrahigh-performance liquid chromatography with UV detection,revealing that its selectivity,linearity,accuracy,precision,and quantitation limit meet the acceptance criteria of method validation stipulated by the International Council for Harmonization of Technical Requirements for Pharmaceuticals for Human Use.Thus,the developed method was concluded to be suitable for the routine analysis of solriamfetol substances.

Methodology Validation of Microbial Limit Test of Bupi Qiangli Ointment

[Objective] This study was conducted to establish a microbial limit test method for Bupi Qiangli Ointment. [Method] The conventional method and medium dilution method were used for bacterial, mold and yeast counting in sample recovery test. [Result] The medium dilution method （1：10 test solution, 0.5 ml/plate） could effectively eliminate the inhibition effect of the Bupi Qiangli Ointment, and the recovery of Staphylococcus aureus was greater than 70% in the 3 batches of samples; and the conventional method exhibited the recoveries of E. coil, Bacillus subtilis, Candida albicans and Aspergillus greater than 70% in the 3 batches of samples. [Conclusion] Due to Bupi Qiangli Ointment has strongly antibacterial effect on Staphylococcus au- reus, the medium dilution method was used for bacterial counting, and the conventional method was used for mold and yeast counting; and the conventional method was used for controlled bacterium examination of E. coll.

Oocyte vitrification:A local validation of the method

Is there a really need to validate oocyte vitrification technique in an ART laboratory before establishing it in daily practice? Validation of micromanipulationbased technique, in this case oocyte vitrification, is essential prior to enlarging its use to routine practice. Oocyte vitrification is a new worldwide used technique and legal recently inFrance. This micromanipulation needs to be performed by a skilled and experienced embryologist and requires an internal assessment in each ART unit before any wide use. We designed a prospective study, from September 2011 to July 2012, using sibling oocytes from women who recovered more than 12 Metaphase II oocytes. A part of freshly recovered oocytes underwent immediate ICSI while the remaining oocytes were vitrified. 87 couples undergoing ICSI were selected based on number of mature oocytes available on the recovery day after denudation. A part of fresh MII oocytes were microinjected and the others were vitrified using an open system (Cryotop?). The major criterion of interest was the number of embryo transferred/ number of Metaphase II ratio for after ICSI on fresh oocytes (42/211) versus vitrified/warmed oocytes (51/204) (p > 0.05). Secondary studied criteria were survival rate (80.5% ± 26.3%), fertilization rate (68.9 ± 33.5) and finally, cumulative pregnancy rate obtained in this study is 40.2%. One of the benefits of such practice is the limitation of embryo freezing. However, the study design delays oocytes warming cycles, due to pregnancies triggered by the transfer of fresh derived oocyte embryos and to the priority to transfer all the frozen embryos before starting oocytes warming. Moreover, no data is available about children’ health. Oocyte vitrification represents not only a change in our daily practice to improve cumulative pregnancy rate but also a promising tool to develop egg banking and donation. Clinical Trials Registration number: 209 R02.

Comparative Analysis and Validation Methodologies of GC and HPLC for Analysis of Cholesterol in Meat Products

This study validated different extraction methodologies and compared the quantification of cholesterol by gas chromatography (GC) and high performance liquid chromatography (HPLC) in mg per 100 g of Italian-type salami and traditional bologna. The GC method used was direct saponification of the samples without derivatizations and the HPLC method was used to extract of the lipid samples. The GC limits of detection and quantification obtained for cholesterol were, respectively, 0.001 and 0.003 mg.g–1. The HPLC values were 0.005 mg.g–1 and 0.016 mg.g–1. The GC recovery rate was 97.10 ± 0.13 and that of HPLC was 93.33 ± 0.22. Comparison of the cholesterol quantity found using the two chromatographic techniques shows that both are capable of quantifying cholesterol in the foods. With regard to costs, analysis time, the cost/benefit relationship was better with gas chromatography than that obtained with high performance liquid chromatography.

Comparison of Crop Model Validation Methods

In this paper, the many indices used in validation of crop models, such as RMSE （root mean square errors）, Sd （standard error of absolute difference）, da （mean absolute difference）, dap （ratio of da to the mean observation）, r （correlation）, and R2 （determination coefficient）, are compared for the same rice architectural parameter model, and their advantages and disadvantages are analyzed. A new index for validation of crop models, dap between the observed and the simulated values, is proposed, with dap〈5% as the suggested standard for precision of crop models. The different kinds of validation methods in crop models should be combined in the following aspects：（1） calculating da and dap; （2） calculating the RMSE or Sd; （3） calculating r and R2, at the same time, plotting 1：1 diagram.

Experimental validation method of elastic thin rod model for simulating the motional cable harness

To analyze the spring disturbance torque caused by motionai cable harness in a stabilized platform, the Kirchhoff theory based cable harness model has been previously developed to dynamically simulate the motional cable harness. In this paper, this model was validated by comparing the simulation results with the experiment results （ both the spring force and the deformed profile of the motional cable harness）. In the experiment, a special optical measuring instrument based on binocular vision was developed and the motion and deformation of cable harness were measured. A simpli- fied stabilized platform system was constructed, and the absolute value of spring disturbance force during the motion of this simplified frame was obtained by using a force gauge （0. 02 N precision）. The physical parameters of experimental specimen were also measured. The experimental and simulated results showed good agreement. These results should be useful for better motional cable harness layout design and reliable evaluation of the spring disturbance torque.

Influence of in vitro Methods, Receptor Fluids on Percutaneous Absorption and Validation of a Novel in vitro Method

In vitro experiment using excised skin has been valuable for studying the mechanism of percutaneous absorption. Based on previously established static diffusion cell system in this laboratory, a novel model-peifused glass diffusion cell system is desboed. The results of initial comporative study on percutaneous absorption between glass perfused diffusion cell and static diffusion cell, ih vitro and in vivo permeation as well as factors affecting permeation with seven radiolabelled chemicals are presented. The results demonstrate that the peifused diffusion cell system, which used a perfusion nuid betow the suiface of skin to take up the materials which penetrated the skin, is more similar to physiologic condition,convenient and automatic than that of the static cell. It well predicts the in vivo percutaneous absorption if appropriate areptor fluid is chosen. The results also show that the selection of receptor fluid is critical for in vitro permeation of chemicals with different soubility

Nurses’ movements within and between various paths when improving their communication skills—An evaluation of validation method training

Aims and objectives: To explore any changes in nurses’ skills at communicating with residents with dementia disease when using the validation method, as observed in one-to-one videotaped conversations. Background: Communication difficulties due to cognitive impairment among residents with dementia disease may complicate care situations. Training can improve nurses’ communication skills and increase care quality. The validation method aims to facilitate communication with residents with dementia disease through empathic and confirmatory approaches. Evaluations of the validation method have primarily focused on the residents’ perspective, and reports on nurses’ experiences are sparse. Improved communication and relationships with residents after validation method training have been described previously. Videotaped data could provide additional information about these earlier results. Design: A descriptive qualitative design. Methods: Eight nurses participated in a year of validation method training, including videotaped conversations with eleven residents. Videotapes with at least five months between the first and last recording were analysed and compared qualitatively. Results: The analysis revealed an overall pattern: nurses’ movements within and between various paths when improving their communication skills. This was based on three sub-patterns: from controlling communication towards developing attentiveness in communication, from ambiguous communication towards developing coherence in communication, and from being open and attentive towards having a refined attuned communication. Conclusions: All nurses developed their communication skills during the programme, albeit to different degrees. The findings are in congruence with the experiences described by nurses, and so it is reasonable to believe that the programme helped to improve the nurses’ skills in communicating with residents with dementia disease. Relevance to clinical practice: A validation method training programme could give nurses the possibility to develop their skills in communicating with residents with dementia disease.

Quantification of Structurally Alert Mutagenic Impurities in Meropenem Trihydrate Drug Substance by Liquid Chromatography with High Resolution Mass Spectrometer (LC-HRMS)

Potential mutagenic impurities in Active Pharmaceutical Ingredient, Meropenem Trihydrate were assessed and a novel analytical method for their quantification was developed and validated. This Liquid Chromatographic method using High Resolution Mass Spectrometer (LC-HRMS) technique is proved to be suitable for simultaneous quantification of all ten identified impurities with required specificity, sensitivity, resolution, precision, accuracy, and other method characteristics as per ICH Guidelines. The acceptable limit of less than 2.9 μg/g was considered for evaluations, based on drug substance dosage and duration of treatment. The method stands most sensitive with a Limit of Detection of 0.35 μg/g, considering the challenge full acceptance criteria as per current regulatory standards.

Development of an analytical method for multi-residue quantification of 18 anthelmintics in various animal-based food products using liquid chromatography-tandem mass spectrometry

In this study,we developed a simple screening procedure for the determination of 18 anthelmintics(including benzimidazoles,macrocyclic lactones,salicylanilides,substituted phenols,tetrahydropyrimidines,and imidazothiazoles)in five animal-derived food matrices(chicken muscle,pork,beef,milk,and egg)using liquid chromatography-tandem mass spectrometry.Analytes were extracted using acetonitrile/1% acetic acid(milk and egg)and acetonitrile/1% acetic acid with 0.5 mL of distilled water(chicken muscle,pork,and beef),and purified using saturated n-hexane/acetonitrile.A reversed-phase analytical column and a mobile phase consisting of(A)10 mM ammonium formate in distilled water and(B)methanol were used to achieve optimal chromatographic separation.Matrix-matched standard calibration curves(R^(2)≥0.9752)were obtained for concentration equivalent to ×1/2,×1,×2,×3,×4,and×5 fold the maximum residue limit(MRL)stipulated by the Korean Ministry of Food and Drug Safety.Recoveries of 61.2e118.4%,with relative standard deviations(RSDs)of ≤19.9%(intraday and interday),were obtained for each sample at three spiking concentrations(×1/2,×1,and ×2 the MRL values).Limits of detection,limits of quantification,and matrix effects were 0.02e5.5 mg/kg,0.06e10 mg/kg,and -98.8 to 13.9%(at 20 μg/kg),respectively.In five samples of each food matrix(chicken muscle,pork,beef,milk,and egg)purchased from large retailers in Seoul that were tested,none of the target analytes were detected.It has therefore been shown that this protocol is adaptable,accurate,and precise for the quantification of anthelmintic residues in foods of animal origin.

Validation for the Determination of Extractable Metals in Ashes from Biomass Incinerators by ICP-OES

Biomass ashes contain valuable elements for plant growth, but also often high amounts of harmful heavy metals which limit their reuse in forests and on grassland. For monitoring metal concentrations, a fast and reliable method allowing the quantitative determination even at trace levels is mandatory. Therefore an analytical method consisting of an acidic microwave assisted digestion prior to ICP-OES measurement was optimized. The elements of interest were AI, As, B, Ba, Cd, Co, Cr, Cu, Fe, Mg, Mn, Na, Ni, Pb, Sb, St, Ti, V, and Zn. A validation including the following parameters was performed for each analyte： limit of detection, limit of quantification, recovery by spiking experiments, accuracy by analyzing a certified reference material, reproducibility （wihtin-24 hours, 4 and 7 days）, linearity, and uncertainty of measurement. Two standards series were used for calibration - one with 1 M HNO3 and one with additionally 2 g/L Mg（NOD2 to investigate the elimination or decrease of possible matrix effects. Both calibration methods showed very good recoveries. The method optimized allows the determination of the relevant elements covered by the recommendation of the Austrian ministry for agriculture and represents thus an appropriate tool for monitoring the metal contents in biomass ashes.

Stability Indicating HPLC Method for Quantification of Solifenacin Succinate &Tamsulosin Hydrochloride along with Its Impurities in Tablet Dosage Form

A novel stability-indicating RP-HPLC method was developed and validated for simultaneous determination of Solifenacin Succinate & Tamsulosin Hydrochloride and its impurities in tablet dosage form. The method was developed using L1 column with gradient using the mobile phase consist of solvent-A (pH = 6.6, phosphate buffer + 0.5% Triethylamine) and solvent-B (90% Acetonitrile). The eluted compounds were monitored at 225 nm. Solifenacin Succinate & Tamsulosin Hydrochloride was subjected to oxidative, acid, base, hydrolytic, thermal and photolytic stress conditions. The developed method was validated as per ICH guidelines with respect to specificity, linearity, limit of detection, limit of quantitation, accuracy, precision and robustness. The limit of quantification results was ranged from 0.135 - 0.221 μg/mL for Solifenacin Succinate impurities and 0.043 - 0.090 μg/mL for Tamsulosin Hydrochloride impurities. This method is suitable for the estimation of impurities and assay of Solifenacin Succinate & Tamsulosin Hydrochloride in tablets dosage form.

Systematical approach in evaluation of LC method for determination of raloxifene hydrochloride and its impurities employing experimental design

Method validation presents a detailed investigation of analytical method and provision of the evidence that the method, when correctly applied, produces results that fit to the purpose. In order to achieve the method validation scope efficiently, experimental design presents a very useful tool. The greatest benefits of such approach could be seen in robustness testing through the provision of very useful data about the control of the chromatograp6hic system during the routine application. In this paper, robustness testing of the LC method proposed for the determination of raloxifene hydrochloride and its four impurities was done employing Plackett-Burman design. Applying this design, the effect of five real factors （acetonitrile content, sodium dodecyl sulfate content, column temperature, pH of the mobile phase and flow rate） on the corresponding resolution factors was investigated through twelve experiments. Furthermore, the insignificance intervals for significant factors were calculated and the parameters for system suitability tests were defined. Eventually, the other validation parameters were tested and the effectiveness of the proposed analytical method with a high degree of accuracy was confirmed.

Stability Indicating RP-HPLC Method for Quantification of Impurities in Valsartan and Hydrochlorothiazide FDC Tablet Dosage Form

A stability-indicating RP-HPLC method has been developed and validated for simultaneous determination of Valsartan & Hydrochlorothiazide and their impurities in FDC (Fixed Dose Combination) tablet dosage form. The method was developed using L1 column (250 × 4.6 mm;5 μm) with gradient elution using the mobile phase consisting of solvent-A (0.1% Ortho phosphoric acid) and solvent-B (100% Acetonitrile);the gradient program (Tmin/%B) was set as 0/10, 5/10, 20/60, 40/60, 41/10 and 50/10. The eluted compounds were monitored at 265 nm. The developed method was validated as per ICH guidelines with respect to specificity, linearity, limit of detection, limit of quantitation, accuracy, precision and robustness. The influence of Acid, Alkaline, Oxidative, Photolytic, Thermal and Humidity stress conditions, on drug product was studied. The limit of quantification results of Valsartan, Hydrochlorothiazide and their impurities are, VAL: 0.303 μg/mL, HCTZ: 0.019 μg/mL, VAL RC-B: 0.085 μg/mL, VAL RC-C: 0.327 μg/mL, HCT RC-A: 0.017 μg/mL, CTZ: 0.080 μg/mL and 5-Chloro HCT: 0.047 μg/mL. The proposed method is suitable for the estimation of Valsartan & Hydrochlorothiazide impurities in tablets dosage form.

A New and Enantioselective Chromatographic Method for Linagliptin on Amylose Coated Chiral Stationary Phase

A new and enantioselective liquid chromatographic method was developed for estimation of S-Linagliptin in Linagliptin (LINA) drug substances. The desired enantiomeric separation was achieved on Chiralpak AD-H (250*4.6 mm*5 μm) column with the mobile phase composition of ethanol, methanol and diethylamine in a ratio of 90:10:0.1 (v/v/v) with flow rate of 0.5 mL·min-1 and column oven temperature 30°C and the eluted compounds were monitored at 225 nm. In the proposed chiral method, USP resolutions between both the enantiomers were more than 5.0. Limit of detection and Limit of quantitation of S-LINA was found to be 0.03 μg·mL-1 and 0.10 μg·mL-1 respectively. Linearity study was conducted from LOQ to 150% and correlation coefficient found to be 0.9997. Accuracy was within the range of 98.6% to 101.5%. To prove selectivity power of the method specificity study was conducted by subjecting drug substance to acid, base, hydrolysis, oxidation and photolysis and ensured the peak purity of analyte in degraded samples. Moreover, the method has been fully validated as per ICH guidelines. The proposed method is precise, accurate, linear, rugged, robust and suitable for accurate quantification of S-LINA in LINA drug substance.

Enantioseparation of Palonosetron Hydrochloride and Its Related Enantiomeric Impurities by Computer Simulation and Validation

A rapid, simple and single stereo selective high-performance liquid chromatographic (HPLC) method was developed and validated for enantiomers of palonosetron hydrochloride (PALO) and its process related chiral impurities. A computer simulating software was used for the development of chiral method. The developed method was able to separate not only the enantiomers of palonosetron hydrochloride but also its process related chiral impurities within 12 min. The chromatographic separation was carried out by normal phase chromatography using a 3 μm column of cellulose based chiral stationary phase (Chiralcel-OD 250mm × 4.6mm) with a mobile phase comprised of n-hexane: ethanol: methanol: heptafluoro butyric acid: diethyl amine (70:15:15:0.05:0.1, v/v) at a flow rate of 1.0 mL/min. The effects of additive concentration as well as nature of polar organic modifier, flow rate, and temperature on enantioselectivity were investigated. The limit of detection (LOD) and limit of quantification (LOQ) of the palonosetron isomers and its related chiral impurities were found to be in the range 0.06-0.10 μg/mL and 0.14 - 0.24 μg/mL respectively. The method showed excellent linearity (R2 > 0.998) over a range of 0.14 to 1.125 μg/mL. The percentage recovery of the isomers in bulk drug samples ranged from 87.0 to 116.0.

Key problems in validation of intelligent models

To provide a realistic simulation environment for users,intelligent models have become key components in military simulations. After the analysis of modeling nature of intelligent models,the validation criteria for defining the validation points and validation metrics for measuring the agreements between human experts and intelligent models were presented. Further,such methods as graphical comparison,feature analysis and face validation were discussed according to the characteristics of intelligent models. Based on the validation criteria,validation metrics and validation methods,the intelligent models can be effectively validated,which has been proved in current developed intelligent models.

Analysis Method for Pesticide Residues in Biological Matrices： Gas Chromatography-mass Spectrometry

Pesticides have done a great service to human, but their use is not safe for public health. Apart from pesticides acute toxicity, their chronic toxicity can cause various problems for human health. The objective of this work was to validate a liquid-liquid extraction method, which allows a fairly reliable analysis of pesticides using gas chromatography-spectrometry mass （GC/MS） in toxicology laboratory at National Institute of hygiene, Rabat Morocco. The equipment required to perform these analyzes are the biological matrices （blood, gastric fluid）, in which the authors have doped the Organophosphorus pesticides such as Chlorpyrifos, Dichlorvos and Organochlorine pesticide： dichlorodiphenyltrichloroethane （DDT） and Heptachlor. After extracting the mixture with toluene, the supernatant was collected after centrifugation and concentrated in a small volume of 1.5 mL and then analyzed in GC/MS. After analyzing, the authors found that the yields of each pesticide in samples are significant; respectively they represented 73.4% of Chlorpyrifos, 70.8% of Dichlorvos, 47.8% of DDT and 71.6% of Heptachlor. The blood has a strong link with the most pesticides, where it＇s important to use the GC/MS to identify these products. The extraction with toluene was effective, especially to OP, but it＇s also sensitive to OC.