Quality-by-design-based development and validation of a stability-indicating UPLC method for quantification of teriflunomide in the presence of degradation products and its application to invitro dissolution
A systematic design-of-experiments was performed by applying quality-by-design concepts to determine
design space for rapid quantification of teriflunomide by the ultraperformance liquid chromatography
(UPLC) method in the presence of degradation products. Response surface and central composite
quadratic were used for statistical evaluation of experimental data using a Design-Expert software. The
response variables such as resolution, retention time, and peak tailing were analyzed statistically for the
screening of suitable chromatographic conditions. During this process, various plots such as perturbation,
contour, 3D, and design space were studied. The method was developed through UPLC BEH C18
2.1 � 100 mm, 1.7-μ column, mobile phase comprised of buffer (5 mM K2HPO4 containing 0.1%
triethylamine, pH 6.8), and acetonitrile (40:60 v/v), the flow rate of 0.5 mL min 1 and UV detection at
250 nm. The method was developed with a short run time of 1 min. Forced degradation studies revealed
that the method was stability-indicating, suitable for both assay and in-vitro dissolution of a drug product.
The method was found to be linear in the range of 28–84 μg mL 1, 2.8–22.7 μg mL 1 with a correlation
coefficient of 0.9999 and 1.000 for assay and dissolution, respectively. The recovery values were found in
the range of 100.1–101.7%. The method was validated according to ICH guidelines.
Stability indicating method development and validation for the simultaneous e...pharmaindexing
Stability indicating method development and validation for the simultaneous estimation of rabeprazole sodium and ketorolac tromethamine in bulk and synthetic mixture by RP-HPLC
Development and validation of a stability-indicating HPLC method for the simultaneous determination of Losartan potassium, hydrochlorothiazide, and their degradationproducts
Development and validation of GC-MS method for analysis of chloropyramine hyd...iosrphr_editor
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
Stability indicating method development and validation for the simultaneous e...pharmaindexing
Stability indicating method development and validation for the simultaneous estimation of rabeprazole sodium and ketorolac tromethamine in bulk and synthetic mixture by RP-HPLC
Development and validation of a stability-indicating HPLC method for the simultaneous determination of Losartan potassium, hydrochlorothiazide, and their degradationproducts
Development and validation of GC-MS method for analysis of chloropyramine hyd...iosrphr_editor
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
Bioanalytical RP-HPLC Method Development and Validation for Estimation of Cur...Sagar Savale
The present study was aimed at developing a reversed phase high performance liquid chromatography (RPHPLC) method for determination of curcumin (CRM) in plasma and hydrochlorothiazide was used as an internal
standard. The separation was achieved by using C-18 column (Qualisil BDS C18, 250 mm x 4.6 mm I.D.)
coupled with a guard column of silica, mobile phase was consisting of acetonitrile: water with 0.1% formic acid
(40:60 v/v). The flow rate was 0.3 ml/min and the drug was detected using PDA detector at the wavelength of 423
nm. The experimental conditions, including the diluting solvent, mobile phase composition, column saturation
and flow rate, were optimised to provide high-resolution and reproducible peaks. The developed method was
validated in terms of linearity, recovery, precision, ruggedness, sensitivity (LOD and LOQ) and stability study
(short and long-term stabilities, Freeze/thaw stability and post-preparative).
Stability indicating analytical method development and validation for estimat...SriramNagarajan18
Stability indicating analytical method development and validation for estimation of Sacubitril and Valsartan in bulk and pharmaceutical dosage form using RP-HPLC
Practical Implementation of the New Elemental Impurities Guidelines May 2015SGS
The International Conference on Harmonization (ICH) released its Q3D Guideline for Elemental Impurities in December 2014, initiating reviews and changes in quality testing programs in bio/pharmaceutical companies around the world. In advance of the implementation dates, companies need to assess the risks of potential elemental impurities in their process and materials streams.
In this presentation, experts will review the requirements of elemental impurities guidelines from ICH, the European Pharmacopeia, and United States Pharmacopeia, outline practical recommendations to address implementation challenges, and discuss key considerations for analytical testing programs.
The drug or drug combination may not be official in any pharmacopoeias.
A proper analytical procedure for the drug may not be available in the literature due to patent regulations.
Analytical methods may not be available for the drug in the form of a formulation due to the interference caused by the formulation excipients.
Analytical methods for the quantitation of the drug in biological fluids may not be available.
Analytical methods for a drug in combination with other drugs may not be available.
The existing analytical procedures may require expensive reagents and solvents. It may also involve cumbersome extraction and separation procedures and these may not be reliable.
An emerging way of estimation of Olmesartan medoxomil & Hydrochlorothiazide i...Priyanka Bose
An emerging way of estimation of Olmesartan medoxomil & Hydrochlorothiazide in Bulk & Combined tablet dosage form by Absorbance Correction Method using methanol as solvent system. It is cost effective, highly precised method for estimation.
Determination of Etodolac in Commercial Formulations by HPLC-UV Methodijtsrd
The aim of this study was to develop and verify a simple, rapid and sensitive high performance liquid chromatography method coupled with UV detector HPLC UV method for the quantitative determination of etodolac in bulk and pharmaceutical dosage forms. Chromatographic separation was performed at ambient conditions on a reverse phase ACE C8 analytical column 250 mm x 4.6 mm ID, 5 umm using the mobile phase containing acetonitrile water 80 20, v v at a flow rate of 1.0 mL min 1. A wavelength of 272 nm was used for etodolok and paracetamol IS . A retention time of 4.21 min and 2.02 min were obtained for etodolac and IS, respectively. The method showed linearity in the range of 0.08 10 µg mL 1 for etodolac R = 0.9999 . The linear regression equations obtained by least square regression method were the ratio of peak area of etodolac and IS =1.559 concentration etodolac µg mL 0.139. The intra day and inter day RE and RSD values of the method were =10.0 and =2.65 , respectively. Limit of detection LOD and limit of quantification LOQ were found to be 0.04 and 0.06 µg mL 1 for etodolac, respectively. A new, simple and sensitive high performance liquid chromatography method was developed and validated for etodolac. The method can be applied for the quantification of etodolac without derivatization in bulk solutions and commercial formulations using the internal standard. Tugrul Cagri Akman | Yucel Kadioglu "Determination of Etodolac in Commercial Formulations by HPLC-UV Method" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-4 | Issue-1 , December 2019, URL: https://www.ijtsrd.com/papers/ijtsrd29452.pdfPaper URL: https://www.ijtsrd.com/pharmacy/analytical-chemistry/29452/determination-of-etodolac-in-commercial-formulations-by-hplc-uv-method/tugrul-cagri-akman
Bioanalytical RP-HPLC Method Development and Validation for Estimation of Cur...Sagar Savale
The present study was aimed at developing a reversed phase high performance liquid chromatography (RPHPLC) method for determination of curcumin (CRM) in plasma and hydrochlorothiazide was used as an internal
standard. The separation was achieved by using C-18 column (Qualisil BDS C18, 250 mm x 4.6 mm I.D.)
coupled with a guard column of silica, mobile phase was consisting of acetonitrile: water with 0.1% formic acid
(40:60 v/v). The flow rate was 0.3 ml/min and the drug was detected using PDA detector at the wavelength of 423
nm. The experimental conditions, including the diluting solvent, mobile phase composition, column saturation
and flow rate, were optimised to provide high-resolution and reproducible peaks. The developed method was
validated in terms of linearity, recovery, precision, ruggedness, sensitivity (LOD and LOQ) and stability study
(short and long-term stabilities, Freeze/thaw stability and post-preparative).
Stability indicating analytical method development and validation for estimat...SriramNagarajan18
Stability indicating analytical method development and validation for estimation of Sacubitril and Valsartan in bulk and pharmaceutical dosage form using RP-HPLC
Practical Implementation of the New Elemental Impurities Guidelines May 2015SGS
The International Conference on Harmonization (ICH) released its Q3D Guideline for Elemental Impurities in December 2014, initiating reviews and changes in quality testing programs in bio/pharmaceutical companies around the world. In advance of the implementation dates, companies need to assess the risks of potential elemental impurities in their process and materials streams.
In this presentation, experts will review the requirements of elemental impurities guidelines from ICH, the European Pharmacopeia, and United States Pharmacopeia, outline practical recommendations to address implementation challenges, and discuss key considerations for analytical testing programs.
The drug or drug combination may not be official in any pharmacopoeias.
A proper analytical procedure for the drug may not be available in the literature due to patent regulations.
Analytical methods may not be available for the drug in the form of a formulation due to the interference caused by the formulation excipients.
Analytical methods for the quantitation of the drug in biological fluids may not be available.
Analytical methods for a drug in combination with other drugs may not be available.
The existing analytical procedures may require expensive reagents and solvents. It may also involve cumbersome extraction and separation procedures and these may not be reliable.
An emerging way of estimation of Olmesartan medoxomil & Hydrochlorothiazide i...Priyanka Bose
An emerging way of estimation of Olmesartan medoxomil & Hydrochlorothiazide in Bulk & Combined tablet dosage form by Absorbance Correction Method using methanol as solvent system. It is cost effective, highly precised method for estimation.
An emerging way of estimation of Olmesartan medoxomil & Hydrochlorothiazide i...
Similar to Quality-by-design-based development and validation of a stability-indicating UPLC method for quantification of teriflunomide in the presence of degradation products and its application to invitro dissolution
Determination of Etodolac in Commercial Formulations by HPLC-UV Methodijtsrd
The aim of this study was to develop and verify a simple, rapid and sensitive high performance liquid chromatography method coupled with UV detector HPLC UV method for the quantitative determination of etodolac in bulk and pharmaceutical dosage forms. Chromatographic separation was performed at ambient conditions on a reverse phase ACE C8 analytical column 250 mm x 4.6 mm ID, 5 umm using the mobile phase containing acetonitrile water 80 20, v v at a flow rate of 1.0 mL min 1. A wavelength of 272 nm was used for etodolok and paracetamol IS . A retention time of 4.21 min and 2.02 min were obtained for etodolac and IS, respectively. The method showed linearity in the range of 0.08 10 µg mL 1 for etodolac R = 0.9999 . The linear regression equations obtained by least square regression method were the ratio of peak area of etodolac and IS =1.559 concentration etodolac µg mL 0.139. The intra day and inter day RE and RSD values of the method were =10.0 and =2.65 , respectively. Limit of detection LOD and limit of quantification LOQ were found to be 0.04 and 0.06 µg mL 1 for etodolac, respectively. A new, simple and sensitive high performance liquid chromatography method was developed and validated for etodolac. The method can be applied for the quantification of etodolac without derivatization in bulk solutions and commercial formulations using the internal standard. Tugrul Cagri Akman | Yucel Kadioglu "Determination of Etodolac in Commercial Formulations by HPLC-UV Method" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-4 | Issue-1 , December 2019, URL: https://www.ijtsrd.com/papers/ijtsrd29452.pdfPaper URL: https://www.ijtsrd.com/pharmacy/analytical-chemistry/29452/determination-of-etodolac-in-commercial-formulations-by-hplc-uv-method/tugrul-cagri-akman
IOSR Journal of Pharmacy (IOSRPHR), www.iosrphr.org, call for paper, research...iosrphr_editor
IOSR Journal of Pharmacy (IOSRPHR), www.iosrphr.org, call for paper, research paper publishing, where to publish research paper, journal publishing, how to publish research paper, Call for research paper, international journal, publishing a paper, call for paper 2012, journal of pharmacy, how to get a research paper published, publishing a paper, publishing of journal, research and review articles, Pharmacy journal, International Journal of Pharmacy, hard copy of journal, hard copy of certificates, online Submission, where to publish research paper, journal publishing, international journal, publishing a paper
Development and validation of UPLC method for simultaneous quantification of ...Ratnakaram Venkata Nadh
A methodical design-of-experiments were performed by applying quality-by-design concepts to establish
a design-space for simultaneous and rapid quantification of Carvedilol and Ivabradine by UPLC in the
presence of degradation products. Response-surface, central-composite design, and quadratic model
were employed for statistical assessment of experimental data using the Design-Expert software.
Response variables such as resolution and retention time were analyzed statistically for chromatographic
screening. During DoE study, various plots such as perturbation, contour, 3D and design-space plots were
considered for method optimization. The method was developed using C8 [100 � 2.1 mm, 1.8 μ] UPLC
column, mobile phase comprising 0.5% triethylamine buffer [pH 6.4] and acetonitrile in the ratio of 50:50
v/v, the flow rate of 0.4 mL minute−1 and UV detection at 285 nm for both Carvedilol and Ivabradine.
The method was developed with a short run time of two minutes. The method was found to be linear in
the range of 25.0–199.9 μg mL−1 and 8.9–21.3 μg mL−1 for Carvedilol and Ivabradine, respectively with a
correlation coefficient of 0.9998 in each case. The recovery values were found in the range of 99.7–100.8%
and 98.9–100.9% for Carvedilol and Ivabradine, respectively. The method was validated according to ICH
Q2 (R1) guidelines.
International Journal of Pharmaceutical Science Invention (IJPSI) is an international journal intended for professionals and researchers in all fields of Pahrmaceutical Science. IJPSI publishes research articles and reviews within the whole field Pharmacy and Pharmaceutical Science, new teaching methods, assessment, validation and the impact of new technologies and it will continue to provide information on the latest trends and developments in this ever-expanding subject. The publications of papers are selected through double peer reviewed to ensure originality, relevance, and readability. The articles published in our journal can be accessed online.
Analytical Method Development and Validation for the Estimation of Zolmitript...ijtsrd
In this work the authors have proposed a simple, specific, economic and accurate reverse phase liquid chromatographic method for the estimation of Zolmitriptan as an active pharmaceutical ingredient and in pharmaceutical formulation. The main objective of the current research paper is to To develop simple, precise and accurate RP HPLC method for Zolmitriptan also to validate the developed method as per ICH guideline Q2R1 and to explore the applicability of the method in finished product formulation for estimation of Zolmitriptan during its lifecycle. The objective was achieved by optimized condition with Phonemenex C18 column 150mm×4.6mm , 5µm. And mobile phase Phosphate buffer pH 3.5 85 Methanol 15. The separation was done with a flow rate of 0.9ml min, detection with 224nm. The retention was found to be 3.57 minute. LOD and LOQ were found to be 2.45 and 7.42 respectively. So in order to obtain the correct results various validations methods are performed to get the results. The results obtained from those validation methods are plotted in the form of the charts as well as the different curves. Mr. Rahul M. Sagde | Mr. Pawan N. Karwa | Mr. Vivek M. Thorat | Sanjay S. Jadhav "Analytical Method Development and Validation for the Estimation of Zolmitriptan by RP HPLC Method" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-3 | Issue-5 , August 2019, URL: https://www.ijtsrd.com/papers/ijtsrd26474.pdfPaper URL: https://www.ijtsrd.com/pharmacy/analytical-chemistry/26474/analytical-method-development-and-validation-for-the-estimation-of-zolmitriptan-by-rp-hplc-method/mr-rahul-m-sagde
International Journal of Engineering Research and Applications (IJERA) is an open access online peer reviewed international journal that publishes research and review articles in the fields of Computer Science, Neural Networks, Electrical Engineering, Software Engineering, Information Technology, Mechanical Engineering, Chemical Engineering, Plastic Engineering, Food Technology, Textile Engineering, Nano Technology & science, Power Electronics, Electronics & Communication Engineering, Computational mathematics, Image processing, Civil Engineering, Structural Engineering, Environmental Engineering, VLSI Testing & Low Power VLSI Design etc.
RP-HPLC Method Developed for the Estimation of Etodolac and Thiocolchicoside ...ijtsrd
A new, simple, specific, sensitive, rapid, accurate and precise RP HPLC method was developed for the estimation of Etodolac ETO and Thiocolchicoside THIO in bulk and combined pharmaceutical dosage forms. Etodolac ETO and Thiocolchicoside THIO was chromatographed on a Symmetry ODS C18 4.6mm—250mm, 5µm Column in a mobile phase consisting of Acetonitrile Methanol Acetate Buffer 25 20 55 v v . The mobile phase was pumped at a flow rate of 1.0 ml min with detection at 238 nm. The detector response was linear in the concentration of 200 600µg ml 20 60µg ml for Etodolac and Thiocolchicoside respectively. The intra and inter day variation was found to be less than 2 . The mean recovery of the drug from the solution was 99.98 . The developed method was validated for sensitivity, accuracy, precision, ruggedness and robustness. The RSD results for precision and intermediate precision found less than 2.0 . The LOD and LOQ were found to be 0.86 and 1.18 µg ml for Etodolac and 2.4 and 3.54µg ml for Thiocolchicoside respectively. The proposed method is simple, fast, accurate, precise and reproducible hence it can be applied for routine quality control analysis of Etodolac ETO and Thiocolchicoside THIO in bulk and combined pharmaceutical formulations. Prapulla Putta "RP-HPLC Method Developed for the Estimation of Etodolac and Thiocolchicoside in Bulk and Combined Dosage Forms" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-4 | Issue-1 , December 2019, URL: https://www.ijtsrd.com/papers/ijtsrd29703.pdf Paper URL: https://www.ijtsrd.com/pharmacy/analytical-chemistry/29703/rp-hplc-method-developed-for-the-estimation-of-etodolac-and-thiocolchicoside-in-bulk-and-combined-dosage-forms/prapulla-putta
Development and validation of rphplc method for nsai ds in combined pharmaceu...IJSIT Editor
The present research work includes the development and validation of High performance liquid
chromatography method for simultaneous estimation of Etirocoxib (ETX )and Paracetamol (PCM) which are
very much of useful in multiple therapies rather than the use of single drug formulation, because of multiple
actions, fever side effect ,and quicker relief. We aimed to design a quick and simple method, suitable for the
determination of identity, strength. The UV spectroscopic method were developed on standard ETX and PCM
using methanol as standard solution and also, the standard stock solution of both ETX and PCM were diluted
with 2%hydrochloric acid to obtain concentration 2.0µg/ml and 16.6µg/ml.The spectrum was recorded in the
range of 400-200nm.The RP- HPLC method were developed on standard ETX and PCM which performed on C-
18 column using mobile phase composed by acetonitrile :water 50:50(v/v) at a flow rate of 1 ml/min, with
injection volume 20µl and Ph 6.8 can be adjusted using orthophosphoric acid.
Validation of analytical method for Diabetes Mellitus DhruvkumarPatel25
Development and validation of analytical method
for simultaneous estimation of Teneligliptin hydrobromide hydrate and Metformin
hydrochloride in combined dosage form.
Sagar kanade m pharm indrustial projectSagar Kanade
DEVELOPMENT AND VALIDATION OF STABILITY INDICATING ANALYTICAL METHOD FOR THE ESTIMATION OF MIDODRINE HYDROCHLORIDE ORAL SOLUTION BY RP HPLC METHOD
Similar to Quality-by-design-based development and validation of a stability-indicating UPLC method for quantification of teriflunomide in the presence of degradation products and its application to invitro dissolution (20)
Electrochemical study of anatase TiO2 in aqueous sodium-ion electrolytesRatnakaram Venkata Nadh
In this paper, a basic electro-analytical study on the behavior of anatase TiO2 in aqueous NaOH has been presented using cyclic voltammetry technique (CV). The study has explored the possibility of using TiO2 as anode material for ARSBs in presence of 5 M NaOH aqueous electrolyte. CV profiles show that anatase TiO2 exhibits reversible sodium ion insertion/de-insertion reactions. CV studies of TiO2 anode in aqueous sodium electrolytes at different scan rate shows that the Na+ ion insertion reaction at the electrode is diffusion controlled with a resistive behavior. Proton insertion from aqueous sodium electrolytes into TiO2 cannot be ruled out. To confirm the ion inserted and de-inserted, CV studies are done at different concentration of NaOH and it is found that at lower concentrations of NaOH, proton insertion process competes with Na+ ion insertion process and as the concentration increases, the Na+ ion insertion process becomes the predominant electrode reaction making it suitable anode materials for aqueous sodium batteries in 5 M NaOH.
Validated HPLC Method for Assay and Content Uniformity Testing of Roflumilast...Ratnakaram Venkata Nadh
Roflumilast is a selective enzyme inhibitor of phosphodiesterase-4. This drug is recommended for treatment of patients suffering from
chronic-obstructive-pulmonary-disease with chronic-bronchitis. Roflumilast is not official in pharmacopoeia and the reported methods
are having high chromatographic run times. A short run time HPLC method was developed for assay and content uniformity testing to
determine the roflumilast in blend and tablets. The mobile phase consists of 10 mM sodium dihydrogen phosphate monohydrate buffer
and acetonitrile in the ratio of 45:55 v/v. The HPLC method was developed using accucore-C18 150 × 4.6 mm, 4 μm column with a flow
rate of 1.0 mL min-1, 215 nm wavelength and 10 μL injection volume with run time of 5 min. The method linearity was proved between
5.02-40.17 μg mL-1 and obtained correlation-coefficient value is 1.0000. The mean recovery of roflumilast was 100.6%. The stability
indicating nature was established and performed the validation by considering ICH Q2 (R1) recommendations.
Substrate Inhibition in Ruthenium(III) Catalyzed Oxidation of Propane-1,3-dio...Ratnakaram Venkata Nadh
Ruthenium(III) catalyzed oxidation of propane-1,3-diol by potassium periodate was studied in aqueous perchloric acid medium. Orders
of reaction with respect to concentrations of oxidant, substrate, acid and catalyst were determined. First order in oxidant and catalyst
concentrations, and inverse fractional order in acid medium were observed. In addition, substrate inhibition (i.e. a decrease in reaction rate
with an increase in substrate concentration) was observed. Effect of addition of salt and solvent was studied. Based on the studies of
temperature variation, Arrhenius parameters were calculated. Plausible mechanism was also proposed based on observed kinetics.
Ruthenium(III) Catalyzed Oxidation of Sugar Alcohols by Dichloroisocyanuric A...Ratnakaram Venkata Nadh
Kinetics of ruthenium(III) catalyzed oxidation of biologically important sugar alcohols (myo-inositol,
D-sorbitol, and D-mannitol) by dichloroisocyanuric acid was carried out in aqueous acetic acid—perchloric
medium. The reactions were found to be first order in case of oxidant and ruthenium(III). Zero order
was observed with the concentrations of sorbitol and mannitol whereas, a positive fractional order was found
in the case of inositol concentration. An inverse fractional order was observed with perchloric acid in oxidation
of three substrates. Arrhenius parameters were calculated and a plausible mechanism was proposed
Shift of Reaction Pathway by Added Chloride Ions in the Oxidation of Aromatic...Ratnakaram Venkata Nadh
Role of added chloride ions on the shift of reaction pathway of oxidation of aromatic ketones (acetophenone,
desoxybenzoin) by dichloroisocyanuric acid (DCICA) was studied in aqueous acetic acid—perchloric
acid medium. Participation of enolic and protonated forms of ketones in the rate determining steps is
manifested from zero and first orders with respect to the oxidant in absence and presence of added chloride
ions, respectively. Positive and negative effects of acid and dielectric constant on the reaction rate were
observed. The observations deduce plausible mechanisms involving (i) rate-determining formation of enol
from the conjugate acid of the ketone (SH+) in the absence of added chloride ions and (ii) rapid formation of
molecular chlorine species from HOCl (hydrolytic species of DCICA) in the presence of added chloride ions,
which then interacts with SH+ in a rate-determining step prior to the rapid steps of product formation. The
order of Arrhenius parameters substantiate the proposed plausible mechanisms based on order of reactants
both in presence and absence of added chloride ions.
Kinetics of Ruthenium(III) Catalyzed and Uncatalyzed Oxidation of Monoethanol...Ratnakaram Venkata Nadh
Kinetics of uncatalyzed and ruthenium(III) catalyzed oxidation of monoethanolamine by N-bromosuccinimide
(NBS) has been studied in an aqueous acetic acid medium in the presence of sodium acetate
and perchloric acid, respectively. In the uncatalyzed oxidation the kinetic orders are: the first order in NBS,
a fractional order in the substrate. The rate of the reaction increased with an increase in the sodium acetate
concentration and decreased with an increase in the perchloric acid concentration. This indicates that free
amine molecules are the reactive species. Addition of halide ions results in a decrease in the kinetic rate,
which is noteworthy. Both in absence and presence of a catalyst, a decrease in the dielectric constant of the
medium decreases the kinetic rate pointing out that these are dipole—dipole reactions. A relatively higher
oxidation state of ruthenium i.e., Ru(V) was found to be the active species in Ru(III) catalyzed reactions. A
suitable mechanism consistent with the observations has been proposed and a rate law has been derived to
explain the kinetic orders.
A novel reversed-phase liquid chromatographic method for the simultaneous det...Ratnakaram Venkata Nadh
In the present study 12 impurities of bisoprolol fumarate (BISO) and hydrochlorothiazide (HCTZ) were
separated simultaneously in a single HPLC method. Out of these 12 impurities, five are potential
degradants, which are validated as per The International Council for Harmonisation of Technical
Requirements for Pharmaceuticals for Human Use (ICH) guidelines. As the two active drug substances
BISO and HCTZ have different solubilities and polarities, the most critical parameters in resolving the
components from each other are pH, temperature, and solvents. The method is precise (RSD < 1.0%),
accurate, linear (r2 > 0.999), robust, and stability indicating in the range of limit of quantification (LOQ)
to 150%. The HPLC method is then migrated to ultra-performance liquid chromatography (UPLC) to
further reduce the run time and solvent consumption, and increase the sample throughput
The emergence of multidrug-resistant TB (MDR-TB) against first-line drugs and extensively drug resistant TB (XDRTB)
due to misuse of second-line anti tubercular drugs (ATDs) is a further concern. Recommended treatment involves
long term and multiple drug therapy with severe side effects. Due to this concern nanoparticle-based systems
have significant potential for treatment and prevention of tuberculosis (TB) to overcome the need to administer
ATDs at high and frequent doses, would assist in improving patient compliance and circumvent hepatotoxic ity
and/or nephrotoxicity/ocular toxicity/ototoxicity associated with the prevalent first-line chemotherapy.
Nanostructured delivery systems constitute a wide range of systems varying from liposomes, micelles, micro- and
nanoemulsions, to polymeric nanoparticles (PNPs ) and solid lipid nanoparticles (SLNs). Pulmonary administration
of inhaled nanoparticles in the form of dry powder inhalers offer particular advantages for pulmonary administration
of anti tubercular drugs (ATDs). Present review comprehensively about different approaches of nanobased
drug delivery, devises and techniques for pulmonary delivery of nanoparticle encapsulated ATD.
Kinetic, isotherm and thermodynamics investigation on adsorption of divalent ...Ratnakaram Venkata Nadh
Three novel and distinct agricultural waste materials, viz., Casuarinas fruit powder (CFP), sorghum stem powder
(SSP) and banana stem powder (BSP) were used as low cost adsorbents for the removal of toxic copper(II) from
aqueous solutions. Acid treated adsorbents were characterized by SEM, EDX and FTIR. Different factors effecting
adsorption capacity were analyzed and the effi ciency order was BSP>SSP>CFP. Based on the extent of compatibility
to Freundlich/Langmuir/D-R/Temkin adsorption isotherm and different models (pseudo-fi rst and second order,
Boyd, Weber’s and Elovich), chemisorption primarily involved in the case of CFP and SSP, whereas, simultaneous
occurrence of chemisorption and physisorption was proposed in the case of BSP. Based on the observations, it was
proposed that three kinetic stages involve in adsorption process viz., diffusion of sorbate to sorbent, intra particle
diffusion and then establishment of equilibrium. These adsorbents have promising role towards removal of Cu(II)
from industrial wastewater to contribute environmental protection.
Kinetic, thermodynamic and equilibrium studies on removal of hexavalent chrom...Ratnakaram Venkata Nadh
Removal of Cr(VI) by biosorption on two agro waste materials, casuarinas fruit powder (CFP) and sorghum
stem powder (SSP), has been investigated. The prepared adsorbent materials were characterized by SEM, EDX,
FTIR and BET. These biomaterials effectively removed Cr(VI) with a maximum removal of 93.35% and 63.75% using
15 gL−1 and 5 gL−1 of CFP and SSP, respectively, at 60 oC with 20mgL−1 initial Cr(VI) concentration in solution. In both
cases of adsorbents, kinetic data of adsorption fitted well in pseudo-second-order in terms of correlation coefficient
(R2). This helps in proposing the process of adsorption as chemical coordination, which is correlated with the thermodynamic
study results conducted at different values of temperature. Langmuir, Freundlich and D-R models were evaluated
for description of metal sorption isotherms. Values of coefficients of intra-particle diffusion and mass transfer have
also been determined at different values of temperature.
Novel coumarin isoxazoline derivatives: Synthesis and study of antibacterial ...Ratnakaram Venkata Nadh
A highly efficient and mild protocol for the syntheses of ethyl-3-
[7-benzyloxy-4-methyl-2-oxo-2H-8-chromenyl]-5-aryl-4,5-dihydro-4-
isoxazole carboxylates and ethyl-3-[7-benzyloxy-3-chloro-4-methyl-2-
oxo-2H-8-chromenyl]-5-aryl-4,5-dihydro-4-isoxazole carboxylates in
good yields via [3 þ 2] cycloaddition of in situ–generated nitrile
oxides from 7-benzyloxy-4-methyl-coumarin hydroxymoylchlorides
and 7-benzyloxy-3-chloro-4-methyl-coumarin hydroxymoylchlorides
respectively with ethyl-3-aryl prop-2-enoate has been developed.
The new compounds are screened for antibacterial activity.
Ultra performance liquid chromatographic method for simultaneous quantificati...Ratnakaram Venkata Nadh
Plerixafor (PLX) injections are administered to patients with cancers of lymphocytes
(non-Hodgkin’s lymphoma) and plasma cells (multiple myeloma). The main
objective of the current study was to develop a short reverse phase chromatographic
method for the simultaneous quantification of PLX and its impurities, in an injection
formulation, to reduce the time required for these quality tests. Furthermore, the
present work describes the role of nonalkyl branched nonquaternary ion pair reagent
in improving the peak shape and reducing column equilibration time. The separation
of PLX and its related substances is pH dependent (optimum pH = 2.50) and was
achieved on an octadecylsilyl (C18) column. The method was validated for its intended
purpose in accordance with the current regulatory guidelines for validation. The
proposed method can be applied for quality control, release, and stability analyses of
active pharmaceutical ingredient, PLX, as well as finished products, PLX injections
Caralluma lasiantha: A review on it’s vital role in Indian Traditional MedicineRatnakaram Venkata Nadh
Caralluma is a genus used as traditional medicine. Caralluma lasiantha is medicinally important due
to existence of pregnane glycosides, which may possess various biological activities. This article thoroughly
reviewed about the usage of C. lasiantha in traditional medicinal system, phytochemicals present in it, profile
identification studies, anti-hyperglycemic effect, antibacterial, antifungal, cytotoxic and antioxidant activities
Phytochemical Investigation of Caralluma lasiantha: Isolation of Stigmasterol...Ratnakaram Venkata Nadh
Stigmasterol, a phytosteroid was isolated for the first time from C. lasiantha using n-hexane as a solvent. Stigmasterol was characterized on the basis of physical, chemical and spectral data (IR, 1H NMR, 13C NMR, 1HNMR, DEPT-45, 90 & 135, and MS) analysis as well as by comparing them to their literature data. A sequence of steps was adopted like saponification, fractional crystallization and gradient elution column chromatography to isolate stigmasterol because some phytosterols possess identical physical properties which makes it difficult to isolate the constituents.
Phytochemical Screening of Caralluma lasiantha Isolation of C21 Pregnane SteroidRatnakaram Venkata Nadh
Phytochemical screening of Caralluma lasiantha was carried out and one C21 pregnane steroid was isolated from chloroform extract. Based on spectroscopic studies (IR, 1H NMR, 13C NMR and ESI-MS) the isolated compound is 3b,14b-dihydroxy-14b-pregn-5-en-20-one which was earlier isolated from other species.
Supercritical fluid (CO2) chromatography for quantitative determination of se...Ratnakaram Venkata Nadh
In the present study, two cancer therapeutic drugs (docetaxel and bortezomib) were separated from their
potential impurities on a chromatographic platform by utilizing CO2 gas (supercritical state) and quantified.
The chromatographic separations were achieved on two short columns BEH-2EP (100mm 3mm, 1.7 mm)
and CHIRALPAK AD-3 (100 mm 4.6 mm, 3 mm) for docetaxel and bortezomib, respectively. The present
work describes the role of organic modifiers in the separation of polar compounds by supercritical fluid
chromatography. The two new methods were fully validated in accordance with the current ICH
(International Council for Harmonization of technical requirements for pharmaceuticals for human use)
guidelines. The stability indicating power of the methods was demonstrated from the stress studies
conducted on the injection formulations of the two compounds. The methods are precise with % RSD of
0.4, linear with the correlation coefficient of r2 $ 0.999 and accurate in the range of 50–150% of the
target assay concentration. The two methods can be equally employed for the assay determination of
docetaxel and bortezomib APIs as well.
A convenient new and efficient commercial synthetic route for dasatinib (Spry...Ratnakaram Venkata Nadh
A new and efficient synthetic route for dual-Src/Abl kinase inhibitor
dasatinib (Sprycel®), an anticancer drug, is described. This commercially
viable process yields dasatinib monohydrate free of potential impurities
with consistent yield of 68% in route A and 61% in route B with HPLC
purity >99.80% over four stages.
Utilization of agro-waste for removal of toxic hexavalent chromium: surface i...Ratnakaram Venkata Nadh
Abundantly available agricultural waste materials
(banana bunch, sorghum stem and casuarinas fruit) are
processed with negligible cost and are found to be highly
suitable as biosorbents for chromium(VI) removal from
aqueous environment due to high surface area and functional
groups of adsorbents. The equilibrium data have
been analyzed for the adsorbate–adsorbate/adsorbent
interactions and found to be fitted to the data in the order,
Hill–de Boer C Fowler–Guggenheim % Frumkin[Kiselev.
To determine the characteristic parameters for process
design, mass transfer studies have been carried out using
two-parameter isotherm models (Harkins–Jura, Halsey,
Smith, El-Awady and Flory–Huggins) and three-parameter
isotherm models (Redlich–Peterson and Sips) which are
applied to the experimental data. The fitness of the isotherms
describes that both mono- and multilayer adsorptions
occur in the present studied three biosorbents in
preference to the latter. The mechanism of adsorption has
been studied using diffusion kinetic models (viz. liquid film
diffusion, Dunwald–Wagner intra-particle diffusion model
and moving boundary model) and described the possibility
of diffusion in the order of banana bunch–stem powder[
sorghum stem powder[casuarinas fruit powder in
terms of diffusion coefficients. In essence of all the results,
the selected adsorbents can be used as a potential adsorbent
for the removal of Cr(VI) from aqueous solutions.
Evaluation of in vitro antibacterial activity of Caralluma lasiantha for scie...Ratnakaram Venkata Nadh
Caralluma lasiantha is used as a traditional medicine in India to heal body
heat and inflammations. In order to find out a scientific validation for the Indian
traditional knowledge, antibacterial activity of C. lasiantha extracts was studied
against inflammation causing bacteria (viz., Staphylococcus aureus, Escherichia coli,
Streptococcus Sp., Bacillus subtilis, Enterobacter aerogenes, Klebsiella pneumoniae)
along with other Gram-positive and Gram-negative bacteria. Solvents with different
polarity were used for extraction from dry roots and stems. Minimum inhibitory
concentrations (MIC) were also studied. Differential antibacterial activity was
exhibited by extracts and higher inhibition potential against Gram-positive bacteria
was explained. The observed antibacterial activities were correlated with the chemical
structures of phytochemicals present in C. lasiantha. Anti-inflammation activities
are related to C. lasiantha extracts through their antibacterial activities.
Novel Hybrid Molecules of Isoxazole Chalcone Derivatives: Synthesis and Study...Ratnakaram Venkata Nadh
medicine due to their significant role in the treatment of different health problems.
Methods: We have synthesized new series of isoxazole-chalcone conjugates (14a-m) by the
Claisen-Schmidt condensation of suitable substituted acetophenones with isoxazole aldehydes (12a-d).
In vitro cytotoxic activity of the synthesized compounds was studied against four different selected
human cancer cell lines by using sulforhodamine B (SRB) method.
Results: The adopted scheme resulted in good yields of new series of isoxazole-chalcone
conjugates (14a-m). Potent cytotoxic activity was observed for compounds -14a, 14b, 14e, 14i, 14j
and 14k against prostate DU-145 cancer cell line.
Conclusion: The observed potent cytotoxic activities were due to the presence of 3,4,5-
trimethoxyphenyl group.
ARTIFICIAL INTELLIGENCE IN HEALTHCARE.pdfAnujkumaranit
Artificial intelligence (AI) refers to the simulation of human intelligence processes by machines, especially computer systems. It encompasses tasks such as learning, reasoning, problem-solving, perception, and language understanding. AI technologies are revolutionizing various fields, from healthcare to finance, by enabling machines to perform tasks that typically require human intelligence.
New Directions in Targeted Therapeutic Approaches for Older Adults With Mantl...i3 Health
i3 Health is pleased to make the speaker slides from this activity available for use as a non-accredited self-study or teaching resource.
This slide deck presented by Dr. Kami Maddocks, Professor-Clinical in the Division of Hematology and
Associate Division Director for Ambulatory Operations
The Ohio State University Comprehensive Cancer Center, will provide insight into new directions in targeted therapeutic approaches for older adults with mantle cell lymphoma.
STATEMENT OF NEED
Mantle cell lymphoma (MCL) is a rare, aggressive B-cell non-Hodgkin lymphoma (NHL) accounting for 5% to 7% of all lymphomas. Its prognosis ranges from indolent disease that does not require treatment for years to very aggressive disease, which is associated with poor survival (Silkenstedt et al, 2021). Typically, MCL is diagnosed at advanced stage and in older patients who cannot tolerate intensive therapy (NCCN, 2022). Although recent advances have slightly increased remission rates, recurrence and relapse remain very common, leading to a median overall survival between 3 and 6 years (LLS, 2021). Though there are several effective options, progress is still needed towards establishing an accepted frontline approach for MCL (Castellino et al, 2022). Treatment selection and management of MCL are complicated by the heterogeneity of prognosis, advanced age and comorbidities of patients, and lack of an established standard approach for treatment, making it vital that clinicians be familiar with the latest research and advances in this area. In this activity chaired by Michael Wang, MD, Professor in the Department of Lymphoma & Myeloma at MD Anderson Cancer Center, expert faculty will discuss prognostic factors informing treatment, the promising results of recent trials in new therapeutic approaches, and the implications of treatment resistance in therapeutic selection for MCL.
Target Audience
Hematology/oncology fellows, attending faculty, and other health care professionals involved in the treatment of patients with mantle cell lymphoma (MCL).
Learning Objectives
1.) Identify clinical and biological prognostic factors that can guide treatment decision making for older adults with MCL
2.) Evaluate emerging data on targeted therapeutic approaches for treatment-naive and relapsed/refractory MCL and their applicability to older adults
3.) Assess mechanisms of resistance to targeted therapies for MCL and their implications for treatment selection
Tom Selleck Health: A Comprehensive Look at the Iconic Actor’s Wellness Journeygreendigital
Tom Selleck, an enduring figure in Hollywood. has captivated audiences for decades with his rugged charm, iconic moustache. and memorable roles in television and film. From his breakout role as Thomas Magnum in Magnum P.I. to his current portrayal of Frank Reagan in Blue Bloods. Selleck's career has spanned over 50 years. But beyond his professional achievements. fans have often been curious about Tom Selleck Health. especially as he has aged in the public eye.
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Introduction
Many have been interested in Tom Selleck health. not only because of his enduring presence on screen but also because of the challenges. and lifestyle choices he has faced and made over the years. This article delves into the various aspects of Tom Selleck health. exploring his fitness regimen, diet, mental health. and the challenges he has encountered as he ages. We'll look at how he maintains his well-being. the health issues he has faced, and his approach to ageing .
Early Life and Career
Childhood and Athletic Beginnings
Tom Selleck was born on January 29, 1945, in Detroit, Michigan, and grew up in Sherman Oaks, California. From an early age, he was involved in sports, particularly basketball. which played a significant role in his physical development. His athletic pursuits continued into college. where he attended the University of Southern California (USC) on a basketball scholarship. This early involvement in sports laid a strong foundation for his physical health and disciplined lifestyle.
Transition to Acting
Selleck's transition from an athlete to an actor came with its physical demands. His first significant role in "Magnum P.I." required him to perform various stunts and maintain a fit appearance. This role, which he played from 1980 to 1988. necessitated a rigorous fitness routine to meet the show's demands. setting the stage for his long-term commitment to health and wellness.
Fitness Regimen
Workout Routine
Tom Selleck health and fitness regimen has evolved. adapting to his changing roles and age. During his "Magnum, P.I." days. Selleck's workouts were intense and focused on building and maintaining muscle mass. His routine included weightlifting, cardiovascular exercises. and specific training for the stunts he performed on the show.
Selleck adjusted his fitness routine as he aged to suit his body's needs. Today, his workouts focus on maintaining flexibility, strength, and cardiovascular health. He incorporates low-impact exercises such as swimming, walking, and light weightlifting. This balanced approach helps him stay fit without putting undue strain on his joints and muscles.
Importance of Flexibility and Mobility
In recent years, Selleck has emphasized the importance of flexibility and mobility in his fitness regimen. Understanding the natural decline in muscle mass and joint flexibility with age. he includes stretching and yoga in his routine. These practices help prevent injuries, improve posture, and maintain mobilit
Ethanol (CH3CH2OH), or beverage alcohol, is a two-carbon alcohol
that is rapidly distributed in the body and brain. Ethanol alters many
neurochemical systems and has rewarding and addictive properties. It
is the oldest recreational drug and likely contributes to more morbidity,
mortality, and public health costs than all illicit drugs combined. The
5th edition of the Diagnostic and Statistical Manual of Mental Disorders
(DSM-5) integrates alcohol abuse and alcohol dependence into a single
disorder called alcohol use disorder (AUD), with mild, moderate,
and severe subclassifications (American Psychiatric Association, 2013).
In the DSM-5, all types of substance abuse and dependence have been
combined into a single substance use disorder (SUD) on a continuum
from mild to severe. A diagnosis of AUD requires that at least two of
the 11 DSM-5 behaviors be present within a 12-month period (mild
AUD: 2–3 criteria; moderate AUD: 4–5 criteria; severe AUD: 6–11 criteria).
The four main behavioral effects of AUD are impaired control over
drinking, negative social consequences, risky use, and altered physiological
effects (tolerance, withdrawal). This chapter presents an overview
of the prevalence and harmful consequences of AUD in the U.S.,
the systemic nature of the disease, neurocircuitry and stages of AUD,
comorbidities, fetal alcohol spectrum disorders, genetic risk factors, and
pharmacotherapies for AUD.
micro teaching on communication m.sc nursing.pdfAnurag Sharma
Microteaching is a unique model of practice teaching. It is a viable instrument for the. desired change in the teaching behavior or the behavior potential which, in specified types of real. classroom situations, tends to facilitate the achievement of specified types of objectives.
Anti ulcer drugs and their Advance pharmacology ||
Anti-ulcer drugs are medications used to prevent and treat ulcers in the stomach and upper part of the small intestine (duodenal ulcers). These ulcers are often caused by an imbalance between stomach acid and the mucosal lining, which protects the stomach lining.
||Scope: Overview of various classes of anti-ulcer drugs, their mechanisms of action, indications, side effects, and clinical considerations.
These simplified slides by Dr. Sidra Arshad present an overview of the non-respiratory functions of the respiratory tract.
Learning objectives:
1. Enlist the non-respiratory functions of the respiratory tract
2. Briefly explain how these functions are carried out
3. Discuss the significance of dead space
4. Differentiate between minute ventilation and alveolar ventilation
5. Describe the cough and sneeze reflexes
Study Resources:
1. Chapter 39, Guyton and Hall Textbook of Medical Physiology, 14th edition
2. Chapter 34, Ganong’s Review of Medical Physiology, 26th edition
3. Chapter 17, Human Physiology by Lauralee Sherwood, 9th edition
4. Non-respiratory functions of the lungs https://academic.oup.com/bjaed/article/13/3/98/278874
Report Back from SGO 2024: What’s the Latest in Cervical Cancer?bkling
Are you curious about what’s new in cervical cancer research or unsure what the findings mean? Join Dr. Emily Ko, a gynecologic oncologist at Penn Medicine, to learn about the latest updates from the Society of Gynecologic Oncology (SGO) 2024 Annual Meeting on Women’s Cancer. Dr. Ko will discuss what the research presented at the conference means for you and answer your questions about the new developments.
Lung Cancer: Artificial Intelligence, Synergetics, Complex System Analysis, S...Oleg Kshivets
RESULTS: Overall life span (LS) was 2252.1±1742.5 days and cumulative 5-year survival (5YS) reached 73.2%, 10 years – 64.8%, 20 years – 42.5%. 513 LCP lived more than 5 years (LS=3124.6±1525.6 days), 148 LCP – more than 10 years (LS=5054.4±1504.1 days).199 LCP died because of LC (LS=562.7±374.5 days). 5YS of LCP after bi/lobectomies was significantly superior in comparison with LCP after pneumonectomies (78.1% vs.63.7%, P=0.00001 by log-rank test). AT significantly improved 5YS (66.3% vs. 34.8%) (P=0.00000 by log-rank test) only for LCP with N1-2. Cox modeling displayed that 5YS of LCP significantly depended on: phase transition (PT) early-invasive LC in terms of synergetics, PT N0—N12, cell ratio factors (ratio between cancer cells- CC and blood cells subpopulations), G1-3, histology, glucose, AT, blood cell circuit, prothrombin index, heparin tolerance, recalcification time (P=0.000-0.038). Neural networks, genetic algorithm selection and bootstrap simulation revealed relationships between 5YS and PT early-invasive LC (rank=1), PT N0—N12 (rank=2), thrombocytes/CC (3), erythrocytes/CC (4), eosinophils/CC (5), healthy cells/CC (6), lymphocytes/CC (7), segmented neutrophils/CC (8), stick neutrophils/CC (9), monocytes/CC (10); leucocytes/CC (11). Correct prediction of 5YS was 100% by neural networks computing (area under ROC curve=1.0; error=0.0).
CONCLUSIONS: 5YS of LCP after radical procedures significantly depended on: 1) PT early-invasive cancer; 2) PT N0--N12; 3) cell ratio factors; 4) blood cell circuit; 5) biochemical factors; 6) hemostasis system; 7) AT; 8) LC characteristics; 9) LC cell dynamics; 10) surgery type: lobectomy/pneumonectomy; 11) anthropometric data. Optimal diagnosis and treatment strategies for LC are: 1) screening and early detection of LC; 2) availability of experienced thoracic surgeons because of complexity of radical procedures; 3) aggressive en block surgery and adequate lymph node dissection for completeness; 4) precise prediction; 5) adjuvant chemoimmunoradiotherapy for LCP with unfavorable prognosis.
TEST BANK for Operations Management, 14th Edition by William J. Stevenson, Ve...kevinkariuki227
TEST BANK for Operations Management, 14th Edition by William J. Stevenson, Verified Chapters 1 - 19, Complete Newest Version.pdf
TEST BANK for Operations Management, 14th Edition by William J. Stevenson, Verified Chapters 1 - 19, Complete Newest Version.pdf
Pulmonary Thromboembolism - etilogy, types, medical- Surgical and nursing man...VarunMahajani
Disruption of blood supply to lung alveoli due to blockage of one or more pulmonary blood vessels is called as Pulmonary thromboembolism. In this presentation we will discuss its causes, types and its management in depth.
Title: Sense of Smell
Presenter: Dr. Faiza, Assistant Professor of Physiology
Qualifications:
MBBS (Best Graduate, AIMC Lahore)
FCPS Physiology
ICMT, CHPE, DHPE (STMU)
MPH (GC University, Faisalabad)
MBA (Virtual University of Pakistan)
Learning Objectives:
Describe the primary categories of smells and the concept of odor blindness.
Explain the structure and location of the olfactory membrane and mucosa, including the types and roles of cells involved in olfaction.
Describe the pathway and mechanisms of olfactory signal transmission from the olfactory receptors to the brain.
Illustrate the biochemical cascade triggered by odorant binding to olfactory receptors, including the role of G-proteins and second messengers in generating an action potential.
Identify different types of olfactory disorders such as anosmia, hyposmia, hyperosmia, and dysosmia, including their potential causes.
Key Topics:
Olfactory Genes:
3% of the human genome accounts for olfactory genes.
400 genes for odorant receptors.
Olfactory Membrane:
Located in the superior part of the nasal cavity.
Medially: Folds downward along the superior septum.
Laterally: Folds over the superior turbinate and upper surface of the middle turbinate.
Total surface area: 5-10 square centimeters.
Olfactory Mucosa:
Olfactory Cells: Bipolar nerve cells derived from the CNS (100 million), with 4-25 olfactory cilia per cell.
Sustentacular Cells: Produce mucus and maintain ionic and molecular environment.
Basal Cells: Replace worn-out olfactory cells with an average lifespan of 1-2 months.
Bowman’s Gland: Secretes mucus.
Stimulation of Olfactory Cells:
Odorant dissolves in mucus and attaches to receptors on olfactory cilia.
Involves a cascade effect through G-proteins and second messengers, leading to depolarization and action potential generation in the olfactory nerve.
Quality of a Good Odorant:
Small (3-20 Carbon atoms), volatile, water-soluble, and lipid-soluble.
Facilitated by odorant-binding proteins in mucus.
Membrane Potential and Action Potential:
Resting membrane potential: -55mV.
Action potential frequency in the olfactory nerve increases with odorant strength.
Adaptation Towards the Sense of Smell:
Rapid adaptation within the first second, with further slow adaptation.
Psychological adaptation greater than receptor adaptation, involving feedback inhibition from the central nervous system.
Primary Sensations of Smell:
Camphoraceous, Musky, Floral, Pepperminty, Ethereal, Pungent, Putrid.
Odor Detection Threshold:
Examples: Hydrogen sulfide (0.0005 ppm), Methyl-mercaptan (0.002 ppm).
Some toxic substances are odorless at lethal concentrations.
Characteristics of Smell:
Odor blindness for single substances due to lack of appropriate receptor protein.
Behavioral and emotional influences of smell.
Transmission of Olfactory Signals:
From olfactory cells to glomeruli in the olfactory bulb, involving lateral inhibition.
Primitive, less old, and new olfactory systems with different path
Ozempic: Preoperative Management of Patients on GLP-1 Receptor Agonists Saeid Safari
Preoperative Management of Patients on GLP-1 Receptor Agonists like Ozempic and Semiglutide
ASA GUIDELINE
NYSORA Guideline
2 Case Reports of Gastric Ultrasound
Ozempic: Preoperative Management of Patients on GLP-1 Receptor Agonists
Quality-by-design-based development and validation of a stability-indicating UPLC method for quantification of teriflunomide in the presence of degradation products and its application to invitro dissolution
1. Full Terms & Conditions of access and use can be found at
http://www.tandfonline.com/action/journalInformation?journalCode=ljlc20
Download by: [115.99.18.65] Date: 08 August 2017, At: 07:15
Journal of Liquid Chromatography & Related
Technologies
ISSN: 1082-6076 (Print) 1520-572X (Online) Journal homepage: http://www.tandfonline.com/loi/ljlc20
Quality-by-design-based development and
validation of a stability-indicating UPLC method
for quantification of teriflunomide in the presence
of degradation products and its application to in-
vitro dissolution
Nukendra Prasad Nadella, Venkata Nadh Ratnakaram & N. Srinivasu
To cite this article: Nukendra Prasad Nadella, Venkata Nadh Ratnakaram & N. Srinivasu (2017)
Quality-by-design-based development and validation of a stability-indicating UPLC method for
quantification of teriflunomide in the presence of degradation products and its application to in-
vitro dissolution, Journal of Liquid Chromatography & Related Technologies, 40:10, 517-527, DOI:
10.1080/10826076.2017.1330211
To link to this article: http://dx.doi.org/10.1080/10826076.2017.1330211
Accepted author version posted online: 19
May 2017.
Published online: 19 May 2017.
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3. class-2 compound of biopharmaceutical classification system
due to its low solubility and high permeability.[1–4]
Terifluno-
mide is weakly acidic with pKa 3.1 at room temperature and
having a pH-dependent solubility. In aqueous buffers at 25°C,
the solubility of teriflunomide increases from 0.02 µg mLÀ 1
at pH 1.2 to 8 mg mLÀ 1
at pH 7.6.[3]
Teriflunomide is an
immunomodulatory agent with anti-inflammatory properties,
inhibits dihydroorotate dehydrogenase, a mitochondrial
enzyme involved in de novo pyrimidine synthesis.[5–16]
Teriflu-
nomide contains two degradation products (Figure 1) namely
4-trifluoromethyl aniline (“leflunomide impurity-A Ph. Eur.”)
and 2-cyano-N-[4-(trifluoromethyl) phenyl]-acetamide (“leflu-
nomide impurity-H Ph. Eur.”).[1,17,18]
A stability-indicating method is a quantitative analytical
procedure used to detect a decrease in the amount of active
pharmaceutical ingredient present due to degradation.
According to Food and Drug Administration (FDA) guide-
lines,[19,20]
stability-indicating method is defined as a validated
analytical procedure that accurately and precisely measures
active ingredients free from potential interferences like degra-
dation products, process impurities, excipients, or other
potential degradation products. Quality-by-design (QbD) is a
systematic approach to development that begins with prede-
fined objectives and emphasizes product, process understand-
ing, and process control, based on sound science and quality
risk management. Key benefits of QbD are as follows.[21–23]
. High level of assurance of analytical method.
. The method is designed to meet predefined needs and
performance requirements.
. The impact of different reagents and method parameters
on analytical method quality is understood.
. Development of robust and cost-effective analytical method.
. Regulatory flexibility.
Teriflunomide is not official in pharmacopoeia such as
United States Pharmacopoeia (USP), BP, JP, and Ph. Eur. A
wide variety of analytical methods have been reported in the
literature for analysis of teriflunomide include estimation of
teriflunomide in biological fluids such as human plasma,
rabbit plasma, and human blood by high performance liquid
chromatography (HPLC) and LC–MS.[24–29]
However, there
are no methods reported in a study of the effect of stress on
pharmaceutical dosage forms, and there is no validated ultra-
performance liquid chromatography (UPLC) method on the
basis of QbD approach, which enables a stability-indicating
UPLC method for quantification of teriflunomide in bulk
and pharmaceutical dosage forms. To speed up the analysis,
UPLC method gives faster product development for the
pharmaceutical industry. In general, an UPLC method
provides 3 � higher efficiency and generates a 9 � increase
in throughput with no loss in resolution with sub-2-µ
particle-size columns than 5-µ particle sizes.[30,31]
The primary objective of the proposed research work is to
develop a stability-indicating UPLC method using QbD
approach for rapid estimation of assay content and in-vitro
dissolution release in the presence of degradation products
for pharmaceutical tablet dosage forms and to validate the
method as per ICH guidelines.[32]
Development of shorter
chromatography run time reduces the analysis time, cost effec-
tiveness, low solvent consumption altogether increases the
pharmaceutical productivity in routine quality control.
Experimental
Materials and reagents
Teriflunomide working standard and teriflunomide
film-coated tablets were provided by AET Laboratories Pvt
Ltd, Hyderabad, India. Dipotassium hydrogen phosphate
(K2HPO4), triethylamine, hydrochloric acid, sodium hydrox-
ide (NaOH), phosphoric acid, potassium dihydrogen phos-
phate (KH2PO4), and potassium chloride of Emparta grade
were purchased from Merck, India. Acetonitrile, methanol of
HPLC grade were procured from Merck, India, and Milli-Q-
water was collected from Merck Millipore ELIX 10 system.
Instrumentation
Ultraperformance liquid chromatography system with
Empower-3 software, UPLC BEH-C18 100 � 2.1 mm, 1.7-µ
column was used for chromatographic analysis. UV spectro-
photometer (Lambda 25, Perkin Elmer) was used for the
spectroscopic analysis. Analytical balance (XP-205 dual-range
model, Metler Toledo), dissolution apparatus (TDT-08 L,
Electrolab), Vacuum oven (Thermolab), vacuum filtration unit
(Millivac-Maxi 230 V, Millipore), pH meter (Orion-Star-A211,
Thermo), Rotary shaker (RS-24BL, REMI), water bath
(MSI-8, Meta Lab), photo-stability chamber (NEC103RSPSI,
Newtronics), and sonicator (9L250H, PCI) were used.
Chromatographic conditions
Chromatographies conditions were optimized based on
design-of-experiments (DoE) studies. The chromatographic
Figure 1. Molecular structure of teriflunomide and its degradation products.
518 N. P. NADELLA ET AL.
Downloadedby[115.99.18.65]at07:1508August2017
4. separation was achieved on UPLC BEH-C18 100 � 2.1 mm,
1.7-µ column using mobile phase composed of buffer (5 mM
K2HPO4 containing 0.1% triethylamine, pH 6.8) and acetoni-
trile in the ratio of 40:60 v/v. The flow rate was set at
0.5 mL minÀ 1
, and UV detection wavelength was performed
at 250 nm. Injection volume was 1 µL with a column tempera-
ture of 35°C. Total run time of method was 1 min. Fifty mM
phosphate buffer (pH 6.8) and acetonitrile in the ratio of
60:40 v/v were used as a diluent.
Standard and sample preparation
Weighed and transferred about 56 mg of teriflunomide working
standard into a 100-mL volumetric flask. Added 60 mL of dilu-
ent, sonication was done to dissolve and made up to volume
with diluent. From this solution, 5 mL was diluted to 50 mL
with diluent to obtain a concentration of 56 µg mLÀ 1
of teriflu-
nomide. Taken 20 tablets of the test sample into a mortar and
pestle, and then crushed to a fine powder. Weighed and trans-
ferred powder equivalent to 14 mg of teriflunomide into a
250-mL clean and dry volumetric flask. Then added 150 mL
of diluent and sonication was performed for 10 min with inter-
mittent shaking for extraction of the drug. The volume was
made up to 250 mL with diluent. The sample solution was
filtered through 0.2-µ polyvinylidene difluoride (PVDF) syringe
filter and collected the sample with a concentration of
56 µg mLÀ 1
of teriflunomide. Teriflunomide was quantified
using the following formula, where, “Ax” is the area obtained
from sample chromatogram, “As” is the average area obtained
from standard chromatograms, “Wstd” is weight of teriflunomide
standard in mg, “Wspl” is weight of sample in mg, “AW” is the
average weight of drug product, “LC” is label claim, and “P” is
the percentage assay of teriflunomide standard on as is basis.
Assay % label claimð Þ ¼
Ax
As
�
Wstd
100
�
5
50
�
250
Wspl
�
AW
LC
� P
In-vitro dissolution
The standard solution was prepared at a concentration of
14 µg mLÀ 1
of teriflunomide in phosphate buffer solution of
pH 6.8. The in-vitro dissolution test was performed using a
phosphate buffer solution of pH 6.8 as dissolution media with
a medium volume of 1000 mL and media temperature of 37°C.
The type of dissolution apparatus used is a paddle with a
rotation speed of 50 rpm. The sample was collected after
30 min time point, and samples were filtered through 0.2-µ
PVDF syringe filter.
Method validation
The developed UPLC method for quantification of terifluno-
mide was validated according to a current international con-
ference on harmonization guideline ICH Q2 (R1) validation
of analytical procedures.[32]
The method was validated for its
system suitability, specificity, linearity, accuracy, precision,
and robustness. Before analyzing the sample, system suitability
criteria were performed to verify whether the analytical system
(analytical solutions, UPLC system, column) is suitable or not
for giving accurate and consistent precise results. The system
suitability of the proposed method was evaluated by calculat-
ing parameters such as theoretical plates not less than 2000,
tailing factor not more than 2.0, and percentage relative stan-
dard deviation (RSD) from five standard injections should not
be more than 2.0.[33,34]
The specificity of the analytical method
was determined by verifying the interference of blank/placebo,
impurity peaks at the retention time of teriflunomide. Stan-
dard and sample solutions were prepared at a concentration
of 56 µg mLÀ 1
of teriflunomide. Placebo solution was prepared
similarly as sample preparation by taking a placebo and omit-
ting drug substance. Individual preparations of impurity-A
and impurity-H solutions were performed at a concentration
of 2.8 µg mLÀ 1
, i.e., 5% level with respect to sample concen-
tration. Solutions were injected into the chromatography sys-
tem by giving 200–400-nm wavelength ranges in photodiode
array (PDA) system. Recorded the chromatograms, verified
for peak purity of teriflunomide as well as interferences of
the blank, placebo and impurity peaks.
Forced degradation studies
Performed the forced degradation studies and all the
degradation samples were diluted with diluent after
completion of the degradation process. Blank and placebo
solutions were prepared in the same manner in respective
degradation to exclude any contribution from the process.
All the degradation sample solutions were injected into
UPLC-PDA system and recorded the chromatograms. Peak
purity was determined for teriflunomide and verified for any
interference of placebo/degradation products at the retention
time of teriflunomide. The total percentage of degradation
products and a percentage of the assay were calculated. Mass
balance was performed by adding together the assay value
and total percentage of degradation products to make up to
about 100% of the initial assay value of the drug product.
Linearity and range
The linearity of the analytical procedure is its ability (within a
given range) to obtain test results which are directly
proportional to the concentration of an analyte in the sample.
The linearity of the proposed analytical method was
determined by preparing five concentration levels from 28 to
84 µg mLÀ 1
of teriflunomide. The correlation coefficient (r),
regression coefficient (r2
), y-intercept, and slope of regression
line were calculated. The range of the method was proved by
performing the linearity, precision, and accuracy at the
proposed minimum 50% and maximum 150% concentration
levels with respect to sample concentration. Weighed and
transferred about 56 mg of teriflunomide into a 100-mL
volumetric flask. Dissolved and diluted to volume with diluent
to obtain a concentration of 560 µg mLÀ 1
of teriflunomide
(linearity stock). To prepare 50, 80, 100, 120, and 150% levels,
respectively, 5, 8, 10, 12, and 15 mL of linearity stock solutions
were diluted to 100 mL.
Accuracy
A known amount of teriflunomide drug substance was spiked at
50, 100, and 150% levels with respect to sample concentration
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5. to the placebo and analyzed by the proposed UPLC method.
Percentage recovery was calculated. Weighed and transferred
350 mg of teriflunomide into a 250-mL volumetric flask. Added
150 mL of diluent and sonication was performed to dissolve.
The diluent was added up to the mark to get the concentration
of 1400 µg mLÀ 1
of teriflunomide (accuracy stock solution).
Weighed and transferred placebo equivalent to one tablet
weight into a 250-mL clean and dry volumetric flask. Added
5, 10, and 15 mL of accuracy stock solution to 150 mL of diluent
to obtain 50, 100, and 150% concentration levels, respectively.
Solutions were sonicated for 10 min with intermittent shaking
and made up to volume with diluent.
Precision
The precision of an analytical procedure expresses the
closeness of agreement (degree of scattering) between a series
of measurements obtained from multiple sampling of the same
homogeneous sample under the prescribed conditions.
Precision was performed in repeatability and intermediate
precision methods. Repeatability for assay method was
demonstrated by preparing six assay sample solutions at a
concentration of 56 µg mLÀ 1
(100% level of sample concen-
tration) and injected into an UPLC system over a short
interval of time as per proposed method and calculated the
percentage RSD for assay results. Intermediate precision was
demonstrated by preparing six assay sample solutions at a con-
centration of 56 µg mLÀ 1
on a different day by different
analysts and then injected into a UPLC system as per proposed
method. Percentage RSD of assay results was calculated
between two analyst values.
Robustness
The robustness of an analytical procedure is a measure of
its capacity to remain unaffected by small, but deliberate var-
iations in method parameters, and it provides an indication of
its reliability during normal usage. Method robustness was
established by considering the variations in wavelength
(250 � 2 nm), flow rate (0.5 � 0.1 mL minÀ 1
), column oven
temperature (35 � 5°C), and a mobile phase ratio (40:60 v/v,
38:62 v/v, 42:58 v/v). Solution stability and filter interference
were established.
Results and discussion
Method development and optimization
Method development was initiated based on QbD concepts.
Quality target analytical profile (QTAP) forms the basis of
design for the development of analytical methods. QTAP
includes all the target requirements along with justification
and objectives of the method and are listed in brief in
Table 1.[21,35]
Method quality attributes were derived from
the QTAP. Critical method quality attributes (CMQA) are
the measurable parameters or characteristics of the method
that should be within predefined appropriate limits, ranges
or acceptance criteria. The CMQA for an analytical method
are primarily all the validation parameters of the method
including the robustness requirements.[32,35]
UV spectrum
of teriflunomide was determined in a solution containing
56 µg mLÀ 1
of teriflunomide in a diluent containing 50 mM
phosphate buffer of pH 6.8 and acetonitrile in the ratio of
60:40 v/v. PDA spectrum was collected from 200 to
400-nm wavelength (Figure 2). Wavelength maxima were
observed at 204.8, 248.8, and 295.7 nm. Though the highest
absorption is observed at 295 nm, degradation products
(impurity-A and impurity H) were observed at 250 nm.
Hence, 250 nm was selected as chromatographic detection
wavelength for teriflunomide peak. Since all peak responses
are at 250 nm, the method is suitable for mass balance
studies.
Method optimization
The solubility of teriflunomide was performed in aqueous
media with a pH ranging from 1.2 to 6.8. The solubility of
teriflunomide in water, hydrochloric acid media of pH 1.2,
acetate buffer of pH 4.5 and phosphate buffer of pH 6.8 were
insoluble, insoluble, 0.1, and 3.9 mg mLÀ 1
, respectively. Based
on the solubility of teriflunomide, pH of diluent and the mobile
phase buffer was considered in pH above 6. During preliminary
method development trials, the chromatographic conditions
were used as follows. Mobile phase-A was 10 mM KH2PO4
containing 0.5% v/v of triethylamine with pH 6.5. Acetonitrile
was used as a mobile phase-B. UPLC BEH C18 2.1 � 50 mm,
1.7 µ column was used with a flow rate of 0.5 mL minÀ 1
, the
detection wavelength of 250 nm, and an injection volume of
1 µL. Different solvent compositions of mobile phase were
studied for teriflunomide peak shape. Poor peak shape was
observed in preliminary method development trials and further
proceeded with pH scouting studies to evaluate the peak tailing
and noninterference of degradation products. pH scouting
studies were performed to select the optimum mobile phase
pH, to obtain shorter run time with no interference of placebo
and impurity peaks at the retention time of teriflunomide.
Mobile phases with different pH of buffers were prepared by
mixing buffer and acetonitrile in the ratio of 50:50 v/v. The
results of pH scouting studies were given in Table 2. pH
scouting studies reveal that the retention time of impurity-A
and impurity-H was not changing with the change in pH of
mobile phase buffer. But, the retention time of teriflunomide
peak was moving to lower retention time by increasing pH of
mobile phase buffer. Comparatively, the higher USP plate
count was observed at buffer pH 2.0, but the impurity peaks
were eluted nearby the teriflunomide peak. And also, lower
pH was not suggestible due to poor solubility of teriflunomide.
Hence, pH of mobile phase buffer between 6.0 and 7.0 was
selected for optimization to achieve a lower retention time of
teriflunomide without the interference of degradation products
and to overcome any solubility issues due to lower pH.
Design-of-experiment studies
The design of experiments was executed to select robust and
rugged operational chromatographic conditions within the
design space. Ten mM KH2PO4 containing 0.5% v/v of
triethylamine with pH 6.8 was taken as a mobile phase buffer
520 N. P. NADELLA ET AL.
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6. for DoE studies. DoE study was executed by considering the
mobile phase composition, flow rate, column temperature as
control variables. USP tailing, the retention time of late eluting
impurity peak and resolution of nearby impurity with
teriflunomide were taken as response variables. Design Expert
8.0 software was used for the study. The selected design
parameters are response surface as study type, central
composite as design type and quadratic as design mode.
Proposed ranges of mobile phase solvent composition
50–70%, flow rate 0.4–0.7 mL minÀ 1
, and column temperature
25–45°C were selected to design. This data were fed into the
design expert software and software given twenty experiments.
All the DoE experiments were executed in UPLC and observed
results are tabulated in Table 3. The effect of control variables
on response variables was graphically evaluated in Figure 3.
The observations were derived from perturbation plot and
3D plot which describes the effect of method control variables
on the response variables.
Based on the desirability plot (Figure 4), column
temperature was set at 35°C. Design space plot (Figure 4)
suggests that flow rate of 0.5 mL minÀ 1
and mobile phase
composition of 40:60 v/v (buffer:acetonitrile) for a robust
chromatographic method. Teriflunomide peak was eluted at
0.5 min and run time was set at 1 min. Risk assessment is a
valuable scientifically based process used in quality risk
management. Risk assessment is helpful in identifying the
input material attributes (reagents/chemicals/columns) and
method parameters (flow, temperature, solution stability) that
are affecting CMQA. The first step of the risk-assessment
process involves identification of critical material attributes
and method parameters and performing risk assessments of
their attributes for the subsequent effect on CMQA. Based
on risk assessment, control strategy was established for the
selection of diluent pH at 6.8 for solubility of teriflunomide.
Method validation
Specificity
Diluent, placebo, standard, sample, impurity-A, impurity-H,
and impurity-spiked sample solutions were injected into
Table 1. Quality target analytical profile.
Analytical target profile
element Target/requirement Justification
Status of current study
(yes/no/remarks)
Type of method Quantification of teriflunomide in the
presence of degradation products
To quantify the teriflunomide in the
formulation
Yes
Mode of detection and
chromatography
UV, isocratic The molecule is having chromophoric groups
and can be detected by UV
Yes, UV detection at 250 nm
Specificity Blank, placebo, and impurity
interference should not be observed
ICH Q2 (R1) guideline requirements Yes
Precision of the method,
repeatability/
reproducibility
Should have a precision
withpercentage RSD below 2.0
ICH Q2 (R1) guideline requirements Yes
Accuracy Percentage recoveryshould be
between 98 and 102
The percentage recovery should be good for
the drug product and as per requirements
of ICH guidelines
Yes
Linearity Linearity at different concentration
levels should be obtained
The correlation coefficient should not be less
than 0.99 and as per requirements of ICH
guidelines
Yes
Robustness Assay results should not be affected
by small changes in method
parameters
Results should not be affected Yes
Stability indicating nature The principle peak should be pure
even after forced degradation
Purity angle should be less than purity
threshold and the mass balance should be
close to 100% of initial assay
Yes
Green chromatography Should use minimum percentage of
organic phase as possible
To avoid the use of more organic solvents and
to develop an environment-friendly method
Yes, low solvent consumption with
a shorter run time of 1 min
Figure 2. UV spectrum of teriflunomide.
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7. UPLC-PDA system, and chromatograms were extracted. The
results are summarized in Table 4. The system suitability test
results observed during specificity and precision test are
tabulated in Table 5. Acidic, basic, oxidation, thermal, water,
humidity, and photodegradation samples were injected into
UPLC-PDA system and chromatograms were extracted. No
interference was observed with blank, placebo, and impurities
at the retention time of teriflunomide. Teriflunomide peak
passed the peak purity test for all degradation samples. The
results are summarized in Table 4. Mass balance was achieved
for all forced degradation samples and the results are
summarized in Table 4. The representative chromatogram of
specificity is given in Figures 5 and 6.
Linearity and range
Linearity solutions, i.e., 50, 80, 100, 120, and 150% levels were
injected into UPLC and chromatograms were recorded. The
regression line of analysis shows the linear relationship
between concentration and area response of teriflunomide.
Results of linearity and range are summarized in Table 5.
Accuracy and precision
Teriflunomide-spiked samples of 50, 100, and 150% levels
with respect to sample concentration to the placebo were
analyzed by the proposed UPLC method. Recovery of
teriflunomide was observed from 100.1 to 101.7% and all the
individual results were within the range of 98–102% criteria.
The results are summarized in Table 6. The precision of the
analytical method was determined by repeatability and
intermediate precision. The percentage RSD results for
repeatability, intermediate precision, and between two analyst
values were 1.03, 1.19, and 1.15, respectively (Table 6). Since
the percentage RSD of six assay results is not more than 2.0,
the method is repeatable. The percentage RSD of two analyst’s
assay results is less than 2.0, hence, intermediate precision is
acceptable.
Robustness
Method robustness was established by considering the changes
in wavelength, flow rate, column oven temperature, and mobile
phase ratio. Solution stability and filter interference were
studied. Hydrophilic polypropylene (GHP) and Millipore-
PVDF syringe filters were evaluated for filter interference and
no significant interference was observed. The percentage differ-
ences of area response from the unfiltered area for GHP and
PVDF were 0.28 and 0.38, respectively. The results of robust-
ness are tabulated in Table 5. Robustness test passed as a vari-
ation from initial results is not more than 2.0%.[36–40]
Table 3. Experimental results of DoE study and effect on response variables.
DoE experiments Results of response variables
Run no.
Column temperature
(°C)
Flow rate
(mL minÀ 1
)
Mobile phase
composition (v/v) USP tailing
Retention time of late
eluting impurity (min)
Resolution of nearby
impurity with teriflunomide
1 25 0.6 50:50 1.70 0.997 10.35
2 45 0.6 50:50 1.54 0.944 9.76
3 35 0.4 40:60 1.30 1.030 7.63
4 45 0.4 30:70 1.19 0.806 4.42
5 35 0.6 40:60 1.50 0.681 6.87
6 45 0.4 50:50 1.40 1.428 9.61
7 35 0.5 40:60 1.43 0.822 7.17
8 45 0.5 40:60 1.36 0.806 6.84
9 35 0.5 40:60 1.43 0.822 7.17
10 25 0.4 50:50 1.52 1.539 10.51
11 25 0.5 40:60 1.52 0.840 7.33
12 35 0.5 40:60 1.43 0.822 7.17
13 35 0.5 50:50 1.58 1.175 10.7
14 35 0.5 40:60 1.43 0.822 7.17
15 25 0.6 30:70 1.07 0.549 4.12
16 35 0.5 30:70 1.10 0.657 4.27
17 35 0.5 40:60 1.43 0.822 7.17
18 45 0.6 30:70 1.08 0.536 3.88
19 35 0.5 40:60 1.43 0.822 7.17
20 25 0.4 30:70 1.22 0.831 4.62
Increase of control variable
Effect on response variables
USP tailing RT of late eluting peak Resolution of nearby peak with teriflunomide
Observations from DoE study plots
A: Column temperature Decreases No effect No effect
B: Flow rate Increases Decreases No effect
C: Mobile phase solvent composition Decreases Decreases Decreases
DoE, design-of-experiments.
Table 2. Results of pH scouting experiments.
Buffer pH RT (retention time) of teriflunomide (min) RT of impurity-A (min) RT ofimpurity-H (min) USP tailing USP plate count
pH 2.0 1.388 1.263 1.130 1.59 9251
pH 3.0 0.808 1.322 1.130 2.02 4694
pH 4.0 0.661 1.329 1.133 1.90 3359
pH 5.0 0.590 1.332 1.134 1.90 3151
pH 6.0 0.587 1.327 1.136 1.90 3157
pH 7.0 0.579 1.333 1.134 1.38 3048
522 N. P. NADELLA ET AL.
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8. Comparison with reported methods
Ultraperformance liquid chromatography method was
developed for rapid quantification of teriflunomide in the
presence of degradation products in the pharmaceutical drug
product. The present method uses the shortest run time of
1 min with a retention time of teriflunomide of about
0.5 min compared to earlier reported methods which are
having a run time of minimum 2 min with an RT of
1.43 min (Table 7). The method was developed based on
QbD approach, and optimum chromatographic conditions
were selected based on design space obtained through DoE
studies. The developed method is stability indicating as there
is no interference in force degradation studies (Table 4). The
method was validated as per ICH guidelines and the results
of specificity, linearity, accuracy, precision, and robustness
were found satisfactory. No interference was observed with
blank, placebo, and degradation products at the retention time
of teriflunomide. The purity angle was less than the purity
threshold indicating that teriflunomide peak was free from
interference and passed the peak purity test (Table 4). The
reported methods were given for estimation of teriflunomide
release rate in biological fluids.[24–28]
Present method was
developed for rapid quantification of teriflunomide in the
presence of potential impurities by QbD in pharmaceutical
bulk and finished dosage form. Sample injection volume
(1 µL) in the present method is significantly lower, which helps
to maintain the good column performance compared to earlier
reported methods (minimum of 5 µL). A detailed comparison
of selected procedures with the present method is given in
Table 7. Teriflunomide contains no asymmetric centers,
Figure 3. Affect of control variables A (column temperature), B (flow rate), C (mobile phase solvent composition) on response variables a) USP tailing, b) resolution,
c) retention time of late eluting impurity.
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9. Figure 4. Schematic representations of desirability plot and design space plot.
Table 5. System suitability evaluation, linearity, and robustness results.
Parameter Acceptance criteria[33]
Results of the test
RemarksSpecificity Precision
RSD (%) of area/five injections Not less than 2.0 0.38 0.38 Satisfactory
USP tailing factor Not more than 2.0 1.66 1.51 Satisfactory
Theoretical plates Not less than 2000 8134 9938 Satisfactory
Type of test Assay In-vitro dissolution
Linearity test results
Test concentration (µg mLÀ 1
) 28.02–84.06 2.84–22.70
Correlation coefficient (R) 0.9999 1.0000
Regression coefficient (R2
) 0.9998 0.9999
Slope 10124.6444 19885449.4184
Intercept 1919.441 1646.58
Parameter Change done Results Remarks
Robustness results
Solution stability Initial 100.6 Solutions are stable for 24 hr
After 24 hr 101.1
Wavelength (250 � 2 nm) 250 99.1 No significant variation in results
248 99.2
252 99.3
Flow rate (0.5 � 0.1 mL minÀ 1
) 0.5 99.1
0.4 99.8
0.6 100.1
Column oven temperature (35 � 5°C) 35 99.1
30 98.8
40 99.2
Mobile phase ratio (buffer:acetonitrile, v/v) 40:60 99.1
38:62 99.2
42:58 98.5
Table 4. Specificity, forced degradation studies, and mass balance results.
Sample name Retention time (min) Purity angle Purity threshold Peak purity Assay (%) Impurities (%) Total Mass balance
Impurity-A 0.813 — — — — — — —
Impurity-H 0.722 — — — — — — —
Teriflunomide standard 0.521 22.675 63.543 Pass — — — —
Sample 0.521 20.704 73.281 Pass 99.1 Nil 99.1 Yes
Acid-degradation 0.518 23.388 62.978 Pass 99.3 0.9 100.2 Yes
Alkali-degradation 0.524 1.509 72.977 Pass 99.2 Nil 99.2 Yes
Oxidation with KMnO4 0.517 22.161 90.000 Pass 91.6 6.6 98.2 Yes
Photo degradation-UV light 0.518 23.954 71.828 Pass 100.5 Nil 100.5 Yes
Photo degradation-visible light 0.519 21.555 66.176 Pass 100.2 Nil 100.2 Yes
Thermal-degradation 0.518 16.006 46.652 Pass 99.1 Nil 99.1 Yes
Water-degradation 0.519 21.389 59.804 Pass 96.8 2.3 99.1 Yes
524 N. P. NADELLA ET AL.
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11. therefore no enantiomers are possible, but can exist in E and Z
isomers. E-isomer will not separate in existing developed
method. As per literature review, there are no methods
available for the separation of these isomers. There is a scope
for future research work on separation of E and Z isomers of
teriflunomide.
Conclusions
The UPLC method was developed based on QbD concepts for
rapid quantification of teriflunomide in pharmaceutical drug
products. The developed method was validated as per ICH
guidelines. The method was found to be simple, selective, accu-
rate, precise, and robust. The developed method was stability
indicating as it was showing no interference of degradation pro-
ducts and placebo at the retention time of teriflunomide. Due to
the shorter run time of 1 min, this method provides faster analy-
sis, more work throughput, and reduces the cost of analysis due
to the reduction in solvent consumption. The method can be
used for in-vitro dissolution analysis. Therefore, the developed
method can be used for routine assay and in-vitro dissolution
analysis of quality control samples and stability samples of bulk
and finished pharmaceutical dosage form.
Acknowledgments
The authors wish to thank the management of AET Laboratories Private
Limited, Hyderabad, India for giving us an opportunity to carry out the
dissertation work.
References
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[5] EU Summary of Product Characteristics: Aubagio, Sanofi-Aventis.
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[6] US Prescribing Information: Aubagio, Sanofi. http://products.sanofi.
us/aubagio/aubagio.pdf (accessed December 20, 2015).
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Teriflunomide for Multiple Sclerosis. Cochrane Database Syst. Rev.
2012 (Art. No. CD009882); DOI: 10.1002/14651858.CD009882
[8] Bar-Or, A.; Pachner, A.; Memguy-Vacheron, F.; Kaplan, J.; Wiendl,
H. Teriflunomide and Its Mechanism of Action in Multiple
Sclerosis. Drugs 2014, 74, 659–674.
[9] Miller, A. E.; Teriflunomide: A Once-daily Oral Medication for the
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Table 7. Comparison of selected analytical methods developed for teriflunomide.
S. no.
Column, elution process, mobile phase, flow rate,
injection volume
Sample linear range,
detection
Run time (RT of
teriflunomide) Intended use Refs. no.
1 Phenomenex Luna2 PFP, 2 � 100 mm, 3 µ,
gradient, mobile phase-A (0.1% formic acid),
mobile phase-B À 0.1% formic acid in
methanol:water:acetonitrile (0.5:0.5:9 v/v/v),
300 µL minÀ 1
, 100 µL
0.005–200 µg mLÀ 1
,
LC–MS/MS
4 min (2.18 min) For estimation of release
rate in biological sample
[24]
2 Inertsil-ODS-3 C18 (50 � 4.6 mm, 5 µ); isocratic,
0.02 M CH3COONH4 pH 6.5: methanol (25:75
v/v); 0.8 mL minÀ 1
, 5 µL
10.1–4000 ng mLÀ 1
,
LC–ESI-MS/MS
2.0 min (1.43 min) For estimation of release
rate in biological sample
[25]
3 PolySULFOETHYL aspartamide strong cation
exchange column; isocratic, 10 mmol LÀ 1
KH2PO4 and 100 mmol LÀ 1
KCl in aqueous 25%
acetonitrile, acidified to pH 3 with o-phosphoric
acid. 0.7 mL minÀ 1
, 10 µL
0–200 µg mLÀ 1
, HPLC–
UV at 280 nm
7 min (2.9 min) For estimation of release
rate in biological sample
[26]
4 XTerra-MS-C18 analytical column (100 � 3.9 mm,
5 µ), gradient, mobile phase-A-0.005 M
ammonium formate (pH 9.0) and B-acetonitrile
at 1.0 mL minÀ 1
, 10 µL
1–4000 ng mLÀ 1
,
LC–ESI-MS/MS
15 min (6.7 min) For estimation of release
rate in biological sample
[27]
5 Nucleosil 100-5 C18-column, gradient, mobile
phase A-0.5 mM ammonium acetate in water–
acetonitrile–formic acid (9:5:0.02 v/v/v) and
mobile phase-B-0.5 mM ammonium acetate in
water–acetonitrile–formic acid (5:95:0.02 v/v/v),
0.5 mL minÀ 1
, 60 µL
5–500 µg mLÀ 1
,
LC–ESI-MS/MS
7 min (3.1 min) For estimation of release
rate in biological sample
[28]
6 Acquity-UPLC BEH-C18 2.1 � 50 mm, 1.7 µ
column, isocratic, 5 mM K2HPO4 buffer
containing 0.1% triethylamine (pH 6.8) and
acetonitrile (40:60 v/v), 0.5 mL minÀ 1
, 1 µL
28–84.1 µg mLÀ 1
,
UPLC–UV at 250 nm
1 min (0.5 min) For quantification of
teriflunomide in the
presence of potential
impurities
Present
method
526 N. P. NADELLA ET AL.
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