Analysis of Aspirin (Acetylsalicylic Acid): Development and Validation using RP-HPLC Methods - A Component of Anti-hypertensive Drugs

 

Monika Puri¹, Yogesh Kumar Walia²

¹Research Scholar, Career Point University, Hamirpur (H.P.), India.

²Professor, Department of Chemistry, Career Point University, Hamirpur (H.P.), India.

 

ABSTRACT:

A precise, sensitive, and focused liquid chromatographic method has been developed to assess the quantity of aspirin in its bulk form. The chromatographic conditions involved utilizing a C-18 column (250 x 4.6 mm, 5 µm particle size) with a mobile phase consisting of water containing 0.1% orthophosphoric acid (v/v) at pH 3.0 and acetonitrile in a ratio of 45:55. The flow rate was set at 1 ml per minute, and the total run time was 20 minutes, with a detection wavelength of 237 nm. The retention time was observed to be 4.01 minutes, and the method exhibited an average recovery percentage of 99.9%. The proposed procedure adhered to the guidelines established by the ICH and is applicable for analyzing aspirin as a standalone bulk drug and in combination with other pharmaceuticals.

 

 

 

1. INTRODUCTION: 

Many new drugs are released on the market daily. The period between the launch of these drugs and their inclusion in pharmaceutical protocols is frequently longer¹. This is because of potential problems in the continuous and extensive use of these drugs, new side effects that arise, increased resistance in patients, and better drugs that are offered by rival companies. In such circumstances, probably, Pharmacopoeias do not provide access to established scientific strategies for these pharmaceuticals². In this approach, developing a new explanatory mechanism for such treatments becomes necessary. Additionally, since medications contain life, quality is important in every service or product. The investigation reveals the spatial organization of an ion as a particle and the location or proximity to a particular natural beneficial gathering in a specific compound³.

 

In the first step, which involves choosing a system, care should be taken to select the ideal instrument for a fruitful investigation. Making an incorrect choice right now will result in a pointless investigation. The three main categories of analytical techniques are physical, chemical, and instrumental analysis^4,5. Physical observations include compound descriptions, measurements of its dimensions (size, form), color, and odor, among other things. Part of chemical analysis is the titrimetric analysis of compounds, such as potentiometric, audiometric, argentometric, permagnometric, and so on. Experimental chemistry depends largely on instrumental techniques for chemical analysis⁶.

 

Both new and current products undergo method development. It is more difficult to implement an adequate optimization lesson and improve on the level of familiarity obtained via such a lesson when there are no immediate lit equilibriums between what is desired and what has already been understood.An official analytical method is used to evaluate the characterized normal for a medicine substance or medication item. The candidate suggests logical technique as an alternative to administrative systematic method. Security testing is a pivotial step in the evolution of pharmaceutical products⁷.

 

Modern methods for quantitative examination decision-making include:

 

1.1 Chromatography:

The main goal of chromatographic procedures is the separation of substances in a sample mixture, rather than the separation of colour. A variety of separation methods have emerged to accommodate the many physical and chemical states of sample mixtures that one may desire to separate and study⁸. Chromatography is unique from most other physical and chemical separation techniques because it brings two mutually immiscible phases into contact, one stationary and one mobile. The stationary phase can only be a liquid or a solid, but the mobile phase can also be a gas. Liquid-liquid chromatography is the term for the separation that mostly involves a simple partitioning between two immiscible liquid phases, one stationary and one mobile^9,10. Liquid solid chromatography is the term for the procedure that mainly relies on physical surface forces for the stationary phase’s ability to retain samples. Liquid chromatography has been conducted in a column or on an open bed¹¹.

 

Reversed-phase: This is the most prevalent HPLC mode, also referred to as RP-HPLC, which is the inverse of NP-HPLC, where the stationary phase is more nonpolar than the eluting solvent¹².

 

RP-HPLC generally employs a nonpolar stationary phase, such as C-18 silica, and a moderately polar aqueous mobile phase¹³. A typical stationary phase for RP-HPLC is surface-modified silica, also known as RMe₂SiCl, where R is a straight-chain alkyl group, such as C₁₈H₃₇ or C₈H₁₇^14,15.

 

Nowadays, HPTLC is becoming a standard investigative technique¹⁶ as a result of  its advantages like inexpensive working costs, minimal example cleaning requirements, andlarge specimen throughput^17,18. A key advantage of HPTLC is that, unlike HPLC, a limited number of samples may be run simultaneously using a small amount of a flexible stage¹⁹, reducing the amount of time and money required for each research²⁰.

 

A common medication for various causes of pain and fever is aspirin (Fig1)²¹, or acetylsalicylic acid (ASA). It reduces fever and inflammation. It also prevents platelet aggregation and is used to treat blood clots, stroke, and MI (myocardial infarction)²².

 

Molecular formula of ASA: C₉H₈O₄

 

Molecular weight of ASA: 180.1574g/mol

 

IUPAC Name of ASA: 2-(acetyloxy) benzoic acid

 

Other Names: Acid acetylsalicylique

 

Physical state of ASA: Solid

 

Fig. 1: Aspirin

 

2. EXPERIMENTAL WORK:

2.1 Chemicals and reagents:

Orthophosphoric acid, acetonitrile and HPLC-grade water were purchased from Rankem Standars Aspirin NIST traceable were purchased from Sigma Aldrich. Acetylsalicylic acid (>=99.0%) was purchased from Merck.

 

2.2 Instrumentation:

High performane liquid Chromatograph equipped with autosampler and UV Detector (Waters) was performed at 237nm and Column- C18 (250nm X 4.5mm X 5µm).

 

2.3 Determination ofλ_max of Aspirin:

The λ_max of aspirin was determined by running the spectrum of the drug solution in a double-beam UV spectrophotometer. Aspirin showed good absorbance at 237nm so it is selected as wavelength of detection.

 

2.4 Selection and procedure for preparation of mobile phase:

Initially, various mobile phase ratios were tested to estimate aspirin in a fixed dosage form. The mobile phase that was determined to be the most acceptable for analysis is as follows: pH of HPLC Water 3.00±0.05 with 0.1% orthophosphoric acid (v/v) and mix with Acetonitrile in the ratio of 45:55.

 

2.5 Selection of solvent:

The mobile phase is used as a solvent for preparing standard and sample solutions. The solvent/mobile phase was used as the blank solution.

 

2.6 Standard Solution Preparation Scheme:

2.6.1 Standard Stock Solution: Weigh 37.5mg of Aspirin, in a 50mL volumetric flask and 25mL of solvent/mobile phase and mix well. Sonication was done for five minutes. Cool the solution to room temperature and make up the volume with solvent^23,24,25.

 

2.6.2 Standard Working Solution: Pipet the Standard stock solution (5mL) and dilute to 50mL with solvent.

 

2.6.3 Concentration of working standard: Aspirin – 75ppm (75µg/mL). Filter the working standard solution using the 5-10mL disposable syringe and PTFE filters into a 2-ml Autosampler vial and seal with the caps.

 

2.7 SAMPLE SOLUTION PREPARATION:

2.7.1 Sample Preparation:

1.     Transfer the 75/150mg Aspirin dosage form to the 100/200mL volumetric flask. 50% of the volumetric flask should be filled, and then thoroughly mixed. Until the sample is completely dissolved, sonicate it. Shake ferociously to guarantee complete sample dissolution. Once the solution has reached room temperature, dilute it to create volume.

2.     Pipet 5ml of the above solution and dilute to 50mL with solvent.

3.     Concentration of sample standard - Aspirin – 75ppm (75µg/mL),

4.     Use the 3ml disposable syringe and PTFE filters to filter the sample working solution before transferring it to a 2ml Autosampler vial and capping it.

 

2.8 OPERATING CONDITIONS:

 

Table 1: Typical Instrument Settings

Detector	237nm
Flow rate	1.0 mL/min
Injection volume	20 µL
Mobile phase	Water at pH 3.00 with 0.1 %orthophosphoric acid (v/v) and Acetonitrile (45:55)
Running time	20 min
System suitability	Retention Time:Aspirin- 4 min
Tailing factor	Not More than 1.2 for analyte in standard solution
Theoretical plates	Not Less than 2,500 for all analyses in standard solution
Resolution	Not less than 2.0

 

3. EXPERIMENTAL PROCEDURE:

1.     Permit the instrument to operate in the chosen conditions.

2.     Inject the Blank Solution to verify the system's specificity.

3.     Determine the aspirin retention time for system testing by injecting the working standard solution.

4.     For system appropriateness, inject the Reference Standard five duplicate. The peak area for the five injections' relative standard deviation (RSD) shouldn't be more than 2.0%.

5.     To verify continued system appropriateness, inject the working standard solution after each sequence of ten or fewer injections of samples.

6.     For the entire set of Standard injections, the Global RSD shouldn't be more than 2.0%.

7.     Use the average of five replicate standard injections to inject each sample twice and calculate the sample size.

4. RESULTS AND DISCUSSION:

4.1 SPECIFICITY:

In analytical chemistry, specificity is a term that refers to the ability of a method to differentiate the analyte from other components in the sample. Specificity is the ability to assess an analyte with confidence in the presence of an additional component that may be present²⁶.

 

Table 2: Results of Specificity

Solutions	Retention Time
Blank	0.2
Standard - Aspirin	4.01
Sample - Aspirin	4.01
Capacity Factor - Aspirin	19.1

 

Observations and Conclusions: In blank, no peak observed corresponds to the retention time of the peak of Aspirin in standard solution (Table 2). The analytical method for the estimation of Aspirin in therapeutic dose form is specific.

 

4.2 LINEARITY:

The relationship between the test results and the concentration of analyte in the sample is described by linearity, a term used in analytical chemistry. Linearity is the ability of an analytical method to get test results that are in proportion to the concentration of analyte in the sample within a defined range, either directly or by a clear mathematical transformation²⁷.

 

Fig. 2: Linearity curve of Aspirin

 

Table 3: Linearity Data

Linearity - Aspirin
Injection	Area	Average	Conc.
1	984640	984560	80%
2	984480
1	1107720	1107630	90%
2	1107540
1	1230800	1230700	100%
2	1230600
1	1353880	1353770	110%
2	1353660
1	1476960	1476840	120%
2	1476720
Correlation coefficient		1.0000

 

Observation and Conclusion:

The observed correlation of Co-efficient is 1.000 which meets the acceptance criteria. Hence Analytical method for the estimation of Aspirin, in therapeutic dose formmeets the Linearity criteria (Table 3).

4.3 SYSTEM SUITABILITY:

The precision of an instrument is the degree of agreement among the replicate injection of the standard ²⁸.

 

Table 4: System Suitability Data

Injection	Area of Aspirin
1	1230914
2	1229830
3	1231620
4	1230614
5	1230520
Mean of Data	1230700
Standard-Deviation	649.79
Relative Standard-Deviation (%)	0.05
RT	4.01
RRT	N/A
Theoretical-Plates	3347
Tailing Factor	1.01
Resolution	N/A

 

Observation and Conclusion: The observed RSD of the replicate five standard injections 0is less than 2.0% which meets the system suitability parameter (Table 4). Hence, developedanalytical method for the estimation of Aspirin in therapeutic dose formmeets the system precision criteria.

4.4 PRECISION (REPEATIBILITY):

The degree of agreement or consistency between repeated measurements of the same sample under the same conditions is described by precision, a term used in analytical chemistry²⁹. When the procedure is frequently used on various preparations of a homogenous sample, the precision of an analytical method is the resemblance of the results from separate tests.

 

Table 5: Results of Precision(Repeatability)

Test Preparation	Assays of Aspirin
1	99.2%
2	98.8%
3	97.9%
4	100.3%
5	101.2%
6	100.8%
Mean of Data	99.7%
Standard-Deviation	0.013
Relative Standard-Deviation (%)	1.28

Observation and Conclusion: The observed Related Standard Deviation of the six different determinations is less than 2.0% which meets the acceptance criteria. Hence developed analytical method for the estimation of Aspirin, in therapeutic dose formmeets the method precision (Repeatability) criteria (Table 5).

 

4.5 PRECISION (REPRODUCIBILITY):

When an analytical method is used repeatedly on various preparations of a homogenous sample, its precision is calculated by the degree of agreement between individual test findings ³⁰.

 

Table 6: Results of precision (Reproducibility)

Test Preparation	Assays of Aspirin
1	100.0%
2	99.5%
3	98.6%
4	99.9%
5	99.6%
6	99.9%
Mean of Data	99.6%
Standard-Deviation	0.005
Relative Standard-Deviation (%)	0.52

 

Table 7: Summarised Results of precision

% Aspirin First Set	% Aspirin Second Set	% Difference between the mean
99.7%	99.6%	0.1%

 

Observation and Conclusion: The observed Related Standard Deviation of the six different determinations is less than 2.0% which meets the acceptance criteria and also % difference between an average of repeatability and reproductivity is less than 2.0%. Hence developedanalytical method for the estimation of Aspirin, in therapeutic dose form meets the method precision (Reproducibility) criteria (Table 6,7).

4.6 ACCURACY (RECOVERY):

The closeness of the test results produced using the suggested method to the actual value is the accuracy ³¹ of an analytical procedure. By applying the approach to a Placebo that has known amounts of Analyte added at three concentration levels—80%, 100%, and 120% of test concentration, the method’s accuracy can be measured.

 

Table 8: Accuracy Data

Test ID	Analyte	Wt. standard (gm)	Wt. of Placebo (gm)	Amount of standard added in (ml)	% Added	% Recovered	Recovery %
80%-1	Aspirin	0.0375	0.221	4	27.15	27.25	100.4
80%-2	Aspirin	0.0375	0.222	4	27.03	27.12	100.3
80%-3	Aspirin	0.0375	0.225	4	26.67	26.82	100.6
100%-1	Aspirin	0.0375	0.218	5	34.40	34.39	100.0
100%-2	Aspirin	0.0375	0.220	5	34.49	33.98	99.7
100%-3	Aspirin	0.0375	0.221	5	33.94	33.81	99.6
120%-1	Aspirin	0.0375	0.223	6	40.36	40.31	99.9
120%-2	Aspirin	0.0375	0.218	6	41.28	41.12	99.6
120%-3	Aspirin	0.0375	0.221	6	40.72	40.90	100.4

 

Observation and Conclusion: The observed individual percentage recovery at all the levels i.e. 80%, 100%, and 120% is in the range of 96-104%, and mean recovery at all levels is in the range of 97-103% which meets the acceptance criteria (Table 8). Hence developedanalytical method for the estimation of Aspirin in therapeutic dose formmeets the Accuracy (Recovery) criteria.

 

4.7 RANGE:

The degree to which test findings generated using the advised approach are close to the real value is the analytical procedure's accuracy. It will be possible to assess the method's precision by using Placebo to which known amounts of analyte have been added at three different concentration levels such as 80%, 100%, and 120% of test concentration ³².

Observation and Conclusion:

The accuracy, precision, and linearity meet the acceptance criteria. Hence range of 80-120% of test concentration is assigned to the Analytical method for the determination of Aspirin in therapeutic dosage.

 

4.8 ROBUSTNESS:

The robustness ³³ of an analytical method is its ability to stay unaffected by small but deliberate variations in method parameters, which shows its reliability under normal conditions. The robustness of the approach is evaluated by changing the mobile phase composition, the number of columns, and the flow rate.

 

Observation and Conclusion: With all the changes system suitability parameter was achieved and the observed % variation in the result obtained with deliberate change is less than ± 2% which meets the acceptance criteria (Table 9). Hence developedanalytical method for the determination of Aspirin in therapeutic dose formmeets the Robustness criteria.

 

 

4.9 SOLUTION STABILITY:

The standard and extracted sample solutions (ready to inject) should be kept at room temperature or in a fridge, depending on how stable they are. The solution stability refers to how well the solutions from the sample or matrix can be analyzed by the given method ³⁴.

 

Observation and Conclusion: A 36-hour solution stability investigation found that there was less than 2% variance in the results, which fulfils the standards for acceptance. Hence 24 hours can be employed with an analytical solution created for the measurement of aspirin in therapeutic dose (Table 10).

 

4.10 Reference Chromatogram:

 

Fig. 3: Chromatogram of (a) Blank (b) Aspirin

 

5. CONCLUSION:

For Aspirin the RP-HPLC method was created. Column C-18 (250 x 4.6mm) with particle size 5 um, mobile phase water at pH 3.00 with 0.1% orthophosphoric acid (v/v) and acetonitrile (45:55) were discovered to be the chromatographic requirements for the optimized procedure. For aspirin, the retention-time was determined to be 4.01min, with an average percentage recovery of 99.9%. It was determined that the suggested approaches complied with ICH criteria. These techniques can be used in the future to regularly determine the presence of Aspirin in bulk medications along with other medicines in combination.

 

Table 9: Robustness Data

Test ID	Analyte	% Assay First Set - Method Precision	% Assay First Set - Robustness	% Difference
Buffer pH-2.95	Aspirin	99.70	99.60	0.01
Buffer pH-3.05	Aspirin	99.70	99.40	0.30
Buffer: CAN (40:60)	Aspirin	99.70	99.10	0.60
Buffer: CAN (50:50)	Aspirin	99.70	98.90	0.80
Change in column lot	Aspirin	99.70	99.10	0.60
Change in column particle size	Aspirin	99.70	99.90	0.30
Change in flow rate 0.9 ml	Aspirin	99.70	98.70	1.00
Change in flow rate 1.1ml	Aspirin	99.70	99.40	0.30

Table 10: Results of Solution Stability

Test id	Analyte	Initial	6 Hours	12 Hours	18 Hours	24 Hours	36 Hours
Solution Stability	Aspirin	99.70%	99.50%	99.20%	99.90%	100.00%	100.20%
% Difference	Aspirin	N/A	0.20%	0.50%	-0.20%	-0.30%	0.32%

 

6. REFERENCES:

1.      Allen, L. and Ansel, H.C., 2013. Ansel's pharmaceutical dosage forms and drug delivery systems. Lippincott Williams and Wilkins.

2.      Wright, P.M., Seiple, I.B. and Myers, A.G., 2014. The evolving role of chemical synthesis in antibacterial drug discovery. Angewandte Chemie International Edition, 53(34): 8840-8869.

3.      Yao, J., Yang, M. and Duan, Y., 2014. Chemistry, biology, and medicine of fluorescent nanomaterials and related systems: new insights into biosensing, bioimaging, genomics, diagnostics, and therapy. Chemical Reviews, 114(12): 6130-6178.

4.      Siddiqui, M.R., AlOthman, Z.A. and Rahman, N., 2017. Analytical techniques in pharmaceutical analysis: A review. Arabian Journal of Chemistry, 10: S1409-S1421.

5.      Bakeev, K.A. ed., 2010. Process analytical technology: spectroscopic tools and implementation strategies for the chemical and pharmaceutical industries. John Wiley and Sons.

6.      Barrett, D.M., Beaulieu, J.C. and Shewfelt, R., 2010. Color, flavor, texture, and nutritional quality of fresh-cut fruits and vegetables: desirable levels, instrumental and sensory measurement, and the effects of processing. Critical reviews in food Science and Nutrition. 50(5): 369-389.

7.      Rantanen, J. and Khinast, J., 2015. The future of pharmaceutical manufacturing sciences. Journal of Pharmaceutical Sciences. 104(11): 3612-3638.

8.      Revathi, R., Naga, S.P.V., Rao, M.K., Ethiraj, T. and Rajarajan, S., 2012. Development and validation of RP-HPLC method for content analysis of Didanosine in dosage form. Asian Journal of Pharmaceutical Analysis. 2(4): 118-121.

9.      Haque, A., Shahriar, M., Parvin, M.N. and Islam, S.M. Validated RP-HPLC method for estimation of ranitidine hydrochloride, domperidone and naproxen in solid dosage form. Asian Journal of Pharmaceutical Analysis. 2011; 1(3): 59-63.

10.   Pardeshi, P.J., Gaware, V.M. and Dhamak, K.B. Development and Validation of RP-HPLC Method for the Estimation of Bilastine from bulk and Formulation. Asian Journal of Pharmaceutical Analysis. 2020; 10(2): 109-111.

11.   Rodriguez, E.L., Poddar, S., Iftekhar, S., Suh, K., Woolfork, A.G., Ovbude, S., Pekarek, A., Walters, M., Lott, S. and Hage, D.S. Affinity Chromatography: A review of trends and developments over the past 50 years. Journal of Chromatography B. 2020; 1157: 122332.

12.   Žuvela, P., Skoczylas, M., Jay Liu, J., Ba̧czek, T., Kaliszan, R., Wong, M.W. and Buszewski, B. Column characterization and selection systems in reversed-phase high-performance liquid chromatography. Chemical Reviews. 2019; 119(6): 3674-3729.

13.   Haque, A., Shahriar, M., Parvin, M.N. and Islam, S.M. Validated RP-HPLC method for estimation of ranitidine hydrochloride, domperidone and naproxen in solid dosage form. Asian Journal of Pharmaceutical Analysis. 2011; 1(3): 59-63.

14.   Chavda, N. and Kumar, S. A Review article on Analytical Method Development for the combination of Azelnidipine and Telmisartan. Asian Journal of Pharmaceutical Analysis. 2021; 11(3): 227-234.

15.   Raul, S.K., Padhy, G.K., Krishna, P.R. and Priya, B.U.M. UV Spectrophotometric method development and validation for the estimation of valsartan in bulk and pharmaceutical dosage form. Asian Journal of Pharmaceutical Analysis. 2016; 6(3): 147-150.

16.   Sebaiy, M.M., El-Shanawany, A.A., El-Adl, S.M., Abdel-Aziz, L.M. and Hashem, H.A. Rapid RP-HPLC method for simultaneous estimation of norfloxacin and tinidazole in tablet dosage form. Asian Journal of Pharmaceutical Analysis. 2011; 1(4): 79-84.

17.   Sivagami, B., Nagendramma, D., Chandrasekar, R. and Babu, M.N. RP-HPLC method development and validation for the estimation of seratrodast in bulk and pharmaceutical dosage form. Asian Journal of Pharmaceutical Analysis. 2018; 8(2): 96-102.

18.   Swapna, J., Madhu, C., Srivani, M., Sumalatha, M., Nehalatha, Y. and Anusha, Y. Analytical method development and method validation for the simultaneous estimation of metformin hydrochloride and pioglitazone hydrochloride in tablet dosage form by RP-HPLC. Asian Journal of Pharmaceutical Analysis. 2012; 2(3): 85-89.

19.   Vanaja, N., Preethi, C., Manjunath, S.Y. and Pal, K. Method development and validation for simultaneous estimation of telmisartan and chlorthalidone by RP-HPLC in pharmaceutical dosage form. Asian Journal of Pharmaceutical Analysis. 2015; 5(4): 171-177.

20.   Kumaraswamy, G., Ravindra, N. and Jyothsna, B. Development and validation of a reversed-phase HPLC method for simultaneous determination of Aspirin, Atenolol and Amlodipine in capsules dosage forms. Asian Journal of Pharmaceutical Analysis. 2014; 4(3): 116-120.

21.   Galante, R.N., Visalli, A.J. and Grim, W.M. Stabilized normal‐phase high‐performance liquid chromatographic analysis of aspirin and salicylic acid in solid Pharmaceutical Dosage forms. Journal of Pharmaceutical Sciences. 1984; 73(2): 195-197.

22.   Shah, D.A., Bhatt, K.K., Mehta, R.S., Shankar, M.B., Baldania, S.L. and Gandhi, T.R. Development and Validation of a RP-HPLC Method for Determination of Atorvastatin Calcium and Aspirin in a Capsule Dosage Form. Indian Journal of Pharmaceutical Sciences. 2007; 69(4).

23.   United States Pharmacopoeia, National Formulary, 24th ed. Rockville, MD: Pharmacopeial Convention; 2000: 161.

24.   The Indian Pharmacopoeia. Vol. 1. Ghaziabad: The Indian Pharmacopoeia Commission. 2007: 745.

25.   United States Pharmacopoeia- National Formulary, Toronto, Ontario:TheOfficialCompendiaofStandards,WebcomLimited;2005

26.   Cetinkaya, A., Kaya, S.I. and Ozkan, S.A. An overview of the current progress in green analytical chemistry by evaluating recent studies using greenness assessment tools. TrAC Trends in Analytical Chemistry. 2023; 17330.

27.   Evans, A.M., O’Donovan, C., Playdon, M., Beecher, C., Beger, R.D., Bowden, J.A., Broadhurst, D., Clish, C.B., Dasari, S., Dunn, W.B. and Griffin, J.L. Dissemination and analysis of the quality assurance (QA) and quality control (QC) practices of LC–MS based untargeted metabolomics practitioners. Metabolomics. 2020; 16(10): 113.

28.   Aithal, A. and Aithal, P.S. Development and validation of survey questionnaire and experimental data–a systematical review-based statistical approach. International Journal of Management, Technology, and Social Sciences (IJMTS). 2020; 5(2): 233-251.

29.   Gillings, N., Todde, S., Behe, M., Decristoforo, C., Elsinga, P., Ferrari, V., Hjelstuen, O., Peitl, P.K., Koziorowski, J., Laverman, P. and Mindt, T.L. EANM guideline on the validation of analytical methods for radiopharmaceuticals. EJNMMI Radiopharmacy and Chemistry. 2020; 5(1): 1-29.

30.   Salapaka, A., Bonige, K.B., Korupolu, R.B., K, C.R., N, S., Sharma, H.K. and Ray, U.K. A new stability indicating reverse phase high performance liquid chromatography method for the determination of enantiomeric purity of a DPP‐4 inhibitor drug linagliptin. Electrophoresis. 2019; 40(7): 1066-1073.

31.   Rabti, H., Amrane, M., Lalaouna, A., Flilissa, A. and Benguerba, Y. Optimization and validation of a bioanalytical HPLC–UV technique for simultaneous determination of underivatized phenylalanine and tyrosine in the blood for phenylketonuria diagnosis and monitoring. Biomedical Chromatography, 2023; p.e5758.

32.   Holton, E. and Kasprzyk-Hordern, B. Multiresidue antibiotic-metabolite quantification method using ultra-performance liquid chromatography coupled with tandem mass spectrometry for environmental and public exposure estimation. Analytical and Bioanalytical Chemistry. 2021; 413: 5901-5920.

33.   Zwanenburg, A. Radiomics in nuclear medicine: robustness, reproducibility, standardization, and how to avoid data analysis traps and replication crisis. European Journal of Nuclear Medicine and Molecular Imaging. 2019; 46(13): 2638-2655.

34.   Selvamani, V. Stability studies on nanomaterials used in drugs. In Characterization and biology of nanomaterials for drug delivery 2019 ;425-444.

 

 

 

 

Accepted on 27.04.2024           © RJPT All right reserved

Research J. Pharm. and Tech 2024; 17(8):3963-3968.