Department of Pharmaceutical chemistry, Channabasweshwar Pharmacy College (Degree),Latur.
A common treatment for moderate to severe Alzheimer's disease is the combination of memantine hydrochloride and donepezil hydrochloride. For bulk medications and pharmaceutical formulations to comply with regulations and maintain quality, analytical techniques that are accurate, precise, and stability-indicating are crucial. Because of its selectivity, repeatability, and applicability for stability investigations, reverse phase high performance liquid chromatography (RP-HPLC) continues to be the most often used method for their simultaneous estimation. Physicochemical factors, chromatographic optimization techniques, validation parameters in accordance with ICH Q2 (R1) guidelines, forced degradation studies, and applications in pharmaceutical quality control are all rigorously examined in this review. This article also discusses methods for overcoming the analytical difficulties brought on by memantine poor chromophore.
Alzheimer's disease (AD) is a gradual, irreversible neurological condition marked by functional decline, behavioural abnormalities, memory loss, and cognitive decline. Due to aging populations, it is the most prevalent cause of dementia globally and imposes a substantial public health burden. The main goals of pharmacological treatment for AD are to alleviate symptoms and limit the disease's progression via modifying neurotransmitter systems related to cognition. For moderate to severe stages of the illness, donepezil hydrochloride and memantine hydrochloride, two approved treatment medications, are frequently used either alone or in combination.
By stopping acetylcholine from being broken down in the synaptic cleft, donepezil hydrochloride, a centrally acting, reversible acetylcholinesterase inhibitor, improves cholinergic neurotransmission.[1]
Patients with mild to moderate Alzheimer's disease benefit most from it in terms of improved cognition and overall functioning. By preventing excessive calcium influx into neurons, memantine hydrochloride, an N-methyl-D-aspartate (NMDA) receptor antagonist, controls glutamatergic neurotransmission and lessens excitotoxicity neuronal damage. In advanced stages of Alzheimer's disease, where both glutamatergic dysregulation and cholinergic deficiencies contribute to disease pathology, the complementary mechanisms of action of these two medications justify their combined usage. Reliable, accurate, and validated analytical techniques for the simultaneous measurement of memantine and donepezil in pharmaceutical dosage forms and bulk pharmaceuticals are becoming more and more necessary due to the expanding clinical use of fixed-dose combos including both drugs.[2]
In order to guarantee the identification, potency, purity, and safety of pharmaceutical items, analytical method development is essential. Chromatographic techniques, especially Reverse Phase great Performance Liquid Chromatography (RP-HPLC), are widely used in the pharmaceutical sector because of their great sensitivity, selectivity, reproducibility, and adaptability for complicated combinations. There are particular analytical difficulties in developing an RP-HPLC method for the simultaneous measurement of donepezil hydrochloride and memantine hydrochloride. Memantine lacks a strong chromophoric group, making it a very water-soluble primary amine with low ultraviolet (UV) absorbance. This feature requires the employment of derivatization techniques to increase sensitivity or detection at lower wavelengths. On the other hand, donepezil is easily visible at higher wavelengths due to its aromatic moiety' considerable UV absorption.[3,4]Chromatographic settings, such as mobile phase composition, pH, column selection, flow rate, and detection wavelength, must be carefully optimized due to the variations in physicochemical features, such as polarity, pKa, and UV absorption characteristics. Validation is a necessary prerequisite to show that the analytical process is appropriate for its intended use, in addition to method development. Specificity, linearity, accuracy, precision, robustness, limit of detection (LOD), and limit of quantification (LOQ) are among the qualities that validated methods must establish in accordance with International Council for Harmonization (ICH) guidelines. For combination medicinal products, stability-indicating capacity is especially crucial for correct quantification in the presence of degradation products created under stress.[5]For routine quality control, stability testing, and regulatory compliance, a reliable RP-HPLC method for memantine hydrochloride and donepezil hydrochloride must be developed and validated. The objective of this review is to provide a thorough discussion of the analytical techniques, optimization factors, and validation needs related to RP-HPLC methodologies for the simultaneous estimation of these therapeutically important compounds.[6]
For the simultaneous measurement of memantine hydrochloride and donepezil hydrochloride in pharmaceutical dosage forms and bulk, a number of analytical techniques have been described; RP-HPLC is the most commonly used approach because of its sensitivity, accuracy, and repeatability.
A Bheema Naik, et. al. 2023 [1]
Using an Altima C18 column (4.6 mm × 150 mm, 5.0 µm), this work revealed a novel, quick, and sensitive RP-HPLC method for the simultaneous quantitative measurement of memantine and donepezil. A mobile phase consisting of buffer, methanol, and acetonitrile in the ratio of 65:35:10 (% v/v/v) at a flow rate of 1.0 mL/min was used to accomplish the chromatographic separation. A UV-visible detector operating at 234 nm was used for detection, which offered sufficient sensitivity for both analytes. With retention durations of 2.088 minutes for memantine and 6.068 minutes for donepezil, effective separation was achieved under ideal conditions, demonstrating good resolution in a brief analytical run time.
The invention of a straightforward, quick, accurate, and precise RP-HPLC technology that can be used for routine quality control analysis of memantine and donepezil in both approved pharmaceutical dosage forms and bulk medicines is highlighted in this paper. The method is beneficial for high-throughput analysis in pharmaceutical laboratories because to the short retention period for memantine and acceptable resolution from donepezil. Overall, the literature consistently demonstrates that RP-HPLC is a reliable and effective analytical tool for the simultaneous estimation of donepezil hydrochloride and memantine hydrochloride, with validated methods showing suitability for routine industrial applications and compliance with regulatory requirements.
Ram S. Sakhare, et.al. 2023 [2]
Using Design Expert® (11.0.0) modeling software for response surface methodology (RSM), this study used a Quality by Design (QbD) strategy to systematically optimize critical method parameters (CMPs) through a central-composite design (CCD). Using an Agilent C18 column (50 × 2.1 mm, 1.7 μM) and UV detection at 230 nm, chromatographic separation was accomplished. Acetonitrile and 0.01N potassium dihydrogen phosphate in a 55.2:44.8 v/v ratio, pumped at 1 mL/min, made up the optimum mobile phase. ICH Q2 (R1) recommendations were followed in the validation process, which confirmed linearity, precision, accuracy, specificity, and robustness. According to the study's findings, the QbD-based RP-HPLC method has enhanced method repeatability, robustness, and predictive performance, making it appropriate for regular pharmaceutical analysis of donepezil and memantine in dosage forms and bulk.
Syeda Noorain Amena, et.al. 2015 [3]
A stability-indicating RP-HPLC technique was devised and validated for the simultaneous analysis of donepezil hydrochloride and memantine hydrochloride in a recent study by the authors. Using a mobile phase made of sodium dihydrogen orthophosphate and acetonitrile in a 30:70 v/v ratio and supplied at a flow rate of 1 mL/min with UV detection, chromatographic separation was accomplished on a Hypersil BDS column (4.6 × 150 mm, 5 µm). Memantine and donepezil were found to have retention durations of 2.833 and 4.777 minutes, respectively, indicating successful separation in a brief analysis period. Recovery studies demonstrated great accuracy with 99.62% and 99.45% recovery, respectively, and the method demonstrated linearity throughout concentration ranges of 40–120 µg/mL for memantine and 20–60 µg/mL for donepezil.
There was little interference from excipients, as evidenced by the reported percentage purity of 98.5% for memantine and 98.6% for donepezil. With limits of detection of 3.69 µg/mL for memantine and 2.72 µg/mL for donepezil, as well as limits of quantification of 11.13 µg/mL and 8.25 µg/mL, respectively, the method also showed good sensitivity. Studies on stress degradation in acidic, basic, oxidative, thermal, and photolytic environments revealed no discernible deterioration for either medication. The authors came to the conclusion that the developed RP-HPLC method is appropriate for routine quality control and stability analysis of memantine and donepezil in bulk and dose forms since it is straightforward, sensitive, accurate, and acceptable as a stability-indicating approach.
Kamepalli Sujana, et. al. 2012 [4]
In order to estimate memantine (MEM) and donepezil (DPZ) simultaneously in the presence of their degradation products, the study looked into the creation of a straightforward, quick, accurate, precise, and affordable stability-indicating HPLC method using diode array detection (DAD). Using an Inertsil ODS column (4.6 × 250 mm, 5 µm) and a mobile phase made up of acetonitrile and 0.1% orthophosphoric acid in a 10:90 ratio at a flow rate of 1.0 mL/min, chromatographic separation was accomplished at 271 nm. Within 3.2 minutes, the medicines were successfully separated from all degradants using this approach. For both medications, linearity was seen across the 10–50 µg/mL concentration range. The method's accuracy, precision, robustness, limit of detection, and limit of quantification were all validated in accordance with ICH recommendations.
Studies on forced degradation, such as acid and base hydrolysis, oxidation, heat stress, and photolytic degradation by sunlight, revealed that both medications only slightly deteriorated in all stress scenarios. Overall, the study found that even in the presence of their degradation products, this stability-indicating approach is dependable, effective, and appropriate for routine quantification of memantine and donepezil in tablet formulations.
Hyun Tae Jang, et. al. 2016 [5]
The paper described the creation and validation of a straightforward, quick, accurate, precise, and an economical stability-indicating HPLC technique with diode array detection (DAD) for the simultaneous measurement of donepezil (DPZ) and memantine (MEM) in the presence of their degradation products. An Inertsil ODS column (4.6 × 250 mm, 5 µm) with a mobile phase of acetonitrile and 0.1% orthophosphoric acid in a 10:90 ratio, a flow rate of 1.0 mL/min, and a detection wavelength of 271 nm was used to accomplish chromatographic separation. In 3.2 minutes, the medicines were completely resolved from all degradants thanks to the technique. For both medications, linearity was seen in the concentration range of 10–50 µg/ml.
The method's accuracy, precision, robustness, limit of detection, and limit of quantification were all validated in accordance with ICH recommendations. Acid and base hydrolysis, oxidation, heat stress, and photolytic degradation under sunlight were among the forced degradation tests performed on the medication samples. The results showed that both pharmaceuticals only experienced minor degradation. Overall, the study found that even in the presence of possible degradation products, the suggested method is dependable, effective, and appropriate for routine quantification of memantine and donepezil in tablet formulations.
Joana A. Loureiro, et. al. 2024 [6]
A high-performance liquid chromatography (HPLC) approach with pre-column derivatization was developed and validated for the quantitative analysis of memantine hydrochloride encapsulated in nanoparticles (NPs), according to the study. 9-fluorenylmethyl chloroformate (FMOC) was used to derivatize memantine. Chromatographic separation produced a mean recovery of 92.9 ± 3.7% with a retention period of 11.393 ± 0.003 min. With intraday precision ranging from 3.6% to 4.6% and interday precision from 4.2% to 9.3%, the method was completely validated in accordance with ICH criteria, exhibiting appropriateness, specificity, linearity over 5–140 µg/mL, sensitivity, precision, accuracy, and robustness. Studies on stability at 4°C and -20°C revealed that memantine did not degrade for up to six months.
The technique for assessing the effectiveness of memantine encapsulation in lipid nanoparticles was straightforward, sensitive, specific, accurate, and precise. The ultimate result for the greenness assessment was 0.45, which indicates a modest level of environmental friendliness. According to the study, this approach may be used in the future to measure memantine in different nano formulations, which could enhance the assessment of medication delivery methods for the treatment of Alzheimer's disease.
Comparative Analysis of Chromatographic Conditions and parameters
|
Parameter |
A Bheema Naik, et.al 2023 |
Ram S. Sakhare, et.al. 2023 |
Syeda Noorain Amena, et.al. 2015 |
Kamepalli Sujana, et. al. 2012 |
Hyun Tae Jang, et. al. 2016 |
Joana A. Loureiro, et. al. 2024 |
|
Column |
Altima C18 (4.6mm × 150mm, 5.0 µm) |
Agilent C18 (50 x 2.1 mm, 1.7µm) |
Hypersil BDS (4.6 x 150 mm, 5μ) |
Agilent C8 (150mm x 4.6mm i.d., 3.5µm particle size) |
Inertsil ODS (4.6 X 250mm, 5µm) |
C18 Cortecs column (4.6 × 150 mm, 2.7 µm) coupled to a C18 Cortecs VanGuard (3.9 × 5 mm, 2.7 µm) |
|
Mobile Phase |
Buffer: Methanol: Acetonitrile (65:25:10% v/v/v)
|
0.01N Potassium dihydrogen phosphate: acetonitrile in 55.2:44.8 v/v proportion.
|
Sodium dihydrogen ortho phosphate: Acetonitrile (30:70v/v)
|
Buffer, water and Acetonitrile (50:5:45 v/v)
|
Acetonitrile with 0.1% orthophosphoric acid at a ratio of 10:90.
|
Ultrapure water with 0.02% TFA (A) and acetonitrile (B). Gradient: 0-5 min (50% B); 5-10 min (50-90% B); 10-12 min (90% B); 12-15 min (90-50% B); 15-20 min (50% B). |
|
Flow Rate |
1.0 ml/min |
1 ml/min |
1 ml/min |
? 0.8 ml/min |
1.0 ml/min |
1 mL/min |
|
Wavelength |
234 nm |
230 nm |
271 nm |
230nm |
271 nm |
265 nm |
|
Injection volume |
10 µL |
20 µL |
Not specified |
20uL |
20uL |
20uL |
|
RT (Memantine and Donepezil) |
Memantine: 2.088 min, Donepezil: 6.068 min |
Memantine: 2.481 min, Donepezil: 3.192 min |
Memantine HCl: 2.833 min, Donepezil HCl: 4.777 min |
Memantine HCl: 6.976 min Donepezil6.976 min |
3.2 min for both |
Memantine: 11.393 ± 0.003 min |
|
Run Time |
14 minutes |
4 minutes |
6 minutes |
Not specified |
3.2 minutes |
20 minutes |
For the simultaneous measurement of memantine and donepezil, the HPLC techniques described by A. Bheema Naik et al. (2023), Ram S. Sakhare et al. (2023), Syeda Noorain Amena et al. (2015), Kamepalli Sujana et al. (2012), Hyun Tae Jang et al. (2016), and Joana A. Loureiro et al. (2024). The majority of techniques use reversed-phase C18 columns, such as Altima C18, Hypersil BDS, Inertsil ODS, and Cortecs C18, suggesting that hydrophobic stationary phases are preferred for successful separation. However, an Agilent C8 column, which has somewhat less hydrophobicity, was utilized by Kamepalli Sujana et al. (2012).
The dimensions of the columns vary as well; Ram S. Sakhare et al. (2023) utilized shorter 50 × 2.1 mm (1.7 µm) columns for quick analysis, while Hyun Tae Jang et al. (2016) used longer 250 mm columns for better resolution. While Joana A. Loureiro et al. (2024) employed a gradient elution method using TFA-modified water and acetonitrile, offering greater control over peak separation and analyte elution, the mobile phase composition also varies greatly, including mixes of phosphate buffer with methanol or acetonitrile.
Additionally, there are differences between the approaches in operational parameters such flow rate, wavelength, injection volume, and run time. With the exception of Kamepalli Sujana et al. (2012), which employed 0.8 mL/min, the majority of studies maintain a flow rate of about 1.0 mL/min. The range of detection wavelengths, which reflect variations in the analytes' ideal UV absorption, is 230–271 nm. Injection volumes are typically 20 µL, although A. Bheema Naik et al. (2023) used 10 µL, and Syeda Noorain Amena et al. (2015) did not specify the value. Memantine and donepezil retention times vary greatly amongst investigations, ranging from roughly two to three minutes in quicker approaches (Ram S. Sakhare et al., Hyun Tae Jang et al.) to more than eleven minutes in the gradient method (Joana A. Loureiro et al., 2024).
As a result, depending on the method design, the entire run time can be as short as 3–4 minutes or as long as 20 minutes, illustrating the trade-off between quick analysis and improved chromatographic resolution.
CONCLUSION
In conclusion, there are discernible variations in chromatographic parameters such column type, mobile phase composition, and detection wavelength between the reported RP-HPLC techniques for the simultaneous quantification of Memantine and Donepezil. The majority of studies have favored C18 reversed-phase columns because of their potent hydrophobic interactions and capacity to effectively separate the analytes. However, the chromatographic performance is greatly affected by differences in column diameters, particle size, and the use of various organic solvents including acetonitrile and methanol. The experiments' detection wavelengths, which vary from 230 to 271 nm, show optimization based on the medications' UV absorption properties.Additionally, depending on the analytical goals, different methods have different operational conditions, such as flow rate, injection volume, retention time, and total run time. Some techniques use longer run durations and gradient elution systems to increase peak resolution and accuracy, while others provide quick analysis with shorter run periods of three to six minutes. Overall, every approach that has been described shows that Memantine and Donepezil may be separated reliably and effectively, demonstrating the versatility of RP-HPLC techniques in pharmaceutical analysis for obtaining precise, accurate, and repeatable results.
REFERENCES
Amol Naragude, Kedar Dongare, Amar Fulsundar, Vishal Kumbhar, Shraddha Belkunde, Recent Advances in RP-HPLC Analysis of Memantine Hydrochloride and Donepezil Hydrochloride, Int. J. of Pharm. Sci., 2026, Vol 4, Issue 4, 2295-2301 https://doi.org/10.5281/zenodo.19591624
10.5281/zenodo.19591624
