Development, Optimization and Evaluation of Thermo-responsive In-situ Nasal Gel of Levodopa and Entacapone for Brain-targeted Drug Delivery

 

K. M. Bhandari, P. L. Pingale*, S. V. Amrutkar

Department of Pharmaceutics, Gokhale Education Society’s Sir Dr. M. S. Gosavi College of Pharmaceutical Education and Research, Nashik, Maharashtra – 422005, India.

 

ABSTRACT:

Parkinson's disease is a neurodegenerative condition that includes mobility problems like decreased function, trouble with movements, stiffness, and shivering.  Levodopa is the medicine of preference to cure Parkinson's; however, it has limited CNS absorption due to substantial degradation in the periphery by aromatic amino acid decarboxylase or catechol-O-methyltransferase. As a result, a peripheral amino acid decarboxylase blocker, Carbidopa, and a catechol-O-methyltransferase inhibitor, Entacapone, are given along with Levodopa. Hence, the presented work's goal was to develop a thermoreversible in-situ nasal gel showing nose-to-brain administration of Levodopa and Entacapone to enhance CNS absorption and limit the enzymatic breakdown of Levodopa in the periphery. To formulate the in-situ nasal gels, Poloxamer-407, Hydroxypropyl methylcellulose, and Carbopol-934 were used as thermos-responsive and gel-forming polymers in varying ratios. The gels containing drugs Levodopa and Entacapone were prepared by the cold technique and optimized using 3² factorial design. Gel strength, drug content, bioadhesion potential, pH, rheology, gelation period and temperature, ex-vivo permeation, kinetic study, and stability study were conducted, and the optimum batch was finalized using optimization, in-vitro study, and stability data. At intranasal temperature, the preparations gelled, while the gelling period was found to be shorter as compared to the mucociliary clearance rate. Because of higher gel strength and bioadhesion, the intranasal retention was observed to be longer. During the ex-vivo trials on the goat’s nasal cavity, the intranasal gel preparations exhibited 98.8% absorption of Levodopa and 97.9% absorption of Entacapone within 12 hours. The kinetic plots obtained indicated that the drug release follows Higuchi and Zero-order models. Also, the results of the stability study showed that the optimum batch of in-situ gel was stable and safe throughout the study. The aim of this work was achieved by successful brain-targeted drug delivery with enhanced CNS absorption, thereby preventing first-pass metabolism or peripheral enzymatic degradation of Levodopa. Despite being in its early stages, the in-situ gelation technique showed great promise as a novel foundation holding the potential to manage Parkinson’s; however, more in-vivo research is needed to support these findings.

 

 

INTRODUCTION: 

Parkinson's disease (PD) and Alzheimer's disease (AD) are the largest global impairments, causing more hospitalizations and extended treatment than other illnesses. PD is the second-highest frequent age-related motor neuron illness, affecting 10 million people globally^1,2, leading to symptoms like tremors, bradykinesia, stiffness, and ocular and smell dysfunction^3-6. The blood-brain barrier makes it difficult for medicines, enzymes, or proteins to penetrate the brain effectively. The olfactory route is being studied for targeted CNS administration of medications with localized therapeutic actions, as it has a rich capillary bed, excellent porosity, and minimal metabolic activity. Systemic delivery of vitamins, hormones, chemotherapeutic agents, and stem cells has already been studied via the intranasal approach^7,8.

 

The BBB acts as a physical barrier for drug transport to the CNS due to the brain’s cell covering and reduced macropinocytosis. Traditional methods are ineffective, leading to new drug distribution alternatives. Intranasal channels offer easy access to the brain, and in-situ polymer gels have gained interest for regulated drug discharge. Bio-adhesives can help increase drug absorption by concentrating the gel in specific areas of the nose passage. Effective nose-to-brain drug transport involves olfactory and trigeminal nerve pathways, with paracellular and transcellular transport being the two major mechanisms^9-11.

 

The rationale of this research is to directly target the drug to the brain via nose-to-brain pathway; reducing its peripheral degradation and side effects and enhancing targeted action.

 

MATERIALS AND METHODS:

Aim:

To develop, optimize, and evaluate the thermoreversible in-situ nasal gel of Levodopa and Entacapone for brain-targeted drug delivery in Parkinson’s disease.

 

Materials:

Levodopa and Entacapone were obtained as gift samples from Alkem Laboratories Ltd., Taloja, Aurobindo Pharma Ltd., Goa, and Poloxamer-407 from Glenmark Pharmaceuticals Ltd., Nasik, India. Other analytical grade polymers and excipients included HPMC K-100, Carbopol-934, PEG-200, benzalkonium chloride, dimethyl sulfoxide (DMSO), phosphate buffer, and distilled water.

 

Methods:

Drug-excipient compatibility study:

The compatibility between APIs (Levodopa, Entacapone) and polymers (Poloxamer-407, HPMC K-100) was determined by Differential Scanning Calorimetry (DSC) and Fourier Transform Infrared (FTIR) spectroscopy.

 

DSC:

In the punctured DSC aluminum pan, approximately 1 to 2 mg of drugs and a combination of drugs and polymers were examined separately at temperatures 40-350°C at a heating velocity of 10°C/min with a standard pan.

 

FTIR:

The drugs, polymers, and their physical combination were examined separately from 4000 - 400 cm^-1. Mixture peaks at 1800 - 600 cm^-1 were contrasted with drug peaks.

 

Formulation and optimization of in-situ gel:

Nine in-situ nasal gels of Levodopa and Entacapone were developed and optimized using 3² factorial design, using various Poloxamer-407 and HPMC K-100 polymeric concentrations. Samples were prepared using the cold method and tested for pH, viscosity, gelation time, temperature, spreadability, mucoadhesive gel strength, drug content, ex-vivo perfusion, and stability¹². Table 1 illustrates the composition and role of ingredients used in the formulation of an in-situ gel wherein HPMC: hydroxypropyl methylcellulose, PEG: polyethylene glycol, DMSO: dimethyl sulfoxide, PBS: phosphate buffer solution, DW: distilled water, q.s.: quantity sufficient.

 

Evaluation of in-situ gel:

In-vitro experiments tested in-situ gel properties, including pH, rheology, spreadability, bioadhesive strength, API concentration, gelling temperature, and gelling time, determining the optimal batch for optimization.

 

pH measurement:

The produced nasal gels (1 gm) were diluted with distilled water, and the pH was measured using a calibrated digital pH meter¹³.

 

Rheology study/viscosity measurement:

The rheology of in-situ gels was examined using a Brookfield viscometer DV-I, recording mean viscosity at 25 ± 0.5°C and 37 ± 0.5°C in triplicate at 100 rpm¹⁴.

 

Table 1: Optimization batches developed by 3² factorial design

Composition (% w/v)	Role	F1	F2	F3	F4	F5	F6	F7	F8	F9
Levodopa	Anti-Parkinson drug	1	1	1	1	1	1	1	1	1
Entacapone	COMT inhibitor	1	1	1	1	1	1	1	1	1
Poloxamer-407	Thermo-responsive polymer	15	15	15	17.5	17.5	17.5	20	20	20
HPMC K-100	Mucoadhesive polymer	0.25	0.5	0.75	0.25	0.5	0.75	0.25	0.5	0.75
Carbopol-934	pH-sensitive gelling agent	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1
PEG-200	Penetration enhancer	2	2	2	2	2	2	2	2	2
Benzalkonium chloride	Anti-microbial agent	0.01	0.01	0.01	0.01	0.01	0.01	0.01	0.01	0.01
DMSO	Co-solvent	25	25	25	25	25	25	25	25	25
PBS (pH 6)	Co-solvent and pH modifier	25	25	25	25	25	25	25	25	25
DW (q.s.)	Solvent	100	100	100	100	100	100	100	100	100

 

Gelation time:

The gelation time was determined by agitating a cold solution with a specified Poloxamer concentration using a magnetic stirrer, recording the moment the magnetic bead stopped rotating, and repeated in triplicates for different concentrations and gelling temperatures^15,16.

 

Gelation temperature:

The gelation temperature is the temperature at which a solution's characteristics change from sol to gel. A 10 ml container filled with cold Poloxamer solution was raised by 2°C, achieving gelation when inclined to 90⁰^15,16.

 

Spreadability Study:

The spreadability index was determined by analyzing the gel’s ‘slip’ and ‘drag’ features, and the weight and duration of the upper slab's detachment from the lower slab were recorded using a fabricated spreadability apparatus¹⁷.

 

Muco-adhesive gel strength (modified balance method):

The study tested ex-vivo muco-adhesive strength using a fresh sheep intranasal mucosal layer. The surface was cleaned, and a 1 cm tissue was mounted on a watch glass. The muco-adhesive strength was computed in dyne/cm square¹⁸.

 

Drug content (UV-VIS method):

The drug content was analyzed using a Shimadzu UV-1800 dual-beam spectrophotometer at 240 and 309nm in triplicate after transferring 1gm of gel to a 10ml volumetric flask¹⁴.

 

Kinetic studies:

Statistical modeling optimizes pharmaceutical products by developing kinematic concepts to represent total API release in formulations. Changes in composition affect drug distribution and in-vivo function, making research easier through easier methods¹⁹. These studies involve various statistical designs like multivariate analysis of variance¹⁸, zero-order design^21,22, first-order design^23,24, Higuchi design^25,26, and Korsmeyer-peppas design⁴.

 

Perfusion/ex-vivo study/Franz-diffusion study:

The Franz-diffusion apparatus was used to evaluate drug perfusion ex-vivo. A sheep nasal membrane was processed using phosphate buffer solution and locked securely. Drug concentration was tested at 240nm and 309nm after dilutions. The release statistics in Zero Order and Higuchi graphs were used to identify drug distribution mechanics. The accuracy of the fit was assessed using MS Excel-2003. The release rate constant and regression coefficients were computed using a linear regression technique^27,28.

 

Stability study (accelerated stability procedure):

A three-month stability test was conducted on an optimum batch (F5) under elevated, room temperature, and frozen conditions of 40±2°C with 75±5% RH, 25 ±2°C with 60±5% RH, and 5±3°C with ambient RH respectively; with aliquots examined in triplicate for various parameters after 0, 1, 2, and 3 months, and averages reported²⁹.

 

RESULTS:

Drug-excipient compatibility study:

DSC:

The study found that the endothermic peaks of Levodopa and Entacapone in a drug-polymer mixture are visible independently, indicating no interaction between drugs and polymers.

 

FTIR:

All important functional group peaks in the FTIR spectra of the mixture are visible, indicating drug-polymer compatibility.

 

Formulation and optimization of in-situ gel:

The development of an in-situ nasal gel was conducted using two methods: “Cold” and “Hot.” The cold method was found to be more successful and stable. The optimization was done using a multilevel categorical linear regression method and an ANOVA study in Stat-Ease 360 software. The optimization of F1 to F9 batches with varying levels of Poloxamer-407 and HPMC K-100 was determined using 3² factorial design, and the F5 batch of an in-situ gel with 17.5% Poloxamer-407 and 0.5% HPMC K-100 was found to be the optimum batch.

 

Table 2: 3² Factorial design with factors and responses

Std	Factor A: HPMC (%)	Factor B: Poloxamer (%)	Response 1: Viscosity (cP)	Response 2: Gelation temperature (℃)	Response 3: Spreadability (gm.cm/sec)
F1	0.25	15	138.1±5.22	34±0.2	33.5±0.5
F2	0.5	15	141.34±6.02	33.5±0.4	33±0.2
F3	0.75	15	155.44±6.71	32±0.3	33±0.6
F4	0.25	17.5	168.56±7.22	33±0.4	30.5±0.1
F5	0.5	17.5	175.02±7.98	32.5±0.2	30±0.5
F6	0.75	17.5	187.12±8.14	32±0.2	29±0.2
F7	0.25	20	194.14±8.54	31.5±0.3	28.5±0.6
F8	0.5	20	211.78±8.83	31±0.4	27.5±0.4
F9	0.75	20	233.84±9.17	30±0.2	25±0.8

 

Figure 1: Surface response, contour, and perturbation plot of response 1 (viscosity)

 

Response 1 viscosity:

The study found significant results with an F-value and P-value of 189.47 and <0.0500, respectively. The Corrected R² was close to the Expected R² of 0.9536, with a variation of <0.2. The Adeq Precision signal-to-noise proportion was 37.211, indicating good output. The 3D surface response plot, contour plot, and perturbation plot showed the effect of varying polymeric concentrations on gel viscosity. HPMC played a major role in determining the optimum gel viscosity, while Poloxamer also had a significant effect. The perturbation plot indicated interference between polymers in determining gel viscosity, with 17.5% Poloxamer having more effect than 0.5% HPMC (Figure 1).

 

Response 2 gelation temperature:

The response to the gelation process was significant with an F-value and P-value of 36.13 and <0.0500, respectively. The Corrected R² was close to the Expected R2 of 0.8088, with a deviation of <0.2. The signal-to-noise proportion was 16.613 on Adeq Precision. The 3D surface response plot, contour plot, and perturbation plot showed the effect of varying polymeric concentrations on the gelation temperature. Both Poloxamer and HPMC played a significant role in determining the gelation temperature. The contour plot suggested different batches with different polymeric ratios, while the perturbation plot showed the interference of polymers in determining the gelation temperature. Both lines are adequately inclined, suggesting that both 17.5% Poloxamer and 0.5% HPMC have an equal effect on gelation temperature (Figure 2).

 

Response 3 spreadability: The response of a gel was found to be significant with a F-value of 160.83 and a P-value of <0.0500. The Corrected R2 was close to the Expected R² of 0.9615, with a deviation of <0.2. The signal-to-noise proportion of 32.863 on Adeq Precision indicated good output. The 3D surface response plot, contour plot, and perturbation plot showed the effect of varying polymeric concentrations on the gel’s spreadability. HPMC played a major role in determining the optimum gel’s spreadability, while 17.5% Poloxamer had a more significant effect. The perturbation plot signifies the interference of both the polymers in deciding the spreadability of the gel. The more inclined blue line suggests that 17.5% Poloxamer has more effect on spreadability over 0.5% HPMC (Figure 3).

 

 

Figure 2: Surface response, contour, and perturbation plot of response 2 (gelation temperature)

Figure 3: Surface response, contour, and perturbation plot of response 3 (spreadability)

 

Desirability:

Desirability is a variable that has a score of unity at the aim and zero outside the limits. The optimization finds the optimal location for the desirability score³⁰. The desirability obtained from the kinetic study data of the optimum F5 batch was found to be equal to unity.

 

Overlay / graphical optimization plot:

It creates one figure that highlights the “perfect zones” for meeting response requirements. It's often employed to demonstrate the system's failing boundaries³⁰. The overlay / graphical optimization plot obtained from the kinetic study data of the optimum F5 batch indicated the permissible component parameters with a vibrant yellow color.

 

Evaluation of in-situ gel:

The in-situ gels were examined for various evaluation parameters to test the effectiveness and stability, and to decide the best batch utilizing selective findings for optimization.

 

pH measurement:

The olfactory cavity typically has a pH of 4.5-6.5³¹, but it can tolerate substances with a pH of 3-10. Lysozyme, essential for killing microorganisms in acidic conditions, is ineffective at basic pH, making the intranasal mucosa vulnerable to pathogen attack. A phosphate buffer solution was used to regulate pH in preparations, resulting in results between 5.6±0.2 to 6.5±0.2, indicating they will not irritate the nasal epithelium.

 

Rheology study:

The viscosity of gels was assessed at 25℃ and 37℃ intranasal passage temperature. Results showed doubled viscosity readings at 37℃, with higher polymeric concentrations resulting in increased viscosity. Gel forms had doubled viscosity compared to sol forms, resulting in successful in-situ gel formation at 37℃ in all batches, making viscosity a major independent factor for formulation selection.

 

Gelation time:

Gelation time is the duration a thermoreversible solution takes to change from a sol form to a gel at a specific temperature. All formulae took 1.5-5.02 minutes to gel, with the optimum batch taking 3 minutes. Poloxamer-407, a thermo-responsive material, causes gel formation due to solubility shifts. However, all formulas had a gelation period less than 5minutes, eliminating the risk of composition clearance due to MCC.

 

Gelation Temperature:

Gelation temperature refers to the temperature at which the sol state changes to the gel state. The biological temperature of the olfactory passage and the MCC rate (21 mins) limit the operational limitations for intranasal in-situ gel distribution³¹. All samples’ gelling temperatures ranged from 30±0.2 to 34±0.2°C. Batches F1 to F3 gelled at temperatures over 32±0.3°C, while batches F4 to F6 gelled at 32±0.3-33±0.4°C. The optimal batch was F5, with a transition temperature of 32.5±0.2°C.

 

Poloxamer-407 is a reversible thermally gelling polymer made of polypropylene oxide and polyethylene oxide. It gels due to enhanced solvation and hydrogen binding at lower temperatures. At higher temperatures, the copolymer’s aqueous strands desolvate, promoting hydrophobic connections between polyoxypropylene structures. HPMC K-100, a non-thermos-responsive material, increases gelation when used as a bioadhesive. Entrapment of HPMC K-100 with Poloxamer-407 or hydrogen bridges may reduce gelation temperature. Table 3 provides mean pH, viscosity, gelation period, and gelation temperature values for F1 to F9 gel formulations.

 

 

Table 3: pH, viscosity, gelation time, and temperature of F1 to F9 batches

Batches	Results of various Parameters
	pH	Viscosity (cP)		Gelation Time (Mins)	Gelation Temp. (℃)
		25℃	37℃
F1	6.3±0.2	138.1±5.22	263.5±0.82	6±0.02	34±0.2
F2	5.7±0.2	141.3±6.02	289.2±9.96	7±0.04	33.5±0.4
F3	5.6±0.2	155.4±6.71	309.1±10.1	2.5±0.03	32±0.3
F4	6.1±0.2	168.5±7.22	326.5±12.5	5±0.02	33±0.4
F5	5.9±0.2	175.0±7.98	347.3±14.2	3±0.01	32.5±0.2
F6	6±0.2	187.2±8.14	364.7±11.7	4±0.04	32±0.2
F7	6.4±0.2	194.1±8.54	388.0±16.3	2±0.03	31.5±0.3
F8	5.8±0.2	211.7±8.83	432.4±7.86	3.5±0.02	31±0.4
F9	6.5±0.2	233.8±9.17	471.9±9.10	1.5±0.04	30±0.2

 

Spreadability study:

The semisolid’s spreadability is a key factor in its resilience, with F1 to F9 samples having a range of 25±0.8 to 33.5±0.5gm.cm/sec. The optimum batch (F5) has a spreadability of 30±0.5gm.cm/sec. The spreadability of the gels decreases with increasing polymeric concentration due to the viscoelastic nature of Poloxamer-407, HPMC K-100, and Carbopol-934, which act as a mucoadhesive gelling matrix (Table 4).

 

Muco-adhesive gel strength:

This test examines the bioadhesive properties of bioadhesive substances like Poloxamer-407, HPMC K-100, and Carbopol-934 in olfactory passages. The bioadhesive ability of the gel increases with the bioadhesive substance’s level but is inversely related to the gelling period and temperature. Batches with higher concentrations of Poloxamer-407 or HPMC K-100 resulted in more mucoadhesive gel strength and a lower gelation period or temperature. The gel strength values ranged from 3852±74.22 to 5270±308.09 dyne/cm², with an optimum value of 4898±132.82 dyne/cm² for the optimized (F5) batch. Poloxamer-407 plays a crucial role in strengthening the gel by promoting hydrophobic connections between polyoxypropylene structures, resulting in gelation and increased mucoadhesive properties.

Drug content:

The percent API concentrations of all intranasal in-situ gel preparations were obtained by UV analysis of the diluted gel samples and were observed to be between 95.9±0.32 to 98.8±0.50% for Levodopa and 95.0±0.92% to 97.9±0.35% for Entacapone (Table 4). The optimized batch (F5) showed a maximum drug content of 98.8±0.50% for Levodopa and 97.9±0.35% for Entacapone. The average spreadability, gel strength, and % drug content for F1 to F9 in-situ gel formulations are explained in Table 4.

 

Perfusion/ex-vivo study:

Ex-vivo penetration tests were conducted on sheep intranasal tissue, determining the percentage of API penetrated. The experiments were conducted in the Franz diffusion unit for 12 hours with the drug in the donor chamber and phosphate buffer in the recipient chamber. The drug penetration was biphasic, with a speedy stage initially and a stable stage thereafter. Factors such as Poloxamer, PEG-200, and DMSO contributed to the initial quick release, and bioadhesive substances increased API penetration.

 

The percent cumulative drug release of Levodopa from F1 to F9 batches in 720 mins is depicted in Table 5.

 

Table 4: Spreadability, gel strength, and percent drug content of F1 to F9 batches

Batches	Results of various Parameters
	Spreadability (gm.cm/sec)	Gel Strength (dyne/cm2)	Drug Content (%)
			Levodopa	Entacapone
F1	33.5±0.5	3852±74.22	96.2±0.54	95.4±0.05
F2	33±0.2	3924±85.54	97.6±0.06	96.7±0.12
F3	33±0.6	4232±92.68	96.7±0.96	95.9±0.54
F4	30.5±0.1	4546±106.52	97.4±0.46	96.6±0.08
F5	30±0.5	4898±132.82	98.8±0.50	97.9±0.35
F6	29±0.2	5006±212.04	97.1±0.35	96.6±0.09
F7	28.5±0.6	5114±246.35	96.8±0.88	95.5±0.67
F8	27.5±0.4	5183±266.31	95.9±0.32	95.2±0.31
F9	25±0.8	5270±308.09	98.0±0.15	95.0±0.92

 

Table 5: Percent cumulative drug release of Levodopa in perfusion study

Time (Mins)	Cumulative drug release of Levodopa (%)
	F1	F2	F3	F4	F5	F6	F7	F8	F9
0	0	0	0	0	0	0	0	0	0
15	11.7	14.5	17.4	20.1	22.5	8.9	5.4	2.8	0
30	17.5	20.3	23.2	24.9	29.2	14.7	11.2	8.9	5.6
60	25.9	28.7	31.6	34.3	37.4	26.1	20	16.3	13.2
120	32.3	35.1	38	41.7	44	29.5	27.9	23.6	20.4
240	46.1	49.9	51.8	55.5	58.7	43.3	40.6	37.4	34.2
360	54	57.8	60.7	63.4	66.9	51.2	48.7	45.4	44.1
480	61.3	66.7	69.4	72.6	75.1	60.2	57.5	54	51.9
600	75.8	78.9	81.3	84.1	87.6	72.2	69	64.7	63.4
720	86.7	88.5	92.6	95.2	98.8	83.9	80	77.1	74.4

 

Table 6: Percent cumulative drug release of Entacapone in perfusion study

 

The percent cumulative drug release of Entacapone from F1 to F9 batches in 720 minutes is depicted in Table 6.

 

Kinetic studies:

The study used MS Excel-2003 for a kinetic best-fit model study. Model-based plots were used to determine the required values for the cumulative drug release. The R² and n values were obtained from these plots, which measure the efficiency of matching up regression projections. The zero-order and Higuchi models with R² values of 0.9926 and 0.9265, respectively (Figure 4), were found to be the best-fit models for optimized in-situ gel, indicating controlled drug release over a prolonged period.

 

 

Figure 4: Higuchi and zero-order plots of kinetic study data

 

Table 7 highlights the pharmacokinetic study data of the optimum batch, indicating time in hours, percent cumulative drug released, and percent drug remaining at respective time intervals, and the square root of time. The values of time, the square root of time, and percent cumulative drug release aid in plotting zero-order and Higuchi kinetic plots. These plots indicate that the formulation fits best in the zero-order and Higuchi models and hence follows a controlled drug release mechanism.

 

Table 7: Kinetic study data of optimized (F5) batch of in-situ gel

Time (Hr)	Cumulative drug release (%)	Drug remaining (%)	Square root time
0	0	100	0
15	18.7	81.3	3.872
30	25.5	74.5	5.477
60	31.9	68.1	7.745
120	40.3	59.7	10.954
240	59.1	40.9	15.491
360	66	34	18.973
480	74.2	25.8	21.908
600	86.6	13.4	24.494
720	98.8	2.2	26.832

 

Stability study:

The stability testing of optimized in-situ gel (F5) for 3 months showed no significant changes in pH, gelation parameters, bioadhesive capacity, rheological and physiological characteristics, content uniformity, and ex-vivo penetration. Table 8a and 8b shows the results obtained after stability assessment of the optimized batch. The results showed that the optimal batch (F5) remained constant with appearance, pH, rheological characteristics, gelation time, and temperature remaining within permissible limits.

 

Table 8b illustrates the results obtained after stability assessment of optimized batch at varying temperature scales for 0-, 1-, 2-, and 3-month intervals. The results obtained show that the spreadability, mucoadhesive strength, and drug content of the optimum batch (F5) remain constant and within permissible limits.

Table 8a: Results of stability test of optimum (F5) batch

Storage duration	Stored at	Color	pH	Viscosity (cP)		Gelation time (Mins)	Gelation temperature (℃)
				At 25℃	At 37℃
0 Month	25℃	Saffron	5.9±0.02	175.0±6.87	347.3±10.54	3±0.04	32.5±0.3
1 Month	40℃	Saffron	5.8±0.02	178.9±5.64	350.5±12.75	4.5±0.02	33.5±0.4
	25℃	Saffron	5.9±0.02	174.5±7.10	345.2±15.51	3±0.03	32.5±0.2
	5℃	Saffron	5.9±0.02	173.6±1.21	343.7±17.32	4±0.04	33±0.5
2 Months	40℃	Saffron	5.8±0.02	180.4±6.08	352.2±18.14	3.5±0.02	34±0.1
	25℃	Saffron	5.9±0.02	175.6±4.75	347.0±14.02	2.5±0.01	32±0.4
	5℃	Saffron	6±0.02	173.9±8.12	345.8±11.69	4.5±0.02	33.5±0.3
3 Months	40℃	Saffron	5.9±0.02	183.5±3.55	358.6±13.30	4±0.03	32±0.2
	25℃	Saffron	5.9±0.02	176.3±4.14	350.2±19.40	2.5±0.04	31.5±0.3
	5℃	Saffron	6±0.02	175.8±5.32	352.1±10.66	5±0.01	31±0.5

 

Table 8b: Results of stability test of optimum (F5) batch

Storage duration	Storage condition	Spreadability (gm.cm/sec)	Gel strength (dyne/cm2)	Drug content (%)
				Levodopa	Entacapone
0 Month	25℃	30±0.5	4898±132.82	98.8±0.50	97.9±0.35
1 Month	40℃	28±0.4	4982±154.10	98.6±0.40	97.6±0.06
	25℃	30±0.5	4860±110.25	98.8±0.21	97.9±0.24
	5℃	32±0.2	4662±102.66	98.7±0.15	97.9±0.33
2 Months	40℃	28±0.4	4963±156.23	98.6±0.08	97.7±0.05
	25℃	30±0.5	4832±135.41	98.8±0.32	97.9±0.16
	5℃	34±0.1	4799±164.02	98.6±0.29	97.8±0.31
3 Months	40℃	26±0.3	4885±140.39	98.7±0.19	97.5±0.17
	25℃	30±0.5	4804±136.08	98.8±0.46	97.9±0.24
	5℃	35±0.3	4747±127.12	98.8±0.37	97.8±0.38

 

DISCUSSION:

This study aimed to develop a thermo-responsive in-situ nasal gel containing Levodopa and Entacapone, which could be used as a brain-focused drug transport strategy in Parkinson's disease (PD). The gel’s viscosity was directly related to its polymer concentration, with the main polymers Poloxamer-407, HPMC K-100, and Carbopol-934 being the main polymers. The pH-sensitive nature of Carbopol in combination with phosphate buffer (pH 6) helped maintain the nasal pH required for gelation. Intranasal drug transport has evolved as a potential mode of drug transport due to the intranasal cavity’s physiology and morphology. The drug’s bioavailability increased with the addition of permeation boosters like PEG. The thermo-responsive polymer Poloxamer-407 was critical in the sol-to-gel transition at 37°C, with the strongest influence at 17.5%. The gel’s-maintained consistency for a long time, allowing prolonged API delivery and rapid gelation. However, brain-targeting via the nasal route may be affected by physiological or formulation-related factors, such as MCC, nasal cavity environment, pathological conditions, dryness, or clogging due to sensitivity to excipients. Further pre-clinical and clinical trials are needed to confirm these studies. Despite its infancy, nose-to-brain drug delivery has great potential to increase drug absorption and bioavailability with reduced side effects.

 

LIST OF SYMBOLS AND ABBREVIATIONS:

AD	Alzheimer’s disease
ANOVA	Analysis of variance
ANS	Autonomic nervous system
API	Active pharmaceutical ingredient
BBB	Blood-brain barrier
CNS	Central nervous system
COMT	Catechol-o-methyl transferase
DMSO	Dimethyl sulfoxide
DSC	Differential scanning calorimetry
FTIR	Fourier transform infrared
HPMC	Hydroxy propyl methyl cellulose
MCC	Mucociliary clearance
PBS	Phosphate buffer solution
PD	Parkinson’s disease
PEG	Polyethylene glycol

 

CONFLICT OF INTEREST:

The authors have no conflicts of interest regarding this investigation.

 

ACKNOWLEDGMENTS:

Authors are thankful to support staff of research centre for their help in completion of this research work.

 

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Received on 26.04.2025      Revised on 22.08.2025 Accepted on 28.10.2025      Published on 01.07.2026 Available online from July 04, 2026 Research J. Pharmacy and Technology. 2026;19(7):2933-2941. DOI: 10.52711/0974-360X.2026.00418 © RJPT All right reserved
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