Formulation, Evaluation and Validation of Oral Film for The Treatment of Antimigraine by Using HPLC Method

Out of various methods, the oral route is the most preferred option for patients. Numerous pharmaceutical companies have focused their research on creating effective oral dosage alternatives for paediatric, geriatric, noncompliant or nauseous patients. Advancements in oral drug delivery have progressed from basic conventional tablets and capsules to modified release forms, oral disintegrating tablets and most recently oral films.  Notably, pain, which is the primary symptom of the condition, is not always the most distressing aspect for all patients at all times. Migraine is characterized by a series of distinct phases often overlapping which include the premonitory (prodromal), aura, pain, and postdrome phases. Improved recognition of these phases has led to the understanding of migraine as a network disorder involving multiple regions of the cortex, subcortex, and brainstem, which together generate a wide range of signs and symptoms. We will explore these regions in detail in the upcoming sections, as they show altered function and structure in individuals with migraine and in animal models of the condition.^[1]

Defination of Migraine   

A migraine is an intense headache characterized by pulsating, throbbing pain primarily on one side of the head. The headache phase of a migraine typically persists for several hours but can extend to an entire day. This pain often intensifies with physical activity, bright light, loud noises, and strong smells. 

Types of Migraine  

There are mainly two types: 

1. Migraine with aura (Classic migraine) 

2. Migraine without aura (Common migraine).  

Migraine Symptoms 

Changes in mood, trouble concentrating, sleep disturbances, fatigue, nausea, increased hunger, thirst and frequent urination, Muscle weakness, alterations in vision, ringing in the ear and heightened sensitivity to touch, sensitivity to light, sound and smells, Stiff neck, sensitivity to light or sound, difficulty concentrating, nausea and dizziness.

Diagnosis

1. When considering a migraine diagnosis

2. Establishing a migraine diagnosis

3. Focus on patient needs and education.^[2,3]

Oral Thin Film

They are also known as oral wafers. In recent years, oral thin films have emerged in the confectionery and oral care industries as oral strips. These innovative products have gained widespread acceptance among consumers for delivering vitamins and personal care items. Currently, oral films are recognized and validated technology for the systemic distribution of active pharmaceutical ingredients (APIs) in over-the-counter (OTC) drugs and are in the initial to mid-development phases for prescription medications. 

Fig 1: Oral film

Classification of Oral Films Consists of Three Types:  

• Flash release/ Fast dissolving films (Placed on the tongue). 

• Mucoadhesive melts away films (in the gingival or buccal region). 

• Mucoadhesive sustained release films (adhere to the buccal mucosa). 

ADVANTAGES 

1. There is no need for a specially trained individual. 

2. They are flexible, portable, making transportation and storage easier. 

3. Dosing is accurately administered. 

4. Enhanced stability and safety are achieved. 

5. Each film contains a precise quantity of medication. 

6. Rapid onset of action and improved bioavailability. 

7. They provide a mouth freshening effect. 

8. Patient adherence is improved. 

9. Each film ensures accurate dose administration compared to liquid forms like drops or syrups. 

10. Patients experiencing frequent vomiting, dysphagia, or motion sickness prefer this dosage form as they find it hard to swallow large volumes of water.  

11. These films can be produced through cost-effective.

DISADVANTAGES 

1. There are limitations on food and drink consumption. 

2. Medications that require lower doses may be administered. 

3. Special packaging is necessary for maintaining product stability and safety. 

4. The variety of polymers available is limited. 

5. The application of thin films can be constrained due to their low drug loading capacity when applying a less potent drug at high doses. 

6. Typically, thin films are hygroscopic, so special care is needed for prolonged storage. 

7. A larger surface area facilitates faster disintegration and dissolution in the oral cavity. ^{[4,5,6,7,8,9]}

Manufacturing or Formulation Techniques of Oral Film

A. Solvent casting method

B. Semi-solid casting method

C. Hot melt extrusion

D. Solid dispersion extrusion

E. Rolling method.  ^[10,11,12]

Aim And Objective

Aim: Formulation, Evaluation and Validation of oral film for the treatment of antimigraine by using HPLC method

Objective:

1. To carry out the preformulation study of drug and formulation.
2. To formulate the oral film.
3. To evaluate the oral film.
4. To study the thickness, folding endurance, disintegration time, content uniformity and In vitro study of oral film.
5. To check the stability of oral film.
6. Simple, precise, accurate and economical HPLC method can be developed.
7. To increase the drug bioavailability, safety, effectiveness.
8. Validation of method according to ICH guideline.

 Drug Profile

Fig 2:  Zolmitriptan

1) Synonyms /Chemical name:  Zolmitriptan, Zomig

2) IUPAC Name: (4S)-4-[[3-[2-(dimethylamino)ethyl]-1H-indol-5-yl]methyl]-1,3-oxazolidin-2-one

3) Molecular Formula: C₁₆H₂₁N₃O₂

4) Molecular weight:  287.3568 g/mol

5) Color /Form: Zolmitriptan is white to Cream, crystalline powder

6) Odour: Odourless

7) Solubility: Slightly soluble in water, freely soluble in methanol, dimethyl sulphoxide and dimethyl formamide soluble in ethanol, acetone and in isopropyl alcohol, slightly soluble chloroform, sparingly soluble in acetonitrile.

8) Melting point: 136 °C

9) Category: Antimigraine

10) Mechanism of Action:  Zolmitriptan attaches with a high affinity to human 5-HT1B and 5-HT1D receptors, which leads to the constriction of cranial blood vessels. The therapeutic effect of zolmitriptan in treating migraine attacks is primarily due to its agonistic actions at the 5-HT1B/1D receptors located on intracranial blood vessels (including the arteriovenous anastomoses) and sensory nerves of the trigeminal system, resulting in vasoconstriction and the suppression of pro-inflammatory neuropeptide secretion. ^[13,14]

Experimental Work / Condition

 Preformulation Study

• To establish the necessary physicochemical characteristics of a new drug substance
• To determine its kinetic release rate profile
• To establish is compatibility with different Excipients.

Selection of Active Pharmaceutical Agents

Zolmitriptan, a serotonin receptor agonist of the 1B and 1D subtypes. It is a triptan, used in the acute treatment of migraine attacks with or without aura and cluster headaches. Zolmitriptan is available as a swallowable tablet, an oral disintegrating tablet, and a nasal spray. The usual dose of Zolmitriptan is 2.5 with a maximum dose of 15 mg in 24 h. The conventional tablet is administered 3or 4 times a day due to its short biological half-life of about 2.5-3h, prolong inpatient with hepatic impairment.

Results: For preparation and evaluation of oral film of Zolmitriptan was selected as active drug.

Organoleptic Properties

Confirmation or identification of drug was carried out by following methods.

• Color -White, crystalline powder
• Odour-Odourless
• Appearance - Amorphous Powder
• Taste - Bitter

Melting Point Determination

Melting point of Zolmitriptan was also measured in the laboratory and found to be in the range of 130-140°c.

 Solubility Determination:

The solubility of pure drug in 10 mg/10 ml of solvent was carried out and it reveals that it was slightly soluble in water and freely soluble in methanol and ethanol (95%)

• Distilled water - Readily Soluble
• Methanol- Freely soluble

Procurement of Drug and Polymer

Drug: Zolmitriptan was received from Zydus Lifesciences Limited as a gift sample.

Polymer and other ingredients were used from research laboratory of college.

Table no 1: The General Formula for Formulation of oral Films Is Given Below

1) The final weight of the film = 50mg

2) Dimension (L×W) = 200cm × 180cm

Procedure For Preparation of Oral Film

1. The polymer (HPMC E15 and HPMC E5) are dissolved in methanol accurately weight and Transfer in glass beaker

2. Then dissolve properly with continuous stirring use magnetic stirrer.

3. Dissolve the API and other processing ingredients in methanol weight accurately and transfer into the another beaker and dissolve properly with continuous stirring.

4. Mix the above two solution to form a thick mass.

5. Taking the above thick mass and then transfer into the petri dish

6. Dry this film mass for 2- 3 days at room temperature.

7. After drying then remove thin film. And cut in above size.

8. Optimization of formulation same procedure is followed for all the formulation F1 to F5 and the formulation with all the acceptable properties and parameters to be selected.

Evaluation Parameter

For evaluation of psychophysical evaluation of the product, special controlled human taste panels are used.

1. Organoleptic evaluation

 Appearance, Shape, and color

The formulated films were checked for their appearance, shape and color.

Table no 2 : The physical parameter of all the formulations

Form the above evaluation it is found that all the formulation so prepared are colorless and rectangular shape.

2. Thickness

Film thickness is crucial for ensuring drug content uniformity. Therefore, it is essential to measure the film's thickness at various key locations using a micrometer screw gauge or calibrated digital Vernier Calipers. 

3. Weight Variation

The films underwent a mass variation study, where randomly selected films were weighed individually, and the average weight of five samples from each batch was calculated. This process was repeated for every batch. 

4. Drug Content Uniformity

The films were tested for content uniformity The acceptable range for drug content uniformity. The film was cut, placed in 100 ml volumetric flask and dissolved in methanol, the volume is made upto 100 ml with methanol. Solution was suitably diluted. The absorbance of the solution was measured at 225 nm. The range is 95% to 105%. 

5. Folding endurance

Folding endurance refers to the number of times the film can be folded at the same location before any noticeable cracks appear, signifying film brittleness. This test was performed by repeatedly folding the film at the same point until a visible crack was observed, and the values were recorded. 

6. Disintegration test

The disintegration of orally fast dissolving films is assessed using the USP disintegration apparatus. The recommended disintegration limit is 30 seconds or less for orally disintegrating tablets, which can also be applied to fast dissolving oral strips. Disintegration times may vary based on the formulation, generally ranging from 5 to 30 seconds. However, there is currently no official guidance specifically for oral fast disintegrating film strips. 

7. In vitro drug release

For in-vitro dissolution studies, each film was carefully placed in a 50 ml glass beaker containing 25 ml of phosphate buffer at pH 6.8 using forceps. The dissolution medium temperature was maintained at 37±0.5ºC and stirred at 50 rpm. Throughout the study, 3 ml aliquots were taken at 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 minutes, and these were replaced with fresh buffer. The amount of drug released into the media was quantified using a UV-Visible Spectrophotometer (Shimadzu-1800) at a wavelength of 225 nm.

8. Surface pH

To determine the surface pH of the films, each film was placed on a 1.5% w/v agar gel, followed by the application of pH paper (pH range 1-11) on top of the films. Any color change in the pH paper was noted and documented. 

9. Stability studies

The optimized batch F5 was packed in a butter paper covered with aluminum foil and was isothermally stressed to study the stability under accelerated temperature and relative humidity conditions carried out at 40°C/75% RH, 25°C/60% RH and 25°C/40% RH for a period of 2 months. Test samples were withdrawn every month and were subjected to various tests including visual inspection of the film, disintegration time and cumulative percent of drug release.

 High Performance Liquid Chromatography:

High-performance liquid chromatography (HPLC) is currently one of the most effective instruments in analytical chemistry. It can separate, identify, and quantify the compounds found in any sample that can be dissolved in a liquid. Today, substances at trace levels as low as parts per trillion (ppt) can be readily detected. HPLC has been utilized in a wide range of samples, including pharmaceuticals, food products, nutraceuticals, cosmetics, environmental samples, forensic evidence, and industrial chemicals.

 Instrumentation

The high-performance liquid chromatography (HPLC) of Shimadzu SCL-10AVP inbuilt with binary pump (LC-10ATVP), UV detector (SPD-10AVP), Rheodyne 20µl loop capacity manual injector (P/N 77251) was used throughout the analysis. The LC-Solution software was used to interpret the HPLC reports. Zic-HILIC Mixed Mode (5µm; 150 x 4.6 mm ID.) column was purchased from Ultrachrom Innovatives Pvt. Ltd was used throughout the analysis. Digital weighing balance (ME-204) purchased from Mettler-Toledo (USA), ultra-sonicator Labman® purchased from UltraChrom Ltd, India. Digital pH meter from Mettler-Toledo was purchased from (Mumbai-India). 50 µ micro-syringe was purchased from Hamilton USA. 0.20µ and 0.45µ nylon membrane filters were purchased from Phenomenex® Mumbai, India.

Reagents And Reference Samples

The reference standard Zolmitriptan was purchased Zydus Lifesciences Limited. Ammonium acetate was purchased from Merck Ltd. (Mumbai-India) HPLC grade acetonitrile, methanol and HPLC grade water were purchased from Merck (Mumbai, India). 0.20µ and 0.45µ nylon membrane filters were used and purchased from Ultra Chrom Innovatives Pvt. Ltd. (India). All other chemicals and reagents were used of HPLC grade.

Standard and Sample Preparation

The Standard and sample solutions were prepared separately by dissolving standard and sample in a solvent mixture of Acetonitrile:Methanol:Water (20:40:40 v/v).

 Optimization of Chromatographic Conditions

The chromatographic conditions were optimized by different analysis. (Using different column, different buffer and different modes of HPLC run, Table 3, Fig. a-d.

Table no 3. Method Development for HPLC Analysis of Zolmitriptan API

Method Validation

In order to determine repeatability, a freshly prepared solution of Zolmitriptan was injected six times to evaluate the consistency of results, focusing on the relative standard deviation (RSD); the findings are displayed in Table 6. To establish both intra-day and inter-day precision of the method, Zolmitriptan was analyzed on a single day as well as over three separate days. The calculations for intra-day and inter-day precision are summarized in Table 7-8.  The robustness of the method was tested by modifying chromatographic conditions, including changes in flow rates, mobile phases, and wavelengths. The robustness of the developed method is indicated by the overall standard deviation among the results under each varied condition, as shown in Table 9. Linearity was evaluated by injecting samples of different concentrations (100, 50, 25, 12.5, 6.25, 3.12 ppm), with the data represented in Table 10.  The accuracy of the prepared formulation was determined at 80%, 100%, and 120%, with results displayed in Table 11. The specificity of the proposed method was examined by analyzing a sample solution of Zolmitriptan. Ruggedness was determine using different analysts. The limit of detection (LOD) and limit of quantification (LOQ) were determined from the linearity study. For system suitability, a freshly prepared solution of Zolmitriptan was injected six times to evaluate the closeness of results achieved for the relative standard deviation (RSD) in percentage.

RESULT AND DISCUSSION

Result of Evaluation Parameter of Oral Film

Table no 4 :  Evaluation Parameters of oral Film of Zolmitriptan

Table no 5 : Stability Studies of oral Film

Figure 3: Photograph of oral Film

Chromatographic condition: To establish a precise, linear, specific, and effective HPLC (HILIC-Hydrophilic Interaction Liquid Chromatography) Mixed Mode technique for the analysis of Zolmitriptan, various chromatographic conditions were tested, and the observed results are shown in Table 3 and Figs. 1a-d. There are various columns available for HPLC, the Zic-HILIC Mixed Mode (150 X 4.6 mm. ID.) column was selected here due to its favourable peak shape, resolution, and absorbance. The Mobile Phase utilized was a mixture of 15mM Ammonium acetate and Acetonitrile in a 50:50 v/v in a ratio.

Table 6: Repeatability Study of Zolmitriptan

Table 7:  Intraday Precision Data of Zolmitriptan

Table 8: Interday/Intermediate Precision Data of Zolmitriptan

Table 9: Robustness data of Zolmitriptan

Table 10: Linearity Data of Zolmitriptan

Table 11 : Accuracy data of Zolmitriptan