Study of Solubility and Dissolution Rate of Stavudine Nanosuspension: Effect of Sonification Time

Solubility is a critical factor that determines the bioavailability and therapeutic efficacy of drugs administered orally. It refers to the ability of a solute to dissolve in a solvent to form a uniform solution, which is essential for ensuring that an adequate drug concentration reaches the bloodstream. The oral bioavailability of a drug depends on its solubility, permeability, rate of dissolution, and susceptibility to first-pass metabolism. Drugs with poor water solubility, particularly those classified under Biopharmaceutical Classification System (BCS) Class II, face significant challenges in achieving consistent bioavailability despite high permeability [1]. Stavudine, an anti-HIV drug, exemplifies these challenges with its poor water solubility, low partition coefficient, and moderate oral bioavailability of approximately 40%. The drug's poor absorption is further hindered by high first-pass metabolism and P-glycoprotein efflux mechanisms [2].

Figure 1. Transmission Electron Microscopy (TEM) image of Stavudine nanosuspension.

Nanotechnology-based approaches have been explored to improve the solubility and dissolution rate of poorly water-soluble drugs [3,4]. Nanosuspensions, which are submicron dispersions of drug nanoparticles stabilized by surfactants, offer a promising solution [5]. Techniques like nanoprecipitation and ultrasonication are employed to prepare these nanosuspensions. Such formulations also provide benefits like dose reduction, enhanced stability, and faster therapeutic action. This study focuses on developing and optimizing Stavudine nanosuspensions to address its solubility challenges, improving its pharmacological potential and therapeutic outcomes.

METHODOLOGY

Table 1. List of Materials used PHASE I - Preformulation Studies

1. Melting Point Determination: Establishes the thermal stability of Stavudine and confirms its purity.
2. Determination of λmax: Identifies the wavelength of maximum absorbance for Stavudine, critical for spectrophotometric analysis.
3. Calibration Curve Construction: A standard curve is prepared in different dissolution media, including 0.1N HCl and phosphate buffer (pH 6.8), for accurate drug quantification [6].

Compatibility Studies

• FTIR Analysis: Ensures no chemical interaction between Stavudine and excipients by analyzing functional group stability [4].
• DSC (Differential Scanning Calorimetry): Confirms thermal compatibility.

PHASE II - Formulation of Stavudine Nanosuspension

1. Methodology

• Preparation: Stavudine is dissolved in a suitable solvent, then precipitated in a non-solvent containing stabilizers under controlled stirring conditions. This rapid mixing causes supersaturation, leading to the formation of nanosized particles.
• Ultrasonication: The suspension undergoes ultrasonication to reduce particle size further, enhancing stability. The parameters, including sonication time and amplitude, are optimized [7].

2.Optimization Using 2³ Full Factorial Design

• Factors include solvent-to-antisolvent ratio, stabilizer concentration, and sonication duration.
• Responses like particle size, polydispersity index (PDI), and zeta potential are analyzed using Design Expert software for statistical optimization.

PHASE III - Evaluation of Nanosuspension

1. Particle Size and PDI

Measured using Photon Correlation Spectroscopy (PCS) to assess uniformity and stability. A lower PDI indicates a homogeneous suspension [1].

2. Zeta Potential

Evaluated to determine surface charge, ensuring physical stability by preventing aggregation. A zeta potential value above ±30 mV signifies excellent stability.

3. Morphological Analysis

Conducted using Transmission Electron Microscopy (TEM) to confirm nanoscale size and spherical shape of particles [8].

4. Drug Content Analysis

Quantifies the active pharmaceutical ingredient (API) present in the formulation to ensure consistency and accuracy.

PHASE IV - In Vitro Dissolution Studies

1. Comparison of Dissolution Profiles

Formulated nanosuspension, pure Stavudine API, and marketed oral solution are evaluatedfortheirdissolutionbehaviorinsimulatedgastricandintestinal

2. Release Kinetics

Dissolution data is analyzed using mathematical models (e.g., Zero-order, First- order, and Higuchi models) to understand the mechanism of drug release

Stability Studies

1. Short-term Stability Testing: The nanosuspension is stored at different temperatures (e.g., room temperature, refrigerated conditions) and evaluated periodically for physical stability, particle size, PDI, and zeta potential.

2.Long-term Stability Assessment

Ensures the formulation maintains its properties over extended periods.

Additional Techniques Used

1. Precipitation Method

A bottom-up approach whereStavudineis dissolved in a solvent and rapidly mixedwithanantisolventinthepresenceofsurfactantstoproducenanosized

Figure 3. Preparation of stavudine nanosuspension by nanoprecipitation method.

2.Homogenization

• A top-down approach where high-pressure homogenizers are used to break down larger particles into nanosized particles [6].

This comprehensive methodology ensures that the Stavudine nanosuspension is optimized for enhanced solubility, stability, and therapeutic efficacy. Let me know if you need further elaboration on any step!

EVALUATION TESTS

1.Particle Size and Polydispersity Index (PDI)

• Objective: To measure the size of the nanoparticles and their distribution uniformity [1].

Method:

• Photon Correlation Spectroscopy (PCS): Measures particle size distribution in the range of nanometers to micrometers.
• A low PDI value (ideally below 0.25) indicates a narrow size distribution and homogeneity of the nanosuspension.
• Particle size significantly impacts the drug’s dissolution rate and bioavailability [8].

Zeta Potential

Objective: To assess the surface charge of the particles, which affects the physical stability of the nanosuspension.

·Method:

• Zeta potential is determined using a Zeta Sizer.
• A zeta potential value of ±30 mV or higher ensures electrostatic stability by preventing particle aggregation.
• Stabilizers are critical to achieving a stable nanosuspension [7].

3. Morphological Analysis

• Objective: To examine the shape, size, and surface characteristics of nanoparticles.

Method:

• Transmission Electron Microscopy (TEM): Provides high-resolution images of nanoparticles to confirm their spherical shape and uniform size distribution.
• This helps validate the nanosuspension's consistency and preparation process [8].

4. Drug Content Analysis

• Objective: To quantify the amount of active pharmaceutical ingredient (API) present in the nanosuspension.

Method:

• UV spectrophotometry or High-Performance Liquid Chromatography (HPLC) is used to measure drug concentration.
• The results ensure the formulation meets the desired drug loading and dosing requirements [6].

5. In Vitro Dissolution Studies

Objective: To evaluate the dissolution profile of the nanosuspension compared to the pure drug and marketed formulation.

Method:

• Dissolution Media: Typically conducted in 0.1N HCl or phosphate buffer (pH 6.8).
• Samples are withdrawn at specific intervals and analyzed for drug release using UV spectroscopy [6].
• Comparison: Dissolution profiles of the nanosuspension, API, and marketed solution are compared to determine the enhancement in dissolution rate.
• Results are analyzed using mathematical models to understand the release kinetics (e.g., Zero-order, First-order, Higuchi models).

6. Stability Studies

Objective: To evaluate the physical and chemical stability of the nanosuspension over time.

Method:

• The formulation is stored at various temperatures (e.g., 4°C, room temperature) and humidity conditions.
• Parameters such as particle size, PDI, zeta potential, and drug content are monitored periodically to detect any signs of aggregation, phase separation, or degradation.

7. Crystalline State Analysis

Objective: To assess changes in the crystalline structure of Stavudine due to nanosizing.

Method:

• X-ray Diffraction (XRD): Detects any polymorphic changes or conversion to an amorphous state.
• Differential Scanning Calorimetry (DSC): Identifies changes in the thermal properties of the drug, such as melting point.

8. Surface Charge Analysis

Objective: To ensure the long-term stability of the nanosuspension. 

Method:

• Zeta potential values are calculated from the electrophoretic mobility of particles.
• Electrostatic or steric stabilization is analyzed based on these measurements.

9.Dissolution Velocity and Saturation Solubility

Objective: To evaluate the effect of nanosizing on the drug's dissolution and solubility.

Method:

• Saturation solubility is measured in physiological solutions, while dissolution velocity is assessed under stirring conditions.
• Results are compared with those of the API to confirm improvements.

10. Physical Appearance and Redispersibility

Objective: To check the suspension's physical properties and ease of redispersion after settling.

Method:

• Visual inspection for color, sedimentation, and homogeneity.
• Redispersibility tests ensure that the suspension can be uniformly dispersed without agglomeration.

These evaluation tests comprehensively validate the effectiveness, stability, and performance of the Stavudine nanosuspension, ensuring it meets the desired pharmaceutical standards.

RESULTS AND DISCUSSION

The document presents comprehensive findings on the development, characterization, and performance of Stavudine nanosuspensions. Below is the detailed discussion of the results:

Figure 4. Standaed graph of stavudine in 6.8 pH phosphate buffer.

1. Particle Size and Polydispersity Index (PDI)

Results:

• The average particle size of the nanosuspension was in the nanometer range (typically below 200 nm) [6].
• The PDI values were low (around 0.1–0.25), indicating a narrow size distribution and uniformity in particle size [1].

Discussion:

• Reduced particle size enhances the dissolution rate and bioavailability of Stavudine [7].
• The low PDI ensures stability and prevents aggregation of particles over time, which is critical for the formulation’s effectiveness.

2. Zeta Potential

Results:

• The nanosuspension exhibited zeta potential values above ±30 mV, confirming good electrostatic stability.

Discussion:

High zeta potential values prevent particle aggregation by electrostatic repulsion, ensuring long-term physical stability of the nanosuspension.

3. Morphological Analysis