Comparative Invitro and Exvivo Evaluation of Transferosomal Patches for Enhanced Drug Delivery of Lisinopril Dihydrate

Background

Transdermal administration of drugs is generally limited by the barrier function of the skin. Vesicular systems are one of the most controversial methods for transdermal delivery of active substances. Permeability of the drug through the skin involves many limitations like the movement of the drug through stratum corneum and physical characters of the drug. These factors led to the incorporation of nanocarries in the preparation of transdermal formulation. These delivery systems were designed in an attempt to penetrate the drug through the skin into the blood. This helps in reducing the unwanted drug effects on the other organs of the body caused by oral drug delivery [1]. Permeability of the drug through the skin involves many limitations like the movement of the drug through stratum corneum and physical characters of the drug. These factors led to the incorporation of nanocarriers in the preparation of transdermal formulation. The nanocarriers have a prominent role in delivery drugs through various route of administration. The transdermal carriers made the drugs not only to move through the stratum corneum but made a wide verity of drugs to be suitable for the transdermal drug delivery. In the present research transdermal drug delivery patches were developed with incorporation of transferosomes as nanocarriers which are suitable for transdermal drug delivery. A comparative study is done among the normal transdermal patches with nanocarrier loaded transdermal patches using lisnopril. Lisinopril dehydrate is an effective blood pressure lowering drug [2],  it  is a correlates with lysine of enalaprilat. As lisnopril has poor oral oral bioavailability, this makes the drug suitable for transdermal drug delivery [3]. The present article aimed in the formulation of the nanocarriers suitable for the transdermal drug delivery and compared with normal transdermal patches the drug release studies and release kinetics were performed. A comparative illustration of both the results is made and the efficacy of nanocarriers was reported.

MATERIALS AND METHODS

Lisnopril dehydrate was obtained as sample from yarrow chem. Products Ghatkopar West, Mumbai, Maharashtra India. Cholesterol, Soya lecithin were purchased from Sigma Aldrich. HPMC E15 was purchased from CDH, New Delhi, India. Span 80, Tween 80 was obtained from Loba Chemie, Mumbai, India. All other chemicals used were of analytical grade.

Preparation of Nanovesicles:

Transferosomes were prepared by using film evaporation method. The surfactant and phospholipids were taken in round bottomed flask. The solvent mixture of methanol and chloroform were added and evaporated with the help of Rota evaporator. The film deposited after evaporation was hydrated with the help of buffer PBS (pH- 7.4). This results in the formation of a colloidal suspension [4]. The suspension formed is then ultrasonicated for the formation of uniform vesicles.

Table No. – 1: Formulation of Nanovesicles

Characterization of transferosomes

Determination of the vesicle size, PDI and zeta potential of the prepared trasferosomes were determined based on laser diffraction using the Malvern Master seizer by diluting the sample using water as dispersant [5].

Determination of Vesicle size

Vesicle size of the nanoparticles was determined by using a Zetasizer 1000 HS (Malvern Instruments, UK). Light scattering was monitored at a 90º angle at 25ºC.

Entrapment efficiency: The amount of the drug entrapped was estimated separation of nanovesicles% Entrapment efficiency =   Total amount of drug added – Nonbound drug

Total amount of drug

Transferosomes containing drug was separated from entrapped drug by centrifugation at 14,000 rpm for 30 min. The supernatant was filtered and assayed [6]

Preparation of transdermal patches with Nano formulation:

The transdermal patches along with nano suspension were prepared using solvent casting method. As mentioned in table no. 2 two different gums HPMC 15cps, HPMC 5cps with variable percentages were used and along with polyethylene glycol- 400 as plasticizer. All the ingredients were weighed accurately and taken in beaker along with nanosuspension [7]. Water is used as solvent and added to the mixture of ingredients with through mixing. The mixture is casted over a Teflon plate of 4cm diameter and was dried in  hot air oven for at 30⁰C for 24hrs. Keeping inverted funnel over the plate to control the rate of drying.

Table– 2: Formulation of Transferosomal Patches:

Preparation of transdermal patches without nanoparticles :

The transdermal patches without nanosuspension was prepared using solvent casting method. As mentioned in table no. 2 two different gums HPMC 15cps, HPMC 5cps with variable percentages were used and along with polyethylene glycol- 400 as plasticizer. All the ingredients were weighed accurately and taken in beaker. Water is used as solvent and added to the mixture of ingredients with through mixing. The mixture is casted over a Teflon plate of 4cm diameter and was dried in hot air oven for at 30⁰C for 24hrs. Keeping inverted funnel over the plate to control the rate of drying [8].

Table 3 : Formulation of transdermal patches without nanovesicles:

Evaluation of transdermal patch

Thickness Variation Test: Vernier calipers was used to measure the thickness of the films at Six points of the patch [9]

Weight Variation Test: The formulated films were prepared in triplicate. Three films from each batch were weighed one after the other and the average weight is noted.

Folding Endurance: Folding endurance of transdermal patches was estimated by folding a strip of film (2cmx2cm) at the one place continuously till it breaks. The number of times of folding at the same place without breaking was the value of folding endurance.

In-vitro diffusion studies: In vitro drug release studies were done by Franz diffusion cell. The cell was fabricated and receptor compartment volume was 25ml. A dialysis membrane was used to perform diffusion studies by placing patch uniformly on dialysis membrane.

Ex-vivo skin permeation studies

The exvivo studies were performed by taking pork skin

Flux (?g/cm2

The following equation:

Flux (Jss) = dMS

Where dM - Amount of drug permeated, S   - Unit cross-section area, t -time(t).

From the equation, the slope of the steady state portion of the permeation curve is known by plotting the cumulative amount of drug permeated (µg) versus time in hours.

Lag Time (hrs): Lag time is the time required for the drug to get released from the nanovesicles. It is calculated by keeping cumulative amount of drug permeated vs time. The x-intercept value gives the lag time.

RESULTS AND DISCUSSION:

Characterization of Lisnopril dihydrate: lisnopril was identified and characterized as per official compendia. The drug purity was determined by IR spectra and melting point was found to be 162°C. The drug was found to be freely soluble in water. The partition coefficient of tolterodine tartrate in n-octanol: pH 7.4 phosphate buffer was found to be 1.62±0.16.

Evaluation of transferosomes:

The prepared transferosomes were formulated as mentioned in Table 1 and optimized by evaluating with different evaluation methods.

Table No. – 4: Evaluation of transferosomal nanoparticles

Note: Sample size (n=3)

From table no 4 it can be concluded that the prepared nanoparticles with labrosol as edge activator has better entrapment efficiency, optimum size and have shown better drug release when compared to other formulations.

Figure No. -2: Bar diagram representation of percentage drug release for different formulations (n=3)

       
           
       
Fig. – 6 : The  zeta potential  results for nanoparticles

Fig -8 fluorescent microscopic images of nanovesicles formulation T3 Edge activator Labrosol, magnification at x 40

Table No.-5: Optimization of transdermal patches with transferosomes (T3)

Note : Sample size (n=3)

The formulations with various percentages of HPMC 5cps (1%w/v,1.5% w/v, 2% w/v, 2.5% w/v) and HPMC 15cps (1%w/v,1.5% w/v, 2% w/v, 2.5% w/v) were prepared using PEG-400 as plasticizer with water as solvent [10]. The nanosuspension of 0.5ml was mixed with the above mixer. The patches prepare were evaluated for weight variation, folding endurance, thickness, diameter, area, surface pH and percentage drug release. The results were tabulated in table no 5.

 From the table no.-5 it was inferred that all the formulations have optimum folding endurance and surface pH. Based on the percentage drug release studies for 24hrs the patches the prepared transdermal patches were optimized [11]

The transdermal patches with 2.5% w/v HPMC 15cps were found to have optimum drug release at 24^th hr in comparison with other patches [12]. The patches with less percentage of HPMC 15Cps were found to deliver the drug prior to 24 hrs and stated decreasing the release at the end of 24 hrs.

Table No. 6: Evaluation of transdermal patches without transferosomes

Note: Sample size (n=3)

The evaluation of transdermal patches without nanovesicles are performed and tabulated in table no.6[14]. From the table it can be inferred that the drug release of the formulations without nanovesicles was found to have very less drug release.

       
           
       
 Fig. 13: A Comparitive invitro drug release profile of transdermal patches without transferosomes.

Table No-7: Exvivo Permeation Studies Using Pork Skin

Note: Sample size (n=3), TL4, TL5- transferosomal patches; L2, L5- patches without

transferosomes.

The Table no.7 depicts the evaluation studies performed using transferosomal patches and and non transferosomal patches[15]. The cumulative amount of the drug permeated, flux and skin content were calculated and it was observed that the transferosomal patches have better Q₂₄ , flux and less skin content compared to non transferosomal patches. The figures 14&15 represent the comparative evaluation of transdermal patches.