Pharmacokinetics and Bio-Distribution Properties of a Self-Emulsifying Drug Delivery System Containing Nevirapine

Research Article

J Drug Discov Develop and Deliv. 2020; 6(1): 1035.

Pharmacokinetics and Bio-Distribution Properties of a Self-Emulsifying Drug Delivery System Containing Nevirapine

Abonyi JI1, Uronnachi EM2*, Umeyor CE2, Chime SA1, Kenechukwu FC1, Attama AA1* and Ibezim EC1

¹Department of Pharmaceutics, University of Nigeria, Nigeria

²Department of Pharmaceutics and Pharmaceutical Technology, Nnamdi Azikiwe University, Nigeria

*Corresponding author: Attama AA, Department of Pharmaceutics, University of Nigeria, 41001 Nsukka, Enugu State, Nigeria

Uronnachi EM, Department of Pharmaceutics and Pharmaceutical Technology, Nnamdi Azikiwe University, Awka, Anambra State, Nigeria

Received: February 05, 2020; Accepted: May 02, 2020; Published: May 09, 2020


Nevirapine has several dose related side effects. Thus necessitating a formulation with increased bioavailability causing a reduction in dosage. The work was aimed at preparing and evaluating a Self-Emulsifying Drug Delivery System (SEDDS) containing nevirapine. Solubility of nevirapine in various vehicles (oil, surfactant and co-surfactant) was determined and those with the highest solubilizing potential for nevirapine were selected. The pseudo ternary phase diagrams composed of lipid (Soya oil) surfactant-co-surfactant mixture (solutol-cremophor EL) water were mapped and combinations with better micro emulsification properties were chosen. Different batches of nevirapine SEDDS were formulated and assessed. The in vivo release profiles of the nevirapine SEDDS were studied using adult albino Wistar rats. The CD4 counts of the animals and biodistribution of nevirapine to various organs was also determined. SEDDS formulation improved the aqueous solubility and bioavailability of nevirapine and resulted in more reproducible blood–time profile. The biodistribution of nevirapine SEDDS was lower to the liver but higher to the brain when compared with conventional nevirapine tablets (p<0.05). Nevirapine SEDDS had a higher time-dependent increase in CD4 cells/μl than the conventional tablets (p<0.05).

Nevirapine SEDDS increases the drug’s aqueous solubility, permeability and bioavailability enabling dose reduction.

Keywords: Antiretroviral; Plasma profile; Emulsification; In vivo kinetics


The oral route is the most convenient means of drug administration to patients. This route however poses problems of absorption for many drugs owing to their poor solubility profiles. Drug discovery data suggest that about 40% of new drug entities introduced into the market have solubility problems [1]. To overcome this challenge, several drug delivery systems have been explored to enhance drug solubility, dissolution in the gastrointestinal tract and, ultimately, absorption into the systemic circulation. Self-Emulsifying Drug Delivery Systems (SEDDS) have great potential in improving drug bioavailability.

SEDDS are mixtures of oils and surfactants, ideally isotropic, and sometimes containing one or more hydrophilic solvents as cosurfactants/ co-solvents, which emulsify spontaneously to produce fine oil in-water emulsions when introduced into aqueous media such as the Gastrointestinal Tract (GIT) under gentle agitation [2,3]. Recently, SEDDS have been formulated using medium chain triglyceride oils and non-ionic surfactants, the latter had been less toxic. SEDDS can be orally administered in soft or hard capsules. Some of the advantages of SEDDs include: Improved drug solubilization, long half-life, improved oral bioavailability, consistency in drug absorption, protection against hydrolysis by enzyme in GIT, reduction of gastrointestinal metabolism of drug prior to systemic absorption, by-pass of hepatic first-pass metabolism, selective targeting of drugs toward specific absorption window in GIT, protection of sensitive drug substances, reduced variability including food effects [4,5].

The process of self-emulsification proceeds through formation of Liquid Crystals (LC) and gel phases, the properties of which significantly affect the formation of droplets and interfaces available for partitioning of drug [6-8]. Lipophilic substances with poor solubility are thus candidates for such formulations. Several SEDD formulations have been explored to enhance the solubility of poorly soluble drugs [9-18].

Nevirapine is a Non-Nucleoside Reverse Transcriptase Inhibitor (NNRTI) which is one of the drugs used in the first line management of HIV/AIDS as a component of the Highly Active Antiretroviral Therapy (HAART) regimen. Its poor solubility and high permeability makes it a Biopharmaceutics Classification System (BCS) class II drug, thus making it a candidate for SEDDS formulation (Figures 1-4).

This study was aimed at determining the ability of a SEDDs formulation prepared using locally available oils, to enhance the solubility and release profile of nevirapine, as well as determine its effect on the pharmacokinetic parameters of the drug in immunecompromised rats.

Materials and Methods


The following materials were used as procured from their manufacturers: Nevirapine, a gift from Fid son Healthcare Ltd, Lagos; Solutol® HS 15 (BASF Ludwigshafen, Germany), Cremophor® EL (Ludwigshafen, Germany), cyclophosphamide (Korea United Pharm Inc), soya oil (processed in our Laboratory), ammonia (Merck, England), hydrochloric acid (Merck, Germany), Monobasic potassium phosphate, ethanol (BDH Chemicals Ltd Poole, England), sodium chloride (Merck, England), sodium hydroxide (Avoadale Laboratories, England), distilled water, and water for injection. All other reagents were of analytical grade and were used as received.


Extraction of soya oil: About 45 kg of soya beans was crushed to powder using a laboratory mill. The resultant powder was soaked in 2.5 L of n-hexane for 48 h and subsequently passed through a muslin cloth to filter it and dried using a rotary evaporator (GmbH, Germany).

Solubility studies of nevirapine in various vehicles: The solubility of nevirapine in various lipids, surfactant and co-surfactants was determined. The method of Shen and Zhong [17] was used with slight modification. An excess amount of nevirapine was introduced into 2 ml of each excipient and the mixture in a capped cuvette was stirred in a water bath at 25oC. A vortex mixer was used to facilitate the solubilization. After standing for 24 h and reaching equilibrium at ambient temperature, each cuvette was centrifuged at 3000 rpm for 10 min using a centrifuge (Sigma 3 k15; Sigma USA). Undissolved nevirapine was removed by filtering in a membrane filter (0.45 μm). The concentration of nevirapine was determined using a UV-Vis spectrophotometer (Jenway 6405, USA) from a calibration curve of the drug in the oils at a predetermined wavelength of 311 nm [9,18] (Tables 1-3).

Construction of pseudo ternary phase diagram: Pseudo ternary phase diagrams consisting of lipid, surfactant, co-surfactant and water were constructed according to the method using the titration method [19]. The non-ionic surfactant, Solutol® HS, and the solubilizer, Cremophor® EL, as co-surfactant were selected. The lipid employed was soya oil. The surfactant was blended with co-surfactant in the ratio of 2:1, 1:1, 3:1 and 1:2 using a magnetic stirrer (IKA, Germany) at 200 rpm for 10 min. Volumes of each surfactant and co-surfactant mixture (Smix) were blended with lipid in a ratio of 1:9, 2:8, 3:7, 4:6, 5:5, 6:4, 7:3, 8:2, 9:1 w/w also at 200 rpm for 10 min using a magnetic stirrer (IKA, Germany). Water was folded in a drop-wise manner to each lipid-Smix with gentle shaking at 37 oC. After equilibrium, the appearance and dispersibility of the formulation were observed, photomicrographs taken and droplet size distribution was analyzed. The amount of water, lipid, surfactant and co-surfactant folded was noted down, and calculated. The pseudo ternary phase diagrams were mapped using Sigma Plot Window 6.1 (USA). The micro emulsion regions in the diagrams were plotted and the ternary diagrams with wider region indicated the better self-micro emulsification efficiency [20].

Stability analysis

Globule size analysis: The particle size/distribution was analysed by computerized image analysis using a motic image analyzer (Moticam, China) attached to a binocular microscope (Weltzer, Germany). The nevirapine SEDDS formulation was dispersed in water on a microscope slide covered with a cover slip and imaged using the motic.

The stability of the micro emulsion was assessed by analyzing droplet size and distribution. Isotropicity test

Stability of the micro emulsions was assessed visually. Changes in droplet size, phase separation and /or precipitation were noted. This was done 24 h after preparation and repeated three months post preparation.

Photomicrographs of the micro emulsion: The structure, droplet size, and size distribution of particles of the micro emulsion were observed using a motic image analyzer (Moticam, China) attached to a binocular microscope (Weltzer, Germany).

Preparation of nevirapine SEDDS: After the pseudotenary phase diagrams were plotted and compared, optimum surfactant, co-surfactant and lipid combination were selected. Nevirapine SEDDS formulations were prepared by firstly dissolving nevirapine into Solutol® HS-Cremophor® EL mixture (Smix) in a test tube heated to 25oC in a water-bath, then, the required weight of the lipid was added in the test tube and mixed properly using a magnetic stirrer (IKA, Germany) at 200 rpm for 10 min. The mixture was filled into capsules and tightly sealed and stored at a temperature of 25oC until required. Different ratios of Smix to oil were used to formulate different concentration of nevirapine SEDDS. The ratios of Smix to oil used were 8:1, 6:1, 4:1 and 2:1.

Emulsification time test: A 1.0 ml volume of each of the different concentrations of nevirapine SEDDS formulation was titrated with water. The emulsification time was noted and recorded. The volume of water used was also recorded. The photomicrograph of the nevirapine micro emulsion was also taken.

Infinite dilution test: A 1 ml volume from each formulation of nevirapine SEDDS was diluted to 10 ml, 100 ml and 1000 ml with distilled water respectively and the degree of phase separation noted.

Absolute drug content: Beer’s calibration curve of nevirapine was obtained fornevirapine in plasma and ethanol at a concentration range of 1 to 5 mg% respectively at a predetermined wavelength of 255 and 291 nm respectively. A 50 mg quantity of nevirapine SEDDS was dissolved in ethanol and analyzed in a spectrophotometer (Jenway 6405, USA) at 291 nm. The determination was replicated three times and the mean taken to obtain the absolute drug content for each batch.

In vivo drug release studies: Prior to commencement of this study, ethical clearance was got from the ethical committee of the institution. The animal studies were conducted in line with the revised Helsinki declaration of 2000 and in accordance with the guidelines set forth in the eight edition of the guide for the care and use of laboratory animals published by the National Academy of Sciences, the National Academies Press, Washington, D.C.

In this study, fifteen adult Albino Wistar rats of both sexes (130- 160 g) were used. The rats were weighed and divided into groups of threes’. They were allowed access to food and water ad libitum for one week to acclimatize. Blood samples of rats were withdrawn to determine the base line CD4 count (Figures 5 and 6). Afterwards, a single dose of 30 mg/kg of cyclophosphamide was administered to the rats Intraperitoneally (IP) to induce immunosuppression [21,22]. The immune suppressed rats were allowed to starve for 24 h with free access to drinking water. At the end of this period, blood samples were withdrawn from the retro-orbital plexus of the rats at time, t=0. Subsequently, 10 mg/kg of pure nevirapine was administered to the rats in group one (served as control) while the remaining four groups received 10 mg/kg of nevirapine SEDDS preparation. After administration, blood samples were withdrawn from the retro-orbital plexus of the animals at intervals of 1, 3, 5, 8 and 15 h respectively with the aid of heparinized capillary tubes and placed in EDTA bottles. The withdrawn blood samples were analyzed for CD4 cells using a CD4 count machine. The blood samples were centrifuged (Abishkar Centrifuge, India) at 5000 rpm for 10 min. The resulting plasma was then carefully collected with the aid of 1 ml syringe and its absorbance read using a UV-Vis spectrophotometer (Jenway 6405, USA)

Biodistribution studies: The distribution of nevirapine from the SEDDS to various organs of the body was checked using healthy albino rats. Three groups of six rats were used. Cyclophosphamide, 30 mg/kg was administered orally to all the rats and denied access to food for 24 h with free access to drinking water. The test group received nevirapine SEDDS dispersed in water and given orally, (equivalent to 10 mg/kg), the control groups received normal saline, while the reference group received nevirapine pure drug (10 mg/kg). After the administration, the rats were sacrificed at intervals from 1 to 1.5 h. The sacrificed animals had their kidneys, livers, brains and spleens harvested. The harvested organs were pulverized, and soaked in ethanol for about 30 min, and filtered. The filtrates were then analysed with a UV-Vis spectrophotometer (Jenway 6405, USA) at a predetermined wavelength of 255 nm for nevirapine content.

Pharmacokinetic and data analysis: For group analysis, Student t-test was used. The pharmacokinetic data was analyzed statistically using the non-compartmental model obtained with the Win Nonlin software (Version 4; Pharsight Inc, Mountain CA). Data from the plasma concentration time curve with 15 h after drug intake were used to obtain the peak plasma concentration (Cmax, mg/ml), time to peak plasma concentration (Tmax, min), Mean residence time (MRT, min).

Results and Discussions


Yield of soya oil extract: The results of the yield of soya oil showed that the percentage yield of the soya oil extracted was 66.5%. The results show that soya bean exhibited high yield of oil thus making it a relatively efficient source of oil.

Solubility of nevirapine in surfactant solutions: The results of the solubility of nevirapine in surfactant solution are shown in Figure 1 and show that the drug exhibited solubility of 260, 200 and 120 mg/ ml in Solutol® HS, Cremophor® EL and Tween® 80 respectively. These results thus revealed that nevirapine exhibited significantly higher solubility in Solutol® HS (p<0.05).