Development and Evaluation of Chloroquine Phosphate Microparticles using Solid Lipid as a Delivery Carrier

Research Article

J Drug Discov Develop and Deliv. 2015;2(1): 1011.

Development and Evaluation of Chloroquine Phosphate Microparticles using Solid Lipid as a Delivery Carrier

Ogbonna JDN¹*, Nzekwe IT², Kenechukwu FC¹, Nwobi CS¹, Amah JI¹ and Attama AA¹

¹Department of Pharmaceutics, University of Nigeria, Nsukka, Nigeria

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

*Corresponding author: Ogbonna John Dike N, Department of Pharmaceutics, University of Nigeria, Nsukka, 410001, Nigeria

Received: February 17, 2015; Accepted: May 05, 2015; Published: May 07, 2015


The aim of the study was to formulate and evaluate in vitro-in vivo Chloroquine (CQ)-loaded Solid Lipid Microparticles (SLMs). CQ-loaded SLMs were prepared by hot homogenization, lyophilized and characterized using particle size, pH stability, Loading Capacity (LC) and Encapsulation Efficiency (EE). In vitro release of CQ was performed in SIF and SGF and in vivo study done using Peter’s Four day in mice, there after kidney and liver of the mice were subjected to histological studies.

The formulations exhibited high entrapment efficiency and yield. Timedependent pH stability studies showed little variations with range from 3.93±0.21- 5, 46±0.23. The release profiles of CQ-loaded SLMs showed a gradual, steady release of the drug at various intervals in both SIF and SGF compared to the commercial CQ samples for 8 h. The in vivo study showed a high percentage reduction in parasitemia with minimal effect on vital organs. The SLMs exhibited sustained release with a pH-dependent release profile as the highest release was obtained in SIF than in SGF. The results showed that the percentage reduction in parasitemia of the optimized SLMs formulation (87.01%) had better activity than the commercial sample (84.12%).

The histological studies revealed that the SLMs formulations have no harmful effects on the organs of the mice. SLMs formulations might be an alternative for delivery of CQ to patients with parasitemia.

Keywords: Solid lipid microparticles; Chloroquine; Parasitemia; Haematological parameters; Histological studies


C0 is SLMs unloaded with CQ; C1 is SLMs loaded with 3 % of CQ; C2 is SLMs loaded with 5 % of CQ; C3 is SLMs loaded with 7 % of CQ


Resistance to antimalarial medicines is a recurring problem in recent years, thus parasitic resistance to artemisinins has been detected and if this resistance to artemisinins develops and spreads to other large geographical areas, the public health consequences could be dire. This treatment failure of the new antimalarials have prompted the quest to formulate CQ in novel alternative delivery systems as it is always the cheapest for the local populace, mostly affected by malaria. Solid Reverse Micellar Solutions-based (SRMS-based) SLMs is a new formulation field with advantages over other carrier systems with high potentials for sustained drug release and gastro-protection [1]. Some proposed mechanisms of action of lipid-based systems to enhance oral bioavailability of compounds includes; increased rate of dissolution into aqueous environment from oil droplets of high surface area, promotion of absorption via intrinsic lipid pathways and enhanced thermodynamic activity via supersaturation of the aqueous environment of the gastrointestinal tract [2,3]. SLMs attract increasing attention in alternative delivery systems as they combine advantages of traditional carriers; for example they can be produced on a large industrial scale, are toxicologically highly acceptable and also allow the control of drug release [4].

Parasitic diseases are of immense global significance as around 30 % of world’s population experience parasitic infections. Malaria, the most life threatening disease among the parasitic infections accounts for 1 to 2 million deaths round the globe every year with the estimate each year that more than 200 million people are infected with malaria worldwide [5]. Malaria, a common problem in areas of Asia, Africa and Central and South America is responsible for 1 in 5 childhood death in Africa and at least fifty per cent of Nigerian population experience one episode of malaria every year [6]. It is the cause of 1/5 of death before the age of 5 years and 1/3 of deaths for children in urban and rural areas of Nigeria respectively [7]. Malaria preferentially affects children younger than 5 years of age, pregnant women, and non-immune individuals [8] becoming more difficult to treat because of multidrug parasite resistance. In humans, malaria is caused by four distinct blood–borne Ampicomplexan parasite species: Plasmodium vivax, Plasmodium malariae, Plasmodium ovale and Plasmodium falciparum being responsible for almost all malaria related deaths as it causes the most severe malaria [9]. Plasmodium berghi transmitted by Anopheles mosquitoes are practical model organism in the Laboratory for the experimental study of human malaria as the symptoms are to a certain degree comparable to symptoms of cerebral malaria in patients infected with the human malaria parasite P. falciparum [10]. In addition, P. berghei is used in research programs for development and screening of anti-malarial drugs and for the development of an effective vaccine against malaria. The recent call for the elimination and eradication of the disease requires research from multiple fronts, including developing strategies for the efficient delivery of new medicines [11].

CQ is a 4-aminoquinoline compound for oral administration. It is indicated for suppressive treatment and acute attacks of malaria due to P. vivax, P. malariae, P. ovale, and susceptible strains of P. falciparum including treatment of extra intestinal amoebiasis as it is rapidly and almost completely absorbed from the gastrointestinal tract [12]. The drug is generally safe and toxicity occurs when very high doses are administered via parenteral route.

Previously, formulation and evaluation of halofantrine-loaded Solid Lipid Microparticles (SLMs) have been done by our research group [13]. The aim of the present work was to formulate CQ-loaded SLMs and assess the antimalaria activity of the formulations both in vitro and in vivo in P. berghei infected mice and compare their activity with commercial CQ sample tablets.

Materials and Methods


The materials used were pure CQ sample (Juhel Pharmaceuticals, Nigeria), commercial CQ sample (Evans Pharmaceuticals, Nigeria), Phospholipon® 90H (P90H) (Phospholipid GmbH, Köln, Germany), sorbic acid, sorbitol, polysorbate 80 (Merck, Darmstadt, Germany), distilled water (Lion Water, Nigeria), goat fat (obtained from a batch processed in our Laboratory). All other reagents and solvents were of analytical grade and were used without further purification.

Parasites: P. berghei NK-65, a strain free of contamination with Eperythrozoon coccoides and sensitive to CQ, was used for in vivo antimalarial study. This strain is known to induce high mortality in mice, providing a good model to estimate antimalarial efficacy in reducing parasitemia, and is sensitive to all currently used antimalarial drugs. It was obtained from Nigerian Institute of Medical Research (NIMR), Lagos, Nigeria.

Animals: Animal experiments were carried out according to the Principles of Laboratory Animal Care and legislation in force in Nigeria. Eperythrozoon-free Swiss albino mice (CD1) weighing 20 to 25 g were obtained from Department of Pharmacology and Toxicology, University of Nigeria.


Extraction of goat fat: The lipid (goat fat) used in the formulation was first extracted from goat fat (Capra hircus) using wet rendering method [14]. Briefly, adipose tissue of goat was grated and subjected to moist heat by boiling with about half its weight of water in a water bath for 45 min. The molten fat was separated from the aqueous phase after filtering with a muslin cloth and stored in a refrigerator until used.

Preparation of lipid matrix: The lipid matrix was prepared using fusion technique according to Friedrich et al. Attama et al. [15,16]. Briefly, a 70 g quantity of the prepared goat fat was weighed (Adventurer, Ohaus, China), melted in a beaker placed in a water bath at a temperature of 60°C;. Thereafter 30g of P90H was added to the melted goat fat and stirred using a magnetic stirrer and hot plate (Jenway 400, EU), until an even mix was obtained. The molten lipid matrix was then placed in a cold water bath for 30 min at room temperature until solidification to obtain the Solidified Reverse Micellar Solution (SRMS).

Preparation of SLMs: The Solid Lipid Microparticles were prepared to contain: lipid matrix (17 % w/w), CQ (0, 3, 5, 7 %w/w), polysorbate 80 (1.5 %), sorbic acid (0.05 %), sorbitol (4 %w/w) and water (to 100 %w/w). The lipid matrix consisted of goat fat and P90H. For each batch, the lipid matrix was placed in a stainless steel bowl and heated at 60°C; until it had melted completely. The remaining excipients were weighed out appropriately and mixed with the corresponding quantity of water at 70 °C;. The excipients mixture with water at 70oC was poured into the lipid matrix to form the lipid matrix-mixture and homogenized at 5000 rpm for 10 min with an Ultra-Turrax homogenizer (IKA® T25, Basic Digital, Germany). The hot emulsion was then poured into a bottle and allowed to recrystallize at room temperature for 24 h and the resultant unloaded SLMs batch C1 obtained. The same procedure was adopted for the CQ-loaded SLMs with varying quantity of the drug (concentrations of 3 %, 5 %, and 7 % for batches C2, C3, and C4 respectively) as shown in (Table 1), except that the drug was poured into the melted matrix and mixed, thereafter the excipients mixture was poured into the lipid matrix-drug mixture. The SLMs obtained after cooling at room temperature were lyophilized to obtain water-free SLMs using a freeze-dryer (Amsco/ Finn-Aqua Lyovac GTZ, Germany) [13,17]. Briefly, lyophilisates of the SLMs are obtained by freezing the formulations at a pressure of 2.7 Pa and temperature of -30 °C; followed by sublimation and drying at 15-25 °C. All these operations took 6-12 h.