A Comparative Study of Oral and Topical Administration of Hesperidin Lipid Nanoparticles in Rheumatoid Arthritis

Research Article

Austin Arthritis. 2016; 1(2): 1010.

A Comparative Study of Oral and Topical Administration of Hesperidin Lipid Nanoparticles in Rheumatoid Arthritiss

Bhalekar MR*, Madgulkar AR, Aswar M, Mariam G and Desale P

Department of Pharmaceutics, AISSMS College of Pharmacy, Kennedy Road, Near RTO, Pune411001, India

*Corresponding author: Mangesh R Bhalekar, AISSMS College of Pharmacy, Kennedy Road, Near RTO, Pune 411001, India

Received: June 24, 2016; Accepted: August 10, 2016; Published: August 12, 2016


Rheumatoid Arthritis (RA) is a chronic, inflammatory autoimmune disease causing synovial proliferation and joint destruction. Hesperidin, a citrus flavanoid has been reported to be effective on reduction of Tumor Necrosis Factor-alpha, key inflammatory mediator in RA. Hesperidin Solid Lipid Nanoparticle (SLN) were prepared and optimised by 32 factorial design using emulsification- probe sonication method. The optimised SLN were found to be spherical in shape with particle size of 279nm, zeta potential of 17.86mV and 29.60% drug content. A comparative study in arthritis induced rats using Complete Freund’s adjuvant (CFA) was performed. The outcome of radiography, histopathology and ELISA study were employed to evaluate the effect of formulation on male wistar rats. Hence topical and oral SLN formulation was successful in treatment of rheumatoid arthritis. But the comparative results of both the formulations so obtained did not differ significantly.

Keywords: Hesperidin; Rheumatoid arthritis; Solid lipid nanoparticle


RA: Rheumatoid Arthritis; SLN: Solid Lipid Nanoparticle; CFA: Complete Freund’s Adjuvant; TNF- α: Tumor Necrosis Factor- Alpha


Rheumatoid arthritis is a chronic, progressive inflammatory autoimmune disease causing synovial inflammation and joint destruction [1]. Pain, fever, malaise, stiffness and swelling of joints constitute the primary symptoms of RA. Synovitis is caused by the release of pro-inflammatory cytokines like TNF-α, IL-6 and IL-1 which further leads to cartilage destruction, bone erosion and finally joint deformities. TNF-α being a key inflammatory mediator triggers an increase in synovial proliferation and also production of other secondary mediators. Overproduction of TNF-α in affected joints causes an increase in synoviocyte proliferation and a cascade of secondary mediators involved in the recruitment of inflammatory cells which leads to joint destruction [2]. TNF-α also induces osteoclast differentiation. Natural flavanoids like Tridax Procumbens Flavonoid (TPFs) has been reported to reduce differentiation of osteoclast [3]. Although the specific cause of RA is unknown up till now but it is reported that patients suffering from this disease share a common amino acid sequence in an antigen–presenting region of Major Histocompatability Complex (MHC) class II human leukocyte antigen molecules are more susceptible to RA. The interference of this complexes results in activation and proliferation of T-cells or Tlymphocytes. Major role of T-cell in RA is stimulation of synovial cells (macrophage like synoviocytes, dendritic cells, fibroblastic synoviocytes and B lymphocytes) and also release of pro-inflammatory cytokines causing destruction of bones and cartilage [4]. IL-2 being one of the pro-inflammatory cytokines regulates the activation of T cells [5]. Steroids, NSAID’s, immunosuppressant’s, DMARD etc. are the mainstay treatment strategies for RA but have their own share of side effects and toxicities which has lead to consideration of powerful natural agents as alternative treatment strategy [6].

Natural products play a key role in discovery of several therapeutic substances. According to estimate by World Health Organisation, 80% of world’s population relies on traditional medicine [7].

Hesperidin is a bioflavonoid obtained from orange peel Citrus sinesis family Rutaceae. The flavanone glycoside hesperidin is composed of hesperetin (flavanone) and rutinose (disaccharide). Hesperidin deficiency in body may cause capillary leakiness, night time leg cramps. The pharmacological property of hesperidin includes anti inflammatory, antioxidant, hypolipidaemic, and anti carcinogenic and is also used in the treatment of varicose vein [8]. Hesperidin also possesses anti- rheumatic action. At dose of 160mg/ kg hesperidin causes amelioration of joint destruction, synovial hyperplasia and infiltration of inflammatory cells. In adjuvant arthritis rat (CFA induced rats) hesperidin inhibited the production of IL- 1, IL-6, and TNF-α and restored the suppression of T- lymphocyte proliferation and production of IL-2 at the same time [6]. Hesperidin also exhibits significant anti-inflammatory activity by modulating the prostaglandin synthesis and COX-2 gene expression pathways [9].

The uptake of hesperidin flavonoid both orally and topically is not possible as it possess very limited water solubility, poor bioavailability and also poor permeability. Nanocarrier’s especially solid lipid nanoparticles could be helpful to enhance bioavailability and bioefficacy of this flavonoid [10]. In RA lymphatic uptake of drug plays a major role in the treatment of this disease [11]. SLN being nanosized and lipidic in nature will be one of the best candidate for the lymphatic uptake of the drug to the inflamed synovium of the arthritic joint. Complete Freund’s Adjuvant (CFA) model is composed of inactivated and dried mycobacteria when injected subcutaneously or intraperitoneally stimulate production of TNF-α which results in infiltration of the synovial membrane [12].

SLN are the submicron colloidal carrier composed of a solid lipid core in which lipophilic drug is encapsulated. They are biocompatible and biodegradable in nature and bear potential to carry both lipophilic and hydrophilic drug. It has superiority over other colloidal nanocarrier in terms of control over release kinetics of drug, enhanced drug stability and bioavailability of entrapped drug [13]. Present work is an attempt at formulation optimization of hesperidin solid lipid nanoparticles and in-vivo evaluation of its activity upon oral as well as topical administration.

Materials and Methods


Hesperidin (≥80%) and Complete Freund’s Adjuvant (CFA) were purchased from Sigma-Aldrich. Rat TNF -α ELISA kit was purchased from Ray Biotech. All other excipients used were of analytical standards.


Preparation of hesperidin SLN dispersion: Hesperidin loaded Solid Lipid Nanoparticles (SLN) dispersion was prepared using emulsification – probe sonication method [14]. The lipid (stearic acid) and mixture of hesperidin and dimethyl formamide (DMF) were heated to 10°C above the melting point of lipid (70°C). The aqueous phase was prepared by dissolving tween 80 and distilled water and heated to the same temperature of the oil phase. Both the phases were mixed together and stirred at 500rpm using a mechanical stirrer for 15 minutes maintaining the above mentioned temperature. The resultant dispersion were subjected for probe sonication (Sonics vibra 750) for 30 minutes which was further evaluated for particle size, zeta potential, drug content etc. The concentration of lipid and surfactant were finalised using design expert 9.1 software. The composition of SLN dispersion is shown in (Table 1).