PLGA nanoparticles loaded with the antileishmanial saponin ß-aescin: factor influence study and in vitro efficacy evaluation.

Colloidal carriers are known to improve the therapeutic index of the conventional drugs in the treatment of visceral leishmaniasis (VL) by decreasing their toxicity whilst maintaining or increasing therapeutic efficacy. This paper describes the development of poly(d,l-lactide-co-glycolide) (PLGA) nanoparticles (NPs) for the antileishmanial saponin ß-aescin. NPs were prepared by the W/O/W emulsification solvent evaporation technique and the influence of five preparation parameters on the NPs' size (Z(ave)), zeta potential and entrapment efficiency (EE%) was investigated using a 2(5-2) fractional factorial design. Cytotoxicity of aescin, aescin-loaded and blank PLGA NPs was evaluated in J774 macrophages and non-phagocytic MRC-5 cells, whereas antileishmanial activity was determined in the Leishmania infantum ex vivo model. The developed PLGA NPs were monodispersed with Z(ave)<500 nm and exhibited negative zeta potentials. The process variables 'surfactant primary emulsion', 'concentration aescin' and 'solvent evaporation rate' had a positive effect on EE%. Addition of Tween 80 to the inner aqueous phase rendered the primary emulsion more stable, which in its turn led to better saponin entrapment. The selectivity index (SI) towards the supporting host macrophages increased from 4 to 18 by treating the cells with aescin-loaded NPs instead of free ß-aescin. In conclusion, the in vitro results confirmed our hypothesis.

Solid lipid nanoparticle (SLN) formulations as a potential tool for the reduction of cytotoxicity of saponins.

The present pilot study explored the potential of solid lipid nanoparticles (SLN) to entrap saponins and reduce the membrane toxicity of these compounds. SLN composed of different types of solid lipid were prepared by the cold homogenisation technique. Combinations of anionic, cationic and non-ionic stabilisers were selected in order to obtain negatively, positively and neutrally charged SLN. Mean particle size and zeta potential of blank and saponin-loaded formulations were measured by Dynamic Light Scattering (DLS), Electrophoretic Light Scattering (ELS) and in vitro cytotoxicity on MRC-5 SV2 and J774 cells was assessed using a resazurin-based assay. The type of solid lipid used for the formulation influenced the mean particle size, while the zeta potential mainly depended on the kind of surfactant utilised. Blank SLN composed of hard fat and anionic or non-ionic surfactants did not result in cytotoxicity. After loading with saponin, the anionic hard fat SLN was found to be the optimal formulation.

Occupational asthma related to aescin inhalation.

BACKGROUND: More than 400 agents are recognized as causes of occupational asthma, a work-related disease that can be induced by an immunologic or a nonimmunologic mechanism. OBJECTIVE: To describe a 57-year-old man employed in the pharmaceutical industry who developed bronchial asthma while working with products such as Plantago ovata and aescin, an active ingredient with anti-inflammatory and venotonic properties. METHODS: Various tests were performed, including radiography, total serum IgE titer measurements, skin tests against common pneumoallergens and Plantago species, pulmonary function studies, a methacholine test, and specific inhalation challenge with P. ovata and aescin. RESULTS: The results of these tests, including specific inhalation challenge, confirmed the diagnosis of occupational asthma due to aescin exposure, whereas the results of specific challenge with P. ovata, a known cause of occupational asthma, were negative. CONCLUSIONS: Aescin may represent a new causative agent of occupational asthma in personnel working in the pharmaceutical industry. The mechanism by which aescin can produce asthma is unknown, but analysis of the characteristics of our patient suggests a non-IgE immunologic mechanism, although an irritative mechanism secondary to long-term low-level exposure to aescin, a possible irritant, cannot be ruled out.

Potential carcinogenicity assessment of alpha-aescin and phenbendasol.

Potential carcinogenic activity of alpha-aescin and phenbendasole made by "Polfa" (Poland) as well as phenbendasole produced by "Hoechst" (Germany) was studied using Salmonella/microsome test, DNA repair test and micronucleus assay. None of tested preparations were mutagenic or genotoxic what suggest that none of them possess potential carcinogenic activity. Besides, it was established that alpha-aescin exhibits strong and phenbendasol weak acute systemic toxicity for mice. Alpha-aescin and phenbendasole produced in Poland have been found to be toxic for bone marrow cells of mice but only when administered at a high dose of 80% LD50.

[Studies on toxicity and fertility of escin in the rat (author's transl)]. / Toxizitäts- und Fertilitätsstudien mit Aescin bei der Ratte.

The DL50 of escin was determined after i.p. application. Juvenile male rats were treated with 2X5 mg/kg escin at age 32 days. After they had reached fertility, kidneys, testes and sperm were examined. The high dose of escin used did not affect fertility and a nephrotoxic activity could not be detected either.

Animal experiments on the question of the renal toleration of the horse chestnut saponin aescin.

The possibility that aescin might have a nephrotoxic side effect has been investigated by clearance studies in kidneys of healthy rats and by toleration studies in rats with damaged kidneys. The effect of aescin, both free and albumin-bound, on renal tubular transport processes was studied in the model of the isolated, artificially perfused frog kidney. The rates at which different concentrations of aescin were bound to rat plasma proteins were determined in vitro. The clearance of i.v. aescin was 13% of creatinine clearance and 7% of p-aminohippurate (PAH) clearance; this rules out the tubular secretion of aescin. No deaths occurred among aminonucleoside-damaged rats given i.v. sodium aescinate 2.2 mg/kg, but rats damaged with mercuric chloride or uranyl nitrate had exactly the same mortality rate as those given 2.2 mg/kg i.v. of sodium aescinate alone. The rats received four injections in all of aescin 0.35 mg/kg i.v., given at intervals of two days. Aescin had no effect on renal damage caused by aminonucleoside, mercuric chloride or uranyl nitrate. Aescin concentrations of 0.2 mg/l and 2.0 mg/l in the perfusion fluid increased the excretion of Na+ and glucose by the frog kidney and reduced the reabsorption of both these substances. With a sodium aescinate concentration of 5 mg/l the production of urine ceased. When 1% (w/v) of albumin was added to the perfusion fluid, even sodium aescinate 5 mg/l had no effect on the tubular transport of Na+, glucose and water. The fact that about 50% of aescin was not bound to plasma protein in vitro suggests that some of the small amount of aescin in the glomerular filtrate is reabsorbed in the tubules.

[The prevention and treatment of post-operative and post-traumatic oedema. Chemical laboratory tests on the renal tolerance of beta-aescin (reparil) (author's transl)]. / Postoperative und posttraumatische Odemprophylaxe und-therapie. Laborchemische Untersuchungen über die Nierenverträglichkeit von beta-Aescin (Reparil)

Renal tolerance of the preparation beta-aescin (Reparil) was tested in cases of oedema following fresh soft tissue crush injuries of the extremities and surgical and reconstructive operations on the hand. Forty in-patients with healthy kidneys were treated with 10 mg beta-aescin intravenously twice daily for six days after operations on the hand and after accident surgery. During this period the following tests were carried out every day and two days after cessation of treatment: urea content in urine and serum, creatinine in serum, creatinine clearance, albumin in urine, urine osmolarity and blood and urine sugar. All values obtained were within the normal range. No impairment of total renal function was observed. Taking the stated dosage instructions and contra-indications into consideration, beta-aescin can thus be administered without hesitation.