Purpose Leishmaniasis is an emerging infection affecting more than one billion people worldwide in endemic areas, including Pakistan. Amphotericin B (AmB) is a class IV antibiotic, characterized by poor water solubility and low membrane permeability, and is commonly used for the systemic treatment of various infections, including leishmaniasis, but is associated with a long list of side effects. Since crystal engineering can alter the physicochemical features of active pharmaceutical substances, the approach has been successfully utilized to address the issue of poor solubility and toxicity of AmB with the aim of improving anti-leishmanial potentialaginst both intracellular and extracellular forms of parasites. Method Six co-crystals (2–7) of Amphotericin B (AmB, 1) were successfully synthesized by adopting reflux approach to investigate their effects on the solubility of (AmB) and anti-leishmanial potential. The combined approach of using FTIR and powder XRD, DSC and TGA was utilized to support the formation of co-crystals. The solubility of these co-crystals was found to be significantly improved in various dissolution media, including pH 6.8 and 7.4 phosphate buffers and distilled water. The co-crystals 2–7 were evaluated for anti-leishmanial activity against promastigotes of L. major, and L. tropica, in vitro and amastigotes stages of L. major parasite by using J774.2 cell line (murine macrophage) cell line (J774) followed by AFM analysis to examine the changes in infected macrophage cells. Conclusion and result The synthesized co-crystals (2–7) showed potent anti-leishmanial potential against L. major (IC50 = 1.43 ± 0.04–1.67 ± 0.02 µM), and L. tropica (IC50 = 1.42 ± 0.05–1.80 ± 0.08 µM) promastigotes (extracellular/in blood parasite stage), in comparison to standard drugs Miltefosine (IC50 = 31.8 ± 0.2 µM), Pentamidine (IC50 = 9.23 ± 0.06 µM) and Amphotericin B (IC50 = 1.97 ± 0.06 µM)for L. major and Miltefosine (IC50 = 27.2 ± 0.6 µM), Pentamidine (IC50 = 9.22 ± 0.04 µM), and Amphotericin B (IC50 = 1.68 ± 0.04 µM) for L. tropica. An infected macrophage cellular assay, along with AFM analysis, disclosed that synthesized co-crystals are also effective against the L. major amastigotes (intracellular/in macrophage cell stage) form with no signs of cytotoxicity. The promising anti-leishmanial and improved aqueous solubility of Amphotericin B due to co-crystallization, demonstrating the remarkable abilities of crystal engineering approach to modulate medicinal efficacy of APIs already in market.

Crystal Engineering of Amphotericin B to Obtained New Co-Crystals with Improved Solubility, Cytotoxicity, and Anti-Leishmanial Activity (Amastigotes and Promastigotes)

期刊论文
发表日期：

2026年4月1日

查看全文

Urooj Iqbal,
M. Iqbal Choudhary,
Sammer Yousuf

摘要

Purpose

Leishmaniasis is an emerging infection affecting more than one billion people worldwide in endemic areas, including Pakistan. Amphotericin B (AmB) is a class IV antibiotic, characterized by poor water solubility and low membrane permeability, and is commonly used for the systemic treatment of various infections, including leishmaniasis, but is associated with a long list of side effects. Since crystal engineering can alter the physicochemical features of active pharmaceutical substances, the approach has been successfully utilized to address the issue of poor solubility and toxicity of AmB with the aim of improving anti-leishmanial potentialaginst both intracellular and extracellular forms of parasites.

Method

Six co-crystals (2–7) of Amphotericin B (AmB, 1) were successfully synthesized by adopting reflux approach to investigate their effects on the solubility of (AmB) and anti-leishmanial potential. The combined approach of using FTIR and powder XRD, DSC and TGA was utilized to support the formation of co-crystals. The solubility of these co-crystals was found to be significantly improved in various dissolution media, including pH 6.8 and 7.4 phosphate buffers and distilled water. The co-crystals 2–7 were evaluated for anti-leishmanial activity against promastigotes of L. major, and L. tropica, in vitro and amastigotes stages of L. major parasite by using J774.2 cell line (murine macrophage) cell line (J774) followed by AFM analysis to examine the changes in infected macrophage cells.

Conclusion and result

The synthesized co-crystals (2–7) showed potent anti-leishmanial potential against L. major (IC₅₀ = 1.43 ± 0.04–1.67 ± 0.02 µM), and L. tropica (IC₅₀ = 1.42 ± 0.05–1.80 ± 0.08 µM) promastigotes (extracellular/in blood parasite stage), in comparison to standard drugs Miltefosine (IC₅₀ = 31.8 ± 0.2 µM), Pentamidine (IC₅₀ = 9.23 ± 0.06 µM) and Amphotericin B (IC₅₀ = 1.97 ± 0.06 µM)for L. major and Miltefosine (IC₅₀ = 27.2 ± 0.6 µM), Pentamidine (IC₅₀ = 9.22 ± 0.04 µM), and Amphotericin B (IC₅₀ = 1.68 ± 0.04 µM) for L. tropica. An infected macrophage cellular assay, along with AFM analysis, disclosed that synthesized co-crystals are also effective against the L. major amastigotes (intracellular/in macrophage cell stage) form with no signs of cytotoxicity. The promising anti-leishmanial and improved aqueous solubility of Amphotericin B due to co-crystallization, demonstrating the remarkable abilities of crystal engineering approach to modulate medicinal efficacy of APIs already in market.