Pyronaridine tetraphosphate

Artesunate, mefloquine hydrochloride, amodiaquine dihydrochloride dihydrate, chloroquine diphosphate, primaquine phosphate, quinine hemisulfate, atovaquone, methylene blue, pyronaridine tetraphosphate, doxycycline hyclate, clindamycin and praziquantel were purchased from Sigma-Aldrich. Ferroquine was obtained from Sanofi-Synthelabo, proguanil and cycloguanil from Jacobus Pharmaceutical Company, tigecycline from Wyeth, and mirincamycin hydrochloride enantiomers from Maldevco [45 (link)]. All compounds were dissolved in sterile DMSO except for quinine for which methanol was used and pure M199 medium (without additives) was used to dissolve proguanil, cycloguanil, clindamycin, and pyronaridine. The stock concentration was 50 mM for Artesunate, amodiaquine, chloroquine, atovaquone, quinine, and primaquine, and 100 mM for praziquantel, proguanil, cycloguanil, methylene blue, pyronaridine, clindamycin, doxycycline, and mirincamycin enantiomers, respectively. Mefloquine was dissolved to 24 mM and Ferroquine to 12.5 mM. All stocks were freshly prepared for the study and stored at -20°C. Maximum concentration of the solvent (DMSO, methanol) in the in vitro assays did not exceed 0.8% and did not interfere with parasite viability.

SYBR Green I (SG I) nucleic acid stain (Lonza, USA; 10,000x) was stored at −20 °C and thawed before use. A lysis buffer consisting of Tris (130 mM; pH 7.5), Ethylenediaminetetraacetic acid (EDTA) (10 mM), saponin (0.016%; W/V), and TritonX-100 (1.6%; V/V) was prepared in advance and stored at 4 °C. Diminazene aceturate (Ciba-Geigy Japan Limited, Japan), Pyronaridine tetraphosphate (Sigma-Aldrich Japan) and allicin (iHerb.com) were used in this study to evaluate the validity of fluorescence-based assays for antibabesial drug screening in mice.

SYBR Green I (SGI) nucleic acid stain (Lonza, USA; 10,000x) was stored at -20°C and thawed before use. A lysis buffer consisting of Tris (130 mM; pH 7.5), EDTA (10 mM), saponin (0.016%; W/V), and TritonX-100 (1.6%; V/V) was prepared in advance and stored at 4°C. Diminazene aceturate (Novartis, Japan), luteolin (Sigma-Aldrich, Japan), pyronaridine tetraphosphate (Sigma-Aldrich), nimbolide (BioVision, USA), gedunin (Tocris Bioscience, UK), and enoxacin (Sigma-Aldrich) were prepared as 100 mM or 50 mM stock solutions.