Chemometric assisted natural DES based VA-DLLME-LC-MS/MS method for the quantitative determination of Garcinol in biofluids/tissues: A practical application to pharmacokinetics and biodistribution studies.

Garcinol (GAR) is a polyisoprenylated benzophenone obtained from Garcinia indica used as anti-oxidant and anti-inflammatory in traditional medicine and due to these activities, it possesses anticancer properties. It is considered to be a next generation epigenetic drug. A green solvent based analytical method which is efficient, sophisticated, and highly enriched has been developed for the quantitative analysis of GAR in biological samples (plasma, liver, kidney and spleen) with the use of deep eutectic solvent (DES) for its extraction. A series of 23 DESs were synthesized and out of which, Thymol (Th)-Terpeniol (T), 2:1 molar ratio with a more hydrophobic environment and high interaction efficiency between GAR and DES was identified for the better extraction from mice plasma and tissue samples. The Design of Experiment approaches like placket-burmann design and central composite design were used to optimize the method conditions. The method validation characteristics, such as limit of detection (0.193-0.237 ng/mL), limit of quantification (0.644-0.697 ng/mL), lower limit of quantification (0.5 ng/mL), broad range of linearity with R2 (0.9994-0.9997) with a percent recovery not less than 87% was observed, which are well within the acceptance criteria for a bioanalytical method. The enrichment factor is upto 53-60 folds, with high extraction efficiency (89-97%). The measurement uncertainty was estimated with an expanded uncertainty ranged between 10.9%-19.0%. The method developed and validated was effectively applied to examine the pharmacokinetic and biodistribution patterns for GAR in mice.

Clastogenic, aneugenic, and tubulin polymerization properties of di-(2-ethylhexyl) phthalate and dibutyl phthalate.

Phthalate compounds were found to disrupt the endocrine system and alter transcriptomes during human embryonic development. In our previous work, we have isolated and reported two such phthalates di-(2-ethylhexyl) phthalate (DEHP) and dibutyl phthalate (DBP) from Brevibacterium mcbrellneri bacteria and evaluated their bioactive properties. Naturally derived phthalates might be less toxic compared with synthesized molecules. We have investigated biologically isolated phthalates to understand the possible genotoxic effects in mice and further investigated in silico binding and polymerization of ß-tubulin. Three sub-lethal concentrations of DEHP (150 µM, 175 µM, and 200 µM) and DBP (10 µM, 15 µM, and 30 µM) were studied. The results showed that the phthalates were found to be highly genotoxic in nature. However, the pattern of genotoxic effects was not found to be dose-dependent in the induction of chromosome aberrations (CA), micronuclei (MN), and changes in the mitotic index (MI) in cells. In silico studies of phthalates on polymerization of ß-tubulin suggested that both DBP and DEHP were able to interact with the hydrogen bonds and make strong van der Waals interactions with ß-tubulin thereby possibly causing destabilization of microtubule network. Our study suggests that these phthalates might be playing an important role in normal cell division thereby showing highly genotoxic effects.

Anti-hyperglycemic and genotoxic studies of 1-O-methyl chrysophanol, a new anthraquinone isolated from Amycolatopsis thermoflava strain SFMA-103.

The compound 1-O-methyl chrysophanol (OMC) which belongs to a class of hydroxyanthraquinones was isolated from Amycolatopsis thermoflava strain SFMA-103 and studied for their anti-diabetic properties. OMC was evaluated as an anti-diabetic agent based on in silico studies which initially predicted the binding energy with α-amylase (-188.81 KJ mol-1) and with α-glucosidase (70.53 KJ mol-1). Further, these results were validated based on enzyme inhibition assays where OMC demonstrated enzyme inhibitory activity towards α-amylase (IC50 3.4 mg mL-1) and α-glucosidase (IC50 38.49 µg mL-1). To confirm the anti-diabetic activity, in vivo studies (oral dose in Wistar rats) revealed that OMC inhibited significantly the increase in glucose concentration at 100 mg/kg as compared to starch control (p < 0.05). Further, to understand the safety of OMC as a therapeutic agent, the genotoxic analysis was performed in both in vitro Chinese Hamster Ovary cells (250, 500, and 1000 µM/mL) and in vivo Swiss albino mice (250, 500, and 1000 mg/kg). In vitro results showed that OMC concentration of up to 250 µM/mL did not elicit significant changes in CAs, MI, and MN counts in CHO cells. Similarly, in mice experiments (i.p. injection), no significant changes in CAs, MI, and MN induction were observed till 500 mg/kg of OMC when compared with chrysophanic acid (Cy) (200 mg/kg). In addition, mice that received the lowest dose of OMC (250 mg/kg) did not show any histological changes in liver, kidney, and heart. The study concluded that five times higher therapeutic dose (100 mg/kg) of OMC can be utilized against hyperglycemia with no genotoxic effects.

Efficient anti-tumor nano-lipoplexes with unsaturated or saturated lipid induce differential genotoxic effects in mice.

Cationic lipids are well-known excipients for nanometric liposomal gene delivery systems. However, because of the suspected, collateral toxicity in normal cells, the use of cationic lipids for the treatment of human tumor is largely limited. Recently, we developed a glucocorticoid receptor (GR)-targeted liposomal, anticancer delivery system (DXE nano-lipoplex), which carried cationic lipid of saturated twin aliphatic chains. It exhibited efficient anti-tumor effect in aggressive and drug-resistant tumor models. Toward exploring lipoplex's human clinical use, we incorporated another nano-lipoplex (D1XE) group that carried cationic lipid with one of its aliphatic chain carrying unsaturation and compared in vivo genotoxicological profiling-based safety assessment and the respective anti-tumor efficacy of the lipoplexes. Thus, both the lipoplexes differ only by the chemical identity of one of their constituent cationic lipid. Unsaturated aliphatic chains in lipid generally impart efficient cell surface fusogenic property in lipid formulations. Herein, we report that nanoplex with unsaturated cationic lipid (D1XE) exhibited better physical appearance with less flocculent behavior than nanoplex with saturated lipid (DXE). Upon multiple injections, D1XE nanoplex imparted better tumor regression but most importantly, exhibited much lower overall toxicity (e.g. genotoxicity, weight loss, etc.) than DXE nanoplex. With a higher antitumor effect but a lower genotoxic effect, D1XE is proved to be a better nanoplex than DXE for the potential clinical trial. Thus, this study clearly delineates the importance of incorporating a constituent lipid that carries a single unsaturated aliphatic chain toward developing efficient anti-tumor nano-lipoplexes with reduced genotoxicity.