GC-MS profiling and assessment of antioxidant, antibacterial, and anticancer properties of extracts of Annona squamosa L. leaves.

BACKGROUND: The research and application of plants in food supplements and drugs have attracted great interest. This study aimed to examine the efficiency of several solvents for the extraction of the main compounds from Annona squamosa leaves and to evaluate the antioxidant, antibacterial, and anticancer activities of these extracts. METHODS: Gas chromatography-mass spectrometry was used to screen the bioactive compounds of A. squamosa methanolic extract. The free radical, hydrogen peroxide, and nitric oxide scavenging activities of the extracts were investigated. Furthermore, MTT, nuclear staining, LDH, and monolayer wound repair assays were performed to evaluate the potential anticancer activity of the extracts in colon cancer cells while the antibacterial activity was tested by using a well diffusion assay. RESULTS: A. squamosa leaves extracts were found to contain several bioactive compounds, of which the majority were sesquiterpenes (C15H24). These extracts exhibited strong antioxidant activity and antibacterial potency against both gram-positive and gram-negative bacteria. Different A. squamosa leaves extracts displayed remarkable antiproliferative, cytotoxic, antimigration, and apoptotic activities in colon cancer cells. CONCLUSIONS: A. squamosa leaves contain major bioactive compounds that inhibit the growth of several types of bacteria and colon cancer cell lines, which demonstrated their efficacy as an alternative source of antibiotics and for the development of novel drugs for colon cancer therapy.

Differential Effects of Camel Milk on Insulin Receptor Signaling - Toward Understanding the Insulin-Like Properties of Camel Milk.

Previous studies on the Arabian camel (Camelus dromedarius) showed beneficial effects of its milk reported in diverse models of human diseases, including a substantial hypoglycemic activity. However, the cellular and molecular mechanisms involved in such effects remain completely unknown. In this study, we hypothesized that camel milk may act at the level of human insulin receptor (hIR) and its related intracellular signaling pathways. Therefore, we examined the effect of camel milk on the activation of hIR transiently expressed in human embryonic kidney 293 (HEK293) cells using bioluminescence resonance energy transfer (BRET) technology. BRET was used to assess, in live cells and real-time, the physical interaction between hIR and insulin receptor signaling proteins (IRS1) and the growth factor receptor-bound protein 2 (Grb2). Our data showed that camel milk did not promote any increase in the BRET signal between hIR and IRS1 or Grb2 in the absence of insulin stimulation. However, it significantly potentiated the maximal insulin-promoted BRET signal between hIR and Grb2 but not IRS1. Interestingly, camel milk appears to differentially impact the downstream signaling since it significantly activated ERK1/2 and potentiated the insulin-induced ERK1/2 but not Akt activation. These observations are to some extent consistent with the BRET data since ERK1/2 and Akt activation are known to reflect the engagement of Grb2 and IRS1 pathways, respectively. The preliminary fractionation of camel milk suggests the peptide/protein nature of the active component in camel milk. Together, our study demonstrates for the first time an allosteric effect of camel milk on insulin receptor conformation and activation with differential effects on its intracellular signaling. These findings should help to shed more light on the hypoglycemic activity of camel milk with potential therapeutic applications.

The Arabian camel Camelus dromedarius heat shock protein 90α: cDNA cloning, characterization and expression.

Heat shock protein 90 (Hsp90) is a highly conserved ubiquitous molecular chaperone contributing to assisting folding, maintenance of structural integrity and proper regulation of a subset of cytosolic proteins. In the present study, a heat shock protein 90α full length coding cDNA was isolated and cloned from the Arabian one-humped camel by reverse transcription polymerase chain reaction (RT-PCR). The full length cDNA sequence was submitted to NCBI GeneBank under the accession number KF612338. The sequence analysis of the Arabian camel Hsp90α cDNA showed 2202bp encoding a protein of 733 amino acids with estimated molecular mass of 84.827kDa and theoretical isoelectric point (pI) of 5.31. Blast search analysis revealed that the C. dromedarius Hsp90α shared high similarity with other known Hsp90α. Comparative analyses of camel Hsp90α protein sequence with other mammalian Hsp90s showed high identity (85-94%). Heterologous expression of camel Hsp90α cDNA in E. coli JM109 (DE3) gave a fusion protein band of 86.0kDa after induction with IPTG for 4h.

Purification and functional characterization of pancreatic insulin from camel (Camelus dromedarius).

Large-scale production of insulin still represents the key step in helping diabetic patients throughout the world. Many species and approaches have been used for the production of insulin. In this study, we purified and characterized for the first time pancreatic insulin from the Arabian camel (Camelus dromedarius) using a modified acid-alcohol extraction method. After extraction insulin was purified using a one-step gel filtration on a Sephadex G-50 column leading to a purification yield of 80 mg/kg (20%) of camel pancreas. The purity of camel insulin was assessed by SDS-PAGE and HPLC using insulin from human, bovine and porcine as standards. Molecular weight was determined for purified camel insulin as 5800 Daltons and its amino acid composition is similar to that known for other species. The functional characterization of purified crude camel insulin was demonstrated in vitro by positive competition by radioimmunoassay and in vivo showing camel insulin inducing acute hypoglycaemia in mice. Together, our study reports for the first time the successful purification of functional insulin from camel pancreas with similar properties compared to other insulin species. This is of great interest given that the camel represents considerable economic worth in many countries.

Purification and characterization of membrane-bound peroxidase from date palm leaves (Phoenix dactylifera L.).

Peroxidase from date palm (Phoenix dactylifera L.) leaves was purified to homogeneity and characterized biochemically. The enzyme purification included homogenization, extraction of pigments followed by consecutive chromatographies on DEAE-Sepharose and Superdex 200. The purification factor for purified date palm peroxidase was 17 with 5.8% yield. The purity was checked by SDS and native PAGE, which showed a single prominent band. The molecular weight of the enzyme was approximately 55 kDa as estimated by SDS-PAGE. The enzyme was characterized for thermal and pH stability, and kinetic parameters were determined using guaiacol as substrate. The optimum activity was between pH 5-6. The enzyme showed maximum activity at 55 °C and was fairly stable up to 75 °C, with 42% loss of activity. Date palm leaves peroxidase showed K m values of 0.77 and 0.045 mM for guaiacol and H2O2, respectively. These properties suggest that this enzyme could be a promising tool for applications in different analytical determinations as well as for treatment of industrial effluents at low cost.