An Effective Method of Isolating Honey Proteins

Honey is a natural sweetener composed mostly of sugars, but it contains also pollen grains, proteins, free amino acids, and minerals. The amounts and proportions of these components depend on the honey type and bee species. Despite the low content of honey protein, they are becoming a popular study object, and have recently been used as markers of the authenticity and quality of honey. Currently, the most popular methods of protein isolation from honey are dialysis against distilled water, lyophilization of dialysate, or various precipitation protocols. In this work, we propose a new method based on saturated phenol. We tested it on three popular polish honey types and we proved its compatibility with both 1D and 2D polyacrylamide gel electrophoresis (PAGE) and MS (mass spectrometry) techniques. The elaborated technique is also potentially less expensive and less time-consuming than other previously described methods, while being equally effective.

Assessment of Programmed Cell Death of Aspergillus flavus by Triggered Cysteine-dependent Aspartate-directed Proteases (Meta-caspase3) Lethality Mechanism of Novel Compounds Isolated from Ethyl Acetate Extract of Spondias mombin

Spondias mombin is a plant that has been traditionally noted for its medicinal with a preliminary results report a wide range of antibacterial and antifungal properties. Meta-caspases and Caspases are essential in cells for programmed cell death, in development and most other stages of adult life, and have been termed "executioner" proteins for their roles in the cell. A 12 hours old culture of each microorganism was re-suspended in plant extract at 1000 µg mL in a total volume of 500 µl for 0, 15, 30, 45, 60, and 180 minutes. The cells were pelleted by centrifugation at 5000 g for 5 minutes. The pellets were rinsed twice in phosphate buffer saline (PBS). Then 1/10 volume of 95% ethanol plus 5% saturated phenol were added to the pellets to stabilize cellular RNA. The cells were then re-harvested by centrifugation (8200 g, 4°C and 2 minutes). The supernatant was aspirated and pellets re-suspended in 800 μl of lysis buffer (10 mMTris, adjusted to pH 8.0 with HCl, 1 mM EDTA) and 8.3 U/ml Ready-LyseTM Lysozyme Solution. After the pellets were re-suspended, 80 μl of a 10% SDS solution was added, mixed and incubated for 2 minutes at 64 °C. Then 88 μl of 1 M NaOAc (pH 5.2) was mixed with the lysate followed by an equal volume of water and saturated phenol was added. Total RNA was quantified using Spectrophotometric absorbance at 260 nm DNA was removed with Turbo DNA-free (Ambion, Inc.). Reverse Transcription-PCR reaction was performed in a 15.0 µl final volume (kit number-DNA-PCR739288). Assessment of Polymerase Chain Reaction products (amplicons) were electrophoreses in 0.5% of agarose gel using 0.5 × TBE buffer ( 2.6 g of Tris base, 5 g of Tris boric acid and 2 ml of 0.5M EDTA and adjusted to pH 8.3 with the sodium hydroxide pellet) with 0.5 μl ethidum bromide. The mechanism of action of isolated novel compounds using Metacaspase3 to programme the death of test organism (Aspergillus flavus) between 0 and 180 minutes interval. It was observed that cell (via DNA) were completely destroyed at 180 minutes with all the isolated compounds. The purpose of this research work is to evaluate the programmed cell death (PCD) of Aspergillus flavus by triggered Cysteine-dependent Aspartate-directed proteases (meta-caspase3) lethality mechanism of novel compound isolated from ethyl acetate extract ofSpondias mombin.

Mechanical Property and Analysis of Asphalt Components Based on Molecular Dynamics Simulation

The asphalt-aggregate interface interaction plays a significant role in the overall performances of asphalt mixture. In order to analyze the chemical constitution of asphalt effects on the asphalt-aggregate interaction, the average structure C64H52S2 is selected to represent the asphalt, and the colloid, saturated phenol, and asphaltene are selected to represent the major constitutions in asphalt. The molecular models are established for the three compositions, respectively, and the Molecular Dynamics (MD) simulation was conducted for the three kinds of asphaltene-aggregate system at different presses. Comparing the E value of Young modulus of these three polymers, the maximum modulus value of asphaltene was 2.80 GPa, the modulus value of colloid was secondary, and the minimum modulus of saturated phenol was 0.52 GPa. This result corresponds to conventional understanding.

Attempted Determination of Endogenous Heparin in Blood

A method was developed for the micro-scale determination of heparin in blood plasma. After extraction with water-saturated phenol glycosaminoglycans were recovered from the aqueous phase by ion-exchange chromatography on DEAE-cellulose. The heparin content of the product was determined by a colorimetric antithrombin-activation assay. Recoveries of 40-60% of both radioactivity and biological activity were obtained when the method was applied to the reisolation of 3H-labeled heparin, added at a concentration of 20 ng (about 0,003 unit) per ml blood. In the absence of exogenous heparin no antithrombin-activating material could be detected. The amount of endogenous heparin, if any, in human blood was calculated to less than 2 ng (about 0.0003 unit) per ml.(Supported by grant No. 2309 from the Swedish Medical Research Council)