DEVELOPMENT OF THE TECHNOLOGY FOR THE SEPARATION OF THE COMPONENTS FROM GREEN TEA USING THE PREPARATIVE LIQUID CHROMATOGRAPHY BY DESIGN OF PERIODICAL PROCESS

Leaves of the green tea plant camellis sinensis contain up to 36 % polyphenols. Epigallocatechin gallate (EGCG) is the most interesting polyphenols because it exhibits a strong antioxidant effect. Furthermore, it has been demonstrated that EGCG has an antimutagenic and an anticancer effects, an antibacterial effect and a beneficial effect on cholesterol level in blood. Therefore, is needed to isolate EGCG in a pure form in high yield by simple and commercial process. This method for isolation of EGCG contents the following steps: a) selection of macroporous polar resin for the preparative liquid chromatography; b) selection of polar elution solvent; c) determination the temperature and pressure for adsorbtion and desorbtion of the polyphenols of the green tea. Therefore, it was searched the interconnection between yield of EGCG and the range of temperature and amount of eluent. Finally, the process for the separation of Epigallocatechin gallate from green tea extract was developed in the laboratory and EGCG was isolated with the concentration 92% by the yield 43%. Accordingly, the technology of EGCG production based on the preparative liquid chromatography was launched and introduced on the market.

Advances Achieved by Ionic-Liquid-Based Materials as Alternative Supports and Purification Platforms for Proteins and Enzymes

Ionic liquids (ILs) have been applied in several fields in which enzymes and proteins play a noteworthy role, for instance in biorefinery, biotechnology, and pharmaceutical sciences, among others. Despite their use as solvents and co-solvents, their combination with materials for protein- and enzyme-based applications has raised significant attention in the past few years. Among them, significant advances were brought by supported ionic liquids (SILs), in which ILs are introduced to modify the surface and properties of materials, e.g., as ligands when covalently bond or when physiosorbed. SILs have been mainly investigated as alternative supports for enzymes in biocatalysis and as new supports in preparative liquid chromatography for the purification of high-value proteins and enzymes. In this manuscript, we provide an overview on the most relevant advances by using SILs as supports for enzymes and as purification platforms for a variety of proteins and enzymes. The interaction mechanisms occurring between proteins and SILs/ILs are highlighted, allowing the design of efficient processes involving SILs. The work developed is discussed in light of the respective development phase and innovation level of the applied technologies. Advantages and disadvantages are identified, as well as the missing links to pave their use in relevant applications.

Downstream Processing of Therapeutic Peptides by Means of Preparative Liquid Chromatography

The market of biomolecules with therapeutic scopes, including peptides, is continuously expanding. The interest towards this class of pharmaceuticals is stimulated by the broad range of bioactivities that peptides can trigger in the human body. The main production methods to obtain peptides are enzymatic hydrolysis, microbial fermentation, recombinant approach and, especially, chemical synthesis. None of these methods, however, produce exclusively the target product. Other species represent impurities that, for safety and pharmaceutical quality reasons, must be removed. The remarkable production volumes of peptide mixtures have generated a strong interest towards the purification procedures, particularly due to their relevant impact on the manufacturing costs. The purification method of choice is mainly preparative liquid chromatography, because of its flexibility, which allows one to choose case-by-case the experimental conditions that most suitably fit that particular purification problem. Different modes of chromatography that can cover almost every separation case are reviewed in this article. Additionally, an outlook to a very recent continuous chromatographic process (namely Multicolumn Countercurrent Solvent Gradient Purification, MCSGP) and future perspectives regarding purification strategies will be considered at the end of this review.

Controlled Production of Zearalenone-Glucopyranoside Standards with Cunninghamella Strains Using Sulphate-Depleted Media

In recent years, conjugated mycotoxins have gained increasing interest in food safety, as their hydrolysis in human and animal intestines leads to an increase in toxicity. For the production of zearalenone (ZEN) glycosides reference standards, we applied Cunninghamellaelegans and Cunninghamella echinulata fungal strains. A sulphate-depleted medium was designed for the preferred production of ZEN glycosides. Both Cunninghamella strains were able to produce zearalenone-14-β-D-glucopyranoside (Z14G), zearalenone-16-β-D-glucopyranoside (Z16G) and zearalenone-14-sulphate (Z14S). In a rich medium, Cunninghamellaelegans preferably produced Z14S, while Cunninghamellaechinulata preferably produced Z14G. In the sulphate-depleted medium a dramatic change was observed for Cunninghamellaelegans, showing preferred production of Z14G and Z16G. From 2 mg of ZEN in sulphate-depleted medium, 1.94 mg of Z14G and 0.45 mg of Z16G were produced. Following preparative Liquid Chromatography-Mass Spectrometry (LC-MS) purification, both fractions were submitted to 1H and 13C NMR and High-Resolution Mass Spectrometry (HRMS). These analyses confirmed that the purified fractions were indeed Z14G and Z16G. In conclusion, the presented research shows that a single Cunninghamella strain can be an effective and efficient tool for the controlled biotransformation of ZEN glycosides and other ZEN metabolites. Additionally, the biotransformation method was extended to zearalanone, β-zearalenol and other mycotoxins.

A REVIEW ON FLASH CHROMATOGRAPHY AND ITS PHARMACEUTICAL APPLICATIONS

Earlier column chromatography was used for preparative purposes as well as for reaction control in organic synthesis. Column chromatography is an extremely tedious stage in any laboratory and can quickly become a point of congestion for any process lab. Flash chromatography is a technique developed as a modification of preparative column chromatography. This is an air pressure driven technique comprising of medium and short column chromatography, optimised for rapid separation of organic compounds. Modern flash chromatographic system consists of pre- packed plastic cartridges where in the solvent is pumped through the cartridge. By employing high pressure gas, the efficiency and speed of classical column chromatography can be increased. Flash chromatography is an easy & quick approach that is economical to preparative liquid chromatography. The content mentioned in this article mainly focuses on the various components, general principles, procedures and applications of flash chromatography.