COLUMN CHROMATOGRAPHY

A chemist in the 1950s using column chromatography. The Erlenmeyer receptacles are on the floor.

Column chromatography in chemistry is a method used to purify individual chemical compounds from mixtures of compounds. It is often used for preparative applications on scales from micrograms up to kilograms. The main advantage of column chromatography is the relatively low cost and disposability of the stationary phase used in the process. The latter prevents cross-contamination and stationary phase degradation due to recycling.

The classical preparative chromatography column, is a glass tube with a diameter from 5 mm to 50 mm and a height of 5 cm to 1 m with a tap and some kind of a filter (a glass frit or glass wool plug – to prevent the loss of the stationary phase) at the bottom. Two methods are generally used to prepare a column: the dry method, and the wet method.

• For the dry method, the column is first filled with dry stationary phase powder, followed by the addition of mobile phase, which is flushed through the column until it is completely wet, and from this point is never allowed to run dry.
• For the wet method, a slurry is prepared of the eluent with the stationary phase powder and then carefully poured into the column. Care must be taken to avoid air bubbles. A solution of the organic material is pipetted on top of the stationary phase. This layer is usually topped with a small layer of sand or with cotton or glass wool to protect the shape of the organic layer from the velocity of newly added eluent. Eluent is slowly passed through the column to advance the organic material. Often a spherical eluent reservoir or an eluent-filled and stoppered separating funnel is put on top of the column.

The individual components are retained by the stationary phase differently and separate from each other while they are running at different speeds through the column with the eluent. At the end of the column they elute one at a time. During the entire chromatography process the eluent is collected in a series of fractions. Fractions can be collected automatically by means of fraction collectors. The productivity of chromatography can be increased by running several columns at a time. In this case multi stream collectors are used. The composition of the eluent flow can be monitored and each fraction is analyzed for dissolved compounds, e.g. by analytical chromatography, UV absorption, or fluorescence. Colored compounds (or fluorescent compounds with the aid of an UV lamp) can be seen through the glass wall as moving bands.

Overview

Stationary phase

The stationary phase or adsorbent in column chromatography is a solid. The most common stationary phase for column chromatography is silica gel, followed by alumina. Cellulosepowder has often been used in the past. Also possible are ion exchange chromatography, reversed-phase chromatography(RP), affinity chromatography or expanded bed adsorption(EBA). The stationary phases are usually finely ground powders or gels and/or are microporous for an increased surface, though in EBA a fluidized bed is used. There is an important ratio between the stationary phase weight and the dry weight of the analyte mixture that can be applied onto the column. For silica column chromatography, this ratio lies within 20:1 to 100:1, depending on how close to each other the analyte components are being eluted.

Mobile phase (eluent)

The mobile phase or eluent is either a pure solvent or a mixture of different solvents. It is chosen so that the retention factor value of the compound of interest is roughly around 0.2 - 0.3 in order to minimize the time and the amount of eluent to run the chromatography. The eluent has also been chosen so that the different compounds can be separated effectively. The eluent is optimized in small scale pretests, often using thin layer chromatography (TLC) with the same stationary phase.

There is an optimum flow rate for each particular separation. A faster flow rate of the eluent minimizes the time required to run a column and thereby minimizes diffusion, resulting in a better separation. However, the maximum flow rate is limited because a finite time is required for analyte to equilibrate between stationary phase and mobile phase, see Van Deemter's equation. A simple laboratory column runs by gravity flow. The flow rate of such a column can be increased by extending the fresh eluent filled column above the top of the stationary phase or decreased by the tap controls. Faster flow rates can be achieved by using a pump or by using compressed gas (e.g. air,nitrogen, or argon) to push the solvent through the column (flash column chromatography).

The particle size of the stationary phase is generally finer in flash column chromatography than in gravity column chromatography. For example, one of the most widely used silica gel grades in the former technique is mesh 230 – 400 (40 – 63 µm), while the latter technique typically requires mesh 70 – 230 (63 – 200 µm) silica gel.

A spreadsheet that assists in the successful development of flash columns has been developed. The spreadsheet estimates the retention volume and band volume of analytes, the fraction numbers expected to contain each analyte, and the resolution between adjacent peaks. This information allows users to select optimal parameters for preparative-scale separations before the flash column itself is attempted.

An automated ion chromatography system.

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Typical set up for manual column chromatography

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Boron vapour trail leads to heterofullerenes

The simple route to borafullerenes could open up an interesting new avenue of heterofullerene research © Wiley-VCH

http://www.rsc.org/chemistryworld/2012/12/boron-fullerene-heterofullerene-atom-exchange

A team of scientists has developed a simple way to synthesise heterofullerenes – fullerenes with atoms other than carbon in their structure – by exposing fullerenes to boron vapour during their growth. They found that atom exchange with a carbon takes place to form a derivative known as borafullerene. The team believes the process can be easily scaled up and applied to other all-carbon analogues including nanotubes or graphene.

read all at

http://www.rsc.org/chemistryworld/2012/12/boron-fullerene-heterofullerene-atom-exchange

Nano-Technoloogy Makes Medicine Greener

The ultra small nanoreactors have walls made of lipids. During their fusion events volumes of one billionth of a billionth of a liter were transferred between nanoreactors allowing their cargos to mix and react chemically. We typically carried out a million of individual chemical reactions per cm2 in not more than a few minutes. (Credit: Image courtesy of University of Copenhagen)http://www.sciencedaily.com/releases/2011/11/111103132357.htm

 Researchers at the University of Copenhagen are behind the development of a new method that will make it possible to develop drugs faster and greener. Their work promises cheaper medicine for consumers.

Over the last 5 years the Bionano Group at the Nano-Science Center and the Department of Neuroscience and Pharmacology at the University of Copenhagen has been working hard to characterise and test how molecules react, combine together and form larger molecules, which can be used in the development of new medicine.http://www.sciencedaily.com/releases/2011/11/111103132357.htm

Computational chemistry draws for the first time the “interactive cartographic map" of enzymes during chemical reactions

Enzyme map

Researchers in Spain have used computational chemistry to plot for the first time a "cartographic map" of enzyme behavior during the catalytic process. Features include the moment when a given enzyme is at the point of maximum energy on the way from taking reactants to final product, which all happens within a femtosecond. The study conducted by researchers at the Universitat Jaume I and the University of Valencia was published in Nature Chemistry. "We can [now] make a quantitative estimate of the flexibility of the protein, how much it deforms itself, how much energy you need to deform that protein to generate the reaction that you want," explains UJI's Vicent Moliner.
Computational chemistry draws for the first time the "interactive cartographic map" of enzymes during chemical reactions
http://www.uji.es/UK/ocit/noticies/detall&id_a=33312503

Enzymes Make the World Go 'Round

We often talk about reactions and the molecules that change in those reactions. Those changes don't happen on their own. If you leave a blob of protein in a Petri dish, will it just break down to the amino acids? No. What will do it? Enzymes! Enzymes are the biological substances (proteins) that act as catalysts and help complex reactions occur everywhere in life.

Assembly Line Robots

You all know about cars and the assembly lines where they are made. There are giant robots helping people do specific tasks. Some lift the whole cars, some lift doors, and some just put bolts on. Enzymes are like those giant robots. They grab one or two pieces, do something to them, and then release them. Once their job is done, they move to the next piece and do the same thing again. They are little protein robots inside your cells.

Enzymes complete very, very specific jobs and do nothing else. The robot that was designed to move a car door can't put brakes on the car. The specialized robot arms just can't do the job. Enzymes are the same. They can only work with specific molecules and only do specific tasks. For example, you might have a protein in a cell. Even with hundreds of amino acids in the chain, the overall shape changes if one amino acid is different. That tiny shape change could stop the enzyme from doing its job. Some herbicides are used to block enzyme activity. Plants have adapted by changing one or two amino acids in the enzymes. They can continue to work with the correct proteins and there is no bonding to the herbicides. In the same way that there are specialized robots for different types of cars, there are enzymes for neural cells, intestinal cells, and your saliva.

There are four steps in the process of an enzyme at work:
1. An enzyme and a substrate are in the same area. The substrate is the biological molecule that the enzyme will work on.
2. The enzyme grabs on to the substrate at a special area called the active site. Enzymes are very, very specific and don't just grab on to any molecule. The active site is a specially shaped area of the enzyme that fits around the substrate. The active site is like the grasping handle of the robot on the assembly line. It can only pick up one part.
3. A process called catalysis happens. Catalysis is when the substrate is changed. It could be broken down or combined with another molecule to make something new.
4. The enzyme lets go. This is a big deal. When the enzyme lets go, it returns to normal, ready to work on another molecule of substrate. The first molecule is no longer the same. It is now called the product.

Enzymes

At any given moment, all of the work being done inside any cell is being done by enzymes. If you understand enzymes, you understand cells. A bacterium like E. coli has about 1,000 different types of enzymes floating around in the cytoplasm at any given time.
Enzymes have extremely interesting properties that make them little chemical-reaction machines. The purpose of an enzyme in a cell is to allow the cell to carry out chemical reactions very quickly. These reactions allow the cell to build things or take things apart as needed. This is how a cell grows and reproduces. At the most basic level, a cell is really a little bag full of chemical reactions that are made possible by enzymes!
Enzymes are made from amino acids, and they are proteins. When an enzyme is formed, it is made by stringing together between 100 and 1,000 amino acids in a very specific and unique order. The chain of amino acids then folds into a unique shape. That shape allows the enzyme to carry out specific chemical reactions -- an enzyme acts as a very efficient catalyst for a specific chemical reaction. The enzyme speeds that reaction up tremendously.
For example, the sugar maltose is made from two glucose molecules bonded together. The enzyme maltase is shaped in such a way that it can break the bond and free the two glucose pieces. The only thing maltase can do is break maltose molecules, but it can do that very rapidly and efficiently. Other types of enzymes can put atoms and molecules together. Breaking molecules apart and putting molecules together is what enzymes do, and there is a specific enzyme for each chemical reaction needed to make the cell work properly.

Maltose is made of two glucose molecules bonded together (1). The maltase enzyme is a protein that is perfectly shaped to accept a maltose molecule and break the bond (2). The two glucose molecules are released (3). A single maltase enzyme can break in excess of 1,000 maltose bonds per second, and will only accept maltose molecules.

You can see in the diagram above the basic action of an enzyme. A maltose molecule floats near and is captured at a specific site on the maltase enzyme. The active site on the enzyme breaks the bond, and then the two glucose molecules float away.
You may have heard of people who are lactose intolerant, or you may suffer from this problem yourself. The problem arises because the sugar in milk -- lactose -- does not get broken into its glucose components. Therefore, it cannot be digested. The intestinal cells of lactose-intolerant people do not produce lactase, the enzyme needed to break down lactose. This problem shows how the lack of just one enzyme in the human body can lead to problems. A person who is lactose intolerant can swallow a drop of lactase prior to drinking milk and the problem is solved. Many enzyme deficiencies are not nearly so easy to fix.
Inside a bacterium there are about 1,000 types of enzymes (lactase being one of them). All of the enzymes float freely in the cytoplasm waiting for the chemical they recognize to float by. There are hundreds or millions of copies of each different type of enzyme, depending on how important a reaction is to a cell and how often the reaction is needed. These enzymes do everything from breaking glucose down for energy to building cell walls, constructing new enzymes and allowing the cell to reproduce. Enzymes do all of the work inside cells.

Natural Products from St. John's Wort

Natural Products from St. John's Wort

Enatiomers (−)-hyperione A and (−)-hyperione B can be synthesised from a common precursor in ten steps
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http://www.chemistryviews.org/details/ezine/1238795/Natural_Products_from_St__Johns_Wort.html

Improved R&D Efficiency Through Speedier Method Development (1)

Improved R&D Efficiency Through Speedier Method Development (1)

Various approaches are available for improving the efficiency of analytical...

http://www.pharmaceutical-business-review.com/suppliers/shimadzu-analytical-instrumentation/whitepapers/improved-rd-efficiency-through-speedier-method-development-1

Bruce Roth Awarded 2013 Perkin Medal

Roth

Credit: Genentech

Bruce Roth Awarded 2013 Perkin Medal

Honors: Chemist was the first to synthesize the cholesterol-lowering drug atorvastatin, also known as Lipitor
The Society of Chemical Industry (SCI) has selected Bruce D. Roth, vice president of discovery chemistry at Genentech, as the winner of the 2013 Perkin Medal. The annual award is recognized as the highest honor given for outstanding work in applied chemistry in the U.S.
http://cen.acs.org/articles/91/web/2013/06/Bruce-Roth-Awarded-2013-Perkin.html

A Molecule Of Many Colors-With rigid wings and a flexible core, a new compound can switch between two shapes and glow one of three colors.

 

Flexible And Fluorescent

A molecule combining rigid anthraceneimide wings and a flexible cyclooctatetraene core switches between a flat and a bent V shape. The R groups are either hydrogens or n-butyl groups.

Credit: J. Am. Chem. Soc.

A Molecule Of Many Colors

Organic Chemistry: With rigid wings and a flexible core, a new compound can switch between two shapes and glow one of three colors.

A new, flexible, multi-ring organic compound fluoresces red, green, or blue depending on its environment (J. Am. Chem. Soc. 2013, DOI: 10.1021/ja404198h). The molecule’s combination of rigid wings and a flexible center could serve as a general design strategy for molecular sensors, the researchers say.

The molecule, developed by a team of researchers, including Shohei Saito, Stephan Irle, and Shigehiro Yamaguchi of Nagoya University in Japan, has two rigid anthraceneimide wings on opposite sides of a floppy cyclooctatetraene core

read all at
http://cen.acs.org/articles/91/web/2013/06/Molecule-Colors.html

PROCESS RESEARCH OVERVIEW

Find 3 presentations in the link 

http://www.anthonymelvincrasto.yolasite.com/process-overview.php

Blind trials of Computer-Assisted Structure Elucidation software

Blind trials of Computer-Assisted Structure Elucidation software

Arvin Moser, Mikhail E Elyashberg, Antony J Williams, Kirill A Blinov, Joseph C DiMartinoJournal of Cheminformatics 2012, 4:5 (9 February 2012)READ AT  http://www.jcheminf.com/content/4/1/5/abstract

http://www.jcheminf.com/content/4/1/5/abstract