A Little TLC

Now that we have collected our different fractions, it is time to begin to consider what may be in the different fractions and, in particular, whether there is potentially a new and exciting molecule in one of them.

The first step towards doing this involves a process called thin layer chromatography (TLC).  Before carrying out TLC each fraction is redissolved in a small volume of 'Magic Brew' (a mixture of methanol and dichloromethane).

In TLC a capillary tube is used to add small amounts of each sample onto a TLC plate.  A TLC plate is a thin piece of plastic covered in silica.  The plate has a horizontal line near the bottom and you place a labelled pencil mark where you want each sample to go on the line.  The aim is to get a nice small circle of sample on each location.  To achieve this you 'spot' a small amount of sample, let it dry, and then add some more sample.  (If you put too much sample on at the start it spreads out too far and you will not get good results.)

Once we have spotted all our samples onto the plate, the plate gets lowered into a glass container with a small amount of solvent in the bottom.  The solvent used was 5% methanol in dichloromethane.  A lid is placed on the container so that fumes from the solvent are kept in the container.

The solvent begins to rise up the plate and as it does so it carries molecules with it. The surface of the TLC plate is quite polar. This results in polar molecules sticking to the plate while molecules of lower polarity continue to travel up the plate with the solvent. You end up with the most polar molecules at the bottom of the plate and the least polar molecules at the top.

The plate to the right shows my three P. microcladioides fractions. The 100% fraction contains a number of coloured compounds which have traveled varying distances up the plate.  There appear to be no compounds in the 30% and 75% fractions but this this only because none of them are coloured.  To see more compounds the plate is put under 2 different wavelengths of UV light.

The image to the left shows the plate when placed under a 350 nm UV light source while the image on the right is of the plate under a 254 nm UV light source. As you can see, new spots have shown up on the plate.  These are for molecules that contain chromophores. A chromophore is a part of a molecule that absorbs certain wavelengths of UV or visible light and reflects other wavelengths. (Chromophores often contain alternating single and double bonds.)

You may notice that the spots for the 30% fraction are all fairly low.  This makes sense as you will recall that the molecules in the 30% fraction are the most polar ones and it is these polar molecules that stick to the relatively polar surface of the plate and therefore don't travel very far.

Drying Down

Once we have obtained a number of solutions containing groups of molecules extracted from our red algae, it is time to dry down those solutions.  This involves evaporating off the solvent so that solid compounds remain.  To do this we use a rotary evaporator.

A rotary evaporator works by spinning the solution in a round-bottomed flask. The solution is spread around the inside surface of the flask and this increases the surface area from which the solvent can evaporate.

The rotary evaporator is attached to a pump which removes some of the air from inside the evaporator and therefore creates a partial vacuum. As the gas pressure above the surface of the solution is decreased, the solvent will evaporate off more quickly than it would under normal pressure.

The rotary evaporator to the left has a cold finger. This is the cone shaped object you can see inside the top part of the evaporator.  This is made very cold by filling it with dry ice (by removing the yellow lid at the top).  Solvent gas that has evaporated from the solution condenses into liquid when it comes into contact with the cold finger.  The liquid then drips into the flask at the bottom of the condenser.

As the solvent evaporates from the flask, the flask begins to get very cold and ice may even begin to form on the outside of it.  This is because when liquids evaporate they take heat from their surroundings.  If the flask gets too cold it will slow down the rate at which the solvent evaporates.  To get around this, the flask can be lowered into a water bath which prevents the flask from getting too cold.

On of the tricky things about drying down solutions is that they sometimes become unstable (particularly if they contain a mixture of water and acetone).  This can result in them bumping and spurting material up towards the condenser.  This would be a problem as we would be losing precious material and also contaminating the rotary evaporator.  For this reason, there is a glass trap above the round-bottomed flask. (It is between the red and green clips in the picture.)  Material that spurts gets caught in the trap and can be retrieved from there.


Once the solution has been dried down, you are left with a mixture of solid compounds attached to the the inside of the flask.  As the flask would be bulky to store (and we want to free it up so we can use it again), the next step is to transfer the compounds into a small glass vial. Before doing this we use a balance to find the mass of the vial so that we can later calculate the mass of material in the vial.


To transfer the material we need to redissolve it.  In the lab we use a solvent referred to as Magic Brew.  It is called this as it will dissolve most molecular compounds.  Magic Brew is a mixture of methanol and dichloromethane.  A small amount of Magic Brew is added to the flask in order to redissolve the solid compounds.  A glass pipette is then used to transfer the solution into the glass vial. The solution then needs to be dried down again on the rotary evaporator.  

The final step is to remove any traces of water that may not have evaporated on the rotary evaporator (as water has a higher enthalpy of vaporisation than most of the organic solvents we are using).  Any water needs to first be frozen and this is done by rolling the flask in liquid nitrogen.  The flask is then attached to a freeze dryer. As with the rotary evaporator, a freeze dryer creates a partial vacuum. This causes the frozen water to sublime (turn straight into  gas).  The water vapour is continually removed from the system by the freezer component of the freeze dryer.

Finally, the vial is reweighed and then stored in the fridge until we next want to work on the sample.

Pram Students @ Vic

One of the great things about being at Victoria University is catching up with past students of Paraparaumu College who are now doing Chemistry courses.  I have managed to track down seven of them.

Briana Hunt
Briana was a Year 13 Paraparaumu College student in 2013. She is currently in her first year of study towards a Biomedical Science degree, majoring in molecular pathology. Once she has finished her degree Briana intends to carry on further studies in the medical industry, pursue a career in sport medicine or complete her post-graduate degree. Briana is currently doing the First Year chemistry paper CHEM 113 Concepts of Chemistry. Students doing Biomedical Science need to take the First Year chemistry paper CHEM 114 Principles of Chemistry. CHEM 113, is a extra paper Briana wished to complete to gain further knowledge and skills to prepare her for CHEM 114. Briana is really enjoying Chemistry at Victoria, especially the practical experiments within the laboratories.

Georgina Bird
Georgina is another Biomedical Science student who is currently doing CHEM 113 in preparation for taking CHEM 114 in the second trimester. She was in Year 13 in 2005 and has since traveled and worked overseas. Georgina hopes that her qualification will eventually lead into a career in genetic counselling. She is enjoying being at Vic and getting back into study.


Lizzie Tafili, Rose McLellan & Bryony Ixer
Lizzie, Rose and Bryony are all currently taking CHEM 114 Principles of Chemistry and are in the same laboratory session (hence the combined photo).

Lizzie was in Year 13 last year and is now studying for a Bachelor of Biomedical Science majoring in medicinal chemistry and molecular pharmacology. As well as CHEM 114 she is taking a number of maths and biology papers, and next trimester intends taking CHEM 115 Structure and Spectroscopy. Lizzie is thoroughly enjoying CHEM 114. She likes the fact the paper contains some concepts learnt in physics and calculus as well as building on knowledge from NCEA Level Three chemistry. She describes it as a "really cool paper". Lizzie would like to one day do medical or chemistry related research so that she can "contribute and give back to the world".

Rose is another member of the class of 2013. She is now studying for a Bachelor of Science majoring in chemistry and biotechnology. Like Lizzie, Rose is currently taking CHEM 114 and I will take CHEM 115 next trimester.  She particularly enjoys the lab component of the course "because you get some fun practical experience that helps you understand all the new concepts you learn".  After graduating with her B.Sc. Rose would be interested in going into some form of research. She believes that her area of research will be determined by the components of chemistry that she is most compelled by in the later years of her degree. Teaching is also an option for Rose in the future because she "would love to have that interaction with people and share what I have learnt with them".  "Studying Chemistry at Victoria is really exciting and I know that it will only get better. The workload is hefty as with any university course but so far it has only inspired me to continue forward in Chemistry."

Bryony was in Year 13 at Paraparaumu College in 2012.  Last year she had a gap year and is now at Victoria University studying for a Bachelor of Science majoring in chemistry and economics.  At this stage Bryony is keeping her options open as to whether she eventually pursues a career in chemistry or economics (or maybe even manages to combine the two).  Bryony is currently doing the First Year Chemistry paper CHEM 114 Principles of Chemistry.  She is finding it challenging returning to study after having a gap year but is enjoying her time at Vic.

Lindsay Morris

Lindsay was in Year 13 in 2011 and is now in his final year of a Bachelor of Science majoring in chemistry and statistics. This year he is taking CHEM 301 - Organic Chemistry, CHEM 302 - Inorganic and Materials Chemistry, CHEM 303 - Physical and Process Chemistry and CHEM 305 - Organic Synthesis Laboratory. Next year Lindsay intends moving onto studying at the honors level with a particular focus on Organic Synthesis and Natural Products. If all goes well, his ultimate goal would be to either work on synthesizing molecules of particular biological interest, or discovering new compounds from marine sponges and alga.  "Studying Chemistry at Vic has been extremely challenging, however, it has been even more rewarding. I can't imagine what life would be like without having done Chemistry. I encourage students to take the challenge, and head to Vic to make the most of the best Science department and facilities New Zealand has to offer!"

Charlotte Page

Like Lindsay, Charlotte studied Chemistry at Paraparaumu College up until 2011. With an interest in Science, she has pursued a Bachelor of Biomedical Science specializing in molecular pathology and medicinal chemistry. She is currently in the third and last year of her degree, Charlotte has taken a variety of Chemistry courses, this year focusing on CHEM 301 - Organic Chemistry and CHEM 305 - Organic Synthesis Laboratory. She is considering options in postgraduate study, hopefully working towards a career in clinical pharmacology. "I have thoroughly enjoyed studying at Vic. I feel they offer a wide range of courses in Science, including Chemistry. Although challenging at times, there has always been assistance in a great community of scientists."

Briana, Georgina, Lizzie, Rose, Bryony, Lindsay and Charlotte are all having a great time studying Chemistry at Vic and would love to see more Paraparaumu College students join them in the future. If you would like to find out more information from any of them, send an email to me at mkb@pcol.school.nz and I will pass your message on to the relevant person.