Spinach Lab Report

Chromatography of Spinach Formal Discussion This lab involved the extraction of pigments from spinach leaves which were then analyzed using thin layer chromatography. The first step of this process was to grind up the leaves in order to extract the pigments. Hexanes facilitated this process and afterwards, the solution was dried over sodium sulfate to remove water. Next, the column was packed with a small piece of glass wool, followed by hexanes, and then packing sand. At this point, the column was packed half way. A mixture of 50:50 alumina and hexanes was then swirled into a flurry and poured into the column with an open stopcock. Once all the alumina settled uniformly, the liquid was drained until the meniscus almost touched the top of

Once completed, the results were fairly easy to read on the plate and the results are depicted in table 1. There were no drastic errors which would have dramatically affected the results, however, simple large-grained sand was used to pack the column instead of the packing sand, although this would not drastically affect the resulting TLC plate. There were no further difficulties conducting the experiment as written. Likely sources of error are most likely related to human error in reading the TLC bands or possibly the improper packing of the column so that the alumina band was not spread across the top layer. This could have impacted the pigments obtained which were then used on the TLC plate. The TLC was not too difficult to read, however, especially when looked at under a UV light. For the yellow column, there were faint bands all the way up to the top band which was located at 3cm, as you can see in figure 1. The green bands were much darker and easier to read. There were more than 5 bands preceding the darkest band which was located 2.3cm above the starting location.
Figure

Rf is equal to the distance traveled by the substance divided by the distance traveled by the solvent. Since the solvent used in the developing chamber was hexanes—a non-polar molecule— the more nonpolar the substance was, the stronger it would stick to the plate. This means that the more polar a pigment was, the higher it climbed on the TLC plate and would therefore have a larger Rf. There are 3 major classes of pigments present in spinach: carotenes, xanthophylls, and chlorophylls. Since the solvent is nonpolar, we would expect carotene to have the lowest Rf, then xanthophylls, and chlorophylls would have the highest. As discussed before, this is because chlorophylls are the most polar of the three and carotenes are the least polar of the three. Based on my results, I would say that the packing and running of the column was effective in separating the pigments as there were quite different Rf values for the original than for the other two colors and the band for the yellow was nothing like the bands for the green. There were no limitations in procedure which negatively affected the end results, so there is nothing I would change to it going