my final year project....

chromatography studies on separation of laccase is the title of my final year project which has been given by my supervisor. supposedly, i should b d one who proposed the title but he proposed it for me..h7uhuhuhu..it was barely started at the end of 2nd sem of my 2nd year..at dat time i was so blur..i tried to seek information as much as i can..but everything was so not to my level of understanding...huhu..well when i go trough several series of experiment..now i know the meaning of the tittle..

producing the laccase is not the main objective of my project..but the priority is given on how the laccase is purified by using column chromatography..anyway Laccases are copper-containing oxidase enzymes that are found in many plants, fungi, and microorganisms. The copper is bound in several sites; Type 1, Type 2, and/or Type 3. The ensemble of types 2 and 3 copper is called a trinuclear cluster . Laccases act on phenols and similar molecules, performing a one-electron oxidations, which remain poorly defined. It is proposed that laccases play a role in the formation of lignin by promoting the oxidative coupling of lignols, a family of naturally occuring phenols. Laccases can be polymeric, and the enzymatically active form can be a dimer or trimer.

The classical preparative chromatography column is a glass tube with a diameter from 5 to 50 mm and a height of 50 cm to 1 m with a tap at the bottom. 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. 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...

here are several pictures that i have taken during laboratory work that i've done recently..