Chrom

It’s a method to determine how well different columns work and it describes you the idea of theoretical plates (N)

It’s not a real plate , it represents one equilibrium step between the mobile and stationary phase the more plates you have the better separation there is

H=L/N H = HETP -height equivalent of theoretical plate L= length of column N= number of plates

It means more plates are better separation so more narrow peaks

It means less plates are so poor separation so broader peaks

Eddy diffusion Longitudinal diffusion Mass transfer

Molecules take different paths through packed columns which is caused by irregular particle packing and you have an independent flow rate

Molecules diffuse from a high to low concentration, this happens while analytes sit in the column and it’s worse at low flow rates

Is the time needed for molecules to move from the mobile to the stationary phase and back again and it’s worse with the large stationary phase particles in high flow rates

Detector measure signal intensity vs time producing peaks

Gaussian - bell shaped Symmetrical around the central axis

Peaks broaden with longer retention as a longer the solute is retained on the compound the greater variability of solute column interactions.

Is the time taken for the solute to be elected from the column and it’s a characteristic of a compound when analysed under the same conditions

Difference between the middle of the peak on the start of the graph/difference between another peak and the start of the graph This is very useful if one of the peaks is a known standard compound

It’s to measure how well two peaks are separated

The larger the retention time means the smaller peak width so better resolution

A resolution value of 1.0 shows acceptable separation

It’s where you have no overlap at the baseline and has a value of 1.5

Larger difference in retention time Narrower peak

The peak area integral is related to the concentration of the compound, but not necessarily same for all compound

Peak area is proportional to the amount of compound, different compounds may give different responses and this is why calibration is needed

When the mobile phase composition stays constant. This is best for simple mixtures, they have an easy set up and maintain, we have consistent conditions or lead to more reproduce results and have simpler instrumentation

This is when the mobile phase composition changes over time. this is best for complex samples as you have better separation of compounds with different priorities or chemical properties, sharper peaks and faster analysis and washes strongly retained compounds of the column

UV light passes through the sample Compounds absorbed light Detector measure measures absorbed light vs transmitted light Most common detector is HPLC. The more light absorb the more molecules present.