chromatography Flashcards
what is chromatography
it is a method by which a mixture is separated by distributing its components between two phases
what happens to the stationary phase
it remains fixed in place; it often acts as a constraint and slows down components in the mixture
what happens in the mobile phase
it carries components of the mixture through the medium, components that like this phase move through the medium quicker
what is retention time?
it is the time that passes between a sample injection and the maximum response for a solutes peak
how is retention time represented
TR= ts+tm
what is ts
the time spent in stationary phase
what is tm
the time spent in the mobile phase, it is also the retention time of an unretained compound that spends no time in the stationary phase
what is tR
the retention time
what is the retention factor?
it is k= (tR-tm)/tm
what is another way to think about k?
time in stationary/time in mobile
what is true of tm
tm is the fastest, it does not stop in the stationary phase at all
the amount of retention time increases as
the amount of stationary time increases
a larger retention time means what?
it means the substance favors the stationary phase
do all the substances spend the same amount of time in the mobile phase
yes, that is why we use tm because it is the same across all of the samples
what is the separation due to? why do all the compounds spend the same amount of time in the mobile phase
As all compounds have the same length to travel before they exit the column, they all spend the same amount of time in the mobile phase. Thus, separation is due to the varying amount of time spent in the stationary phase, or how long compounds adhere to the column coating
what is a separation factor
Selectivity is a relative measure of the retention of two solutes
what is the formula for the separation factor?
it is the larger partition coefficient divided by the smaller partition coefficient
what is relative retention?
it is given to us by the separation factor
what does the selectivity factor compare?
the stationary phase of peak 2 over the stationary phase of peak 1 where the lowest factor is 1.00 which is when the two peaks overlap and have the same retention time
what is the formula for the resolution
the change in retention times divided by the average peak width, it is arranged so Rs is always greater than 0
what does the resolution tell us
how well separated something is
do we want a larger resolution or smaller resolution
we want a resolution that is larger where 1 is the minimum and 1.50 is the best for quantitative analysis
How is the concentration and response on the graph related
the higher the concentration, the higher the peak, this changes the area under the curve
what does the retention time tell us
it tells us what the compound is
what is the y axis on a chromatograph
instrument response
in chromatography what is the x axis on the graph
it is the retention time
why do we want narrow peaks?
when the peaks are narrow and high, its easier to separate because its more concentrated. As the analyte travels down the column, the peak gers smaller as the concentration becomes more diluted on the edges of the band
what makes a better separation factor
if there is a greater difference in times, the peaks will be further separated from one another and will have a better separation factor
where does the separation and stationary phase occur in an LC?
in the column
what’s the main difference between liquid chromatography and gas chromatography?
liquid chromatography has a liquid mobile phase and a solid stationary phase
how do LC and HPLC compare
LC involved low pressure and its elution is driven by gravity
it typically has low resolution (worse separation) and large particles
What is different about HPLC and UHPLC
they involve higher pressue and smaller particles
why are smaller particles better in LC?
they allow for better separation
What is the normal mode of separation
it is where the stationary phase is polar and the mobile phase is nonpolar
in a normal mode of separation, what types of analytes will elute first
in a normal mode of separation, the mobile phase is nonpolar so they nonpolar analytes will elute first
In a normal mode of separation, what types of molecules will be retained
the stationary phase is polar so polar molecules will be retained
In a reversed mode of separation, what analytes elute first
the polar analytes will favor the mobile phase
describe the stationary and mobile phases of separation in a reversed mode of separation
the stationary phase is nonpolar
the mobile phase is polar with water and polar organic modifiers like methanol and acetonitrile
what is the mobile phase composed of in a normal mode of separation
it is composed of hexane and polar modifiers: ethanol and acetone
in separation, if the mobile and stationary phases become more similar, what happens to the retention times
the retention times decrease
in separation, if the mobile and stationary phases become less similar, what happens to the retention times
the retention times increase
in a gas chromatography, what is the mobile phase?
they are gaseous
what drives the analyte separation in gas chromatography
higher temperatures force more gases into the mobile gas phase
