Exercise 5 results


Please don’t. To enter this text as an answer results in zero points and a bad reputation of your person in your teacher’s mind.

In another question one of you copied text and added the reference. This is the correct behaviour

Generally you get zero points for answering a yes/no question incorrectly. In some cases where a correct and understood reasoning was added (that was not copied from somewhere) I awarded 0.5 points for that.

Overall performance was 74% which is in line with the previous tests. What I can see is that some of you follow a minimalistic approach with not answering questions that require thinking or working. Some submit correct, most probably copied texts that are correct but lack relationship to the questions. This is dangerous and will result in a FAIL decision.

I have to point out that these cases are a minority. In most of the cases I am delighted to see how skilled you are in gathering information and mostly digesting it to a meaningful conclusion relating to the question 🙂

Fig. 1: Test performance in 50310-01 in 2021. Line shows average and dots individual results. Values in Exercise 2 and 5 are not normally distributed. This suggests the presence of two distinct populations.

Comments to the questions

Questionperfect answerComment
What is the advantage of the high pressure in HPLC?noneThe pressure itself is not an advantage. Please read the questions carefully and answer them. Congratulations go to Florian and Jara for being the only ones to answer this correctly.
Will separation power always increase if you reduce the flow rate?NoThat was to check whether you got the point of the vanDeemter curve. The results were a bit mixed, I have to work on making points..
Selectivity is independent of resolution, true or false?YesEverybody was wrong here except Robin Jane, to whom goes another congratulation. It was sooo easy: Go to slide ‘Chromatographic parameters, Separation and Resolution’, you see that Selectivity is = k2/k1 (the retention factors that have nothing to do with peak shape.
Download Module 5- paper A and B examples. Compare and comment the differences in the HPLC part.A (Mason, 1995): precipitation with ZnSO4/MeOH
HPLC 4.6×250 mm phoshoric acid/ACN, 
16min runtime
UV 355nm
Advantage: easy sample prep, aqueous sample optimal for good peak shape. disadvantage: slow, high solvent comsumption

B (Ni, 2015):
liquid-liquid-extraction with NaOH/ethyl acetate
UPLC 2 x 50mm, TFA/ACN, 
2min runtime
UV 310nm
Advantage: rapid, low solvent consumption
Disadvantage: LLE is more work than precipitation

This question gave totally 4 points because of the higher effort, 2 for the comparison with mention of HPLC/UPLC, run time difference etc., and 2 for a judgment with advantages and disadvantages that was not barely copied from the papers.

Please notice that 16 min runtime and 1.5ml/min flow you consume 24 ml/sample and with UPLC in 2 min with 0.3ml/min 0.6ml per sample – this is a factor of 40 improvement in environmental protection ! Not many of you mentioned this point.
You use a gradient to separate two compounds, but your separation is incomplete.
Might it help to make the gradient steeper?
NoSteep gradients are never good for peak shape (but might be for speed). Was mentioned in the lecture.
Might it help to start at more aqueous conditions and make the run longer?YesMore aqueous starting conditions in Reversed Phase LC concentrate the sample in a narrow band at the head of the column. This is advantageous for narrow peaks. Shallower gradients suffer less from incomplete solvent mixing.
Might it help to take a longer column?YesLonger column = more interactions = better separation. All of you got that. One point I was able to make..

Now I wish you an enjoyable rest of the weekend and hope that you join again on Tuesday. It will be on elemental composition determination by mass spectroscopy and I might need both lessons for all the theory and examples, therefore the test – where you have the role of the analytical contributor in a metabolomics study – might be in the first hour of the lesson on April, 6. Let’s see!

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