One thing you will need to do is verify that the HPLC and fraction collector are "aligned". Some HPLC systems specify tubing lengths from the detector(s) and fraction collector, suggesting their software compensates for the delay time. This link (https://www.waters.com/webassets/cms/support/docs/71500012102re.pdf ) page 86, makes reference to a delaying time (I realize this is for a different fraction collector than yours, but the concept still holds).

I think you can choose the peaks for collection from a prior run.

As Jack suggested you have to allign the two instruments to identify the fraction which has your peak of interest. I have been doing it for quite some time and I always did it manually. And by manually I meant that I start the fraction collection just right after the injection when the UV trace starts. But a lot of HPLC’s software can also control this and will auto start the fraction collection. I never calculated the tubing length and would read through to see if I can improve my runs. By the way, are you doing reverse phase or normal phase?

Collect either manually OR better yet, with an automated fraction collector set up with the properly sized containers for collection and low volume tubing to reduce diffusion and losses to the collection tubes. Automate using simple timed-contact closures on your HPLC system's fraction collector (very easy to set-up and the fraction collector manuals often have simple examples to follow).

To determine the "start" and "stop" times for peak fraction collection, first calculate, then measure the volume from the detector's flow cell (when it is detected) to the fraction collector outlet tip. Once you have calculated it, measure the volume to be 100% certain and use this volume to calculate the needed delay volume (and therefore, the delay TIMES) needed to automatically collect peaks. *See the link below for tubing volumes. *Measure the volume by filling with water, then weigh the water!

This whole process is made much easier by first scaling the method up from an analytical method to a Preparative HPLC method on a Prep system (larger column = large sample volume). This allows you to inject one large volume into the column and column concentrated peaks each time. If this is not possible, then develop your analytical method to use a large volume column to maximize the sample loading (do a loading study first). OPTIMIZE the method for resolution so you have clean. Automate the collection to obtain the amount of material needed. Be sure to reanalyze fraction(s) for purity early in the process so you do not waste time collecting impure samples (which will invalidate your results).

Do not start the collection process until long after you have optimized the HPLC method. The method needs to be reproducible, follow good chromatography fundamentals and show excellent resolution to make the job of isolating and collecting peaks efficient and useful.

• "Do your HPLC Methods Meet Basic Good Chromatography Requirements? HPLC Training: RETAIN, SEPARATE and RESOLVE"; https://hplctips.blogspot.com/2019/12/do-your-hplc-methods-meet-basic.html

BTW: You did not indicate how much of each sample you need to collect, and for what purpose (normally, we collect fractions for ID using orthogonal methods so need a specific minimum amount of each for this purpose. Often several mgs are needed for each technique tried. ).

• CAPILLARY TUBING CONNECTION VOLUMES; https://www.hplctools.com/capvolume.htm
• HPLC COLUMN VOLUMES: https://www.hplctools.com/columnvolume.htm

PS: Put the "Waters 2424 ELS Detector" back on the shelf and do not use it.