HP (Agilent) 1050 - HPLC

So, I scored an eBay bargain: an old HP 1050 HPLC setup, for 99 pence. I did end up having to pay a courier £114 to get it to me, but even at £114.99 it's a bargain.


The biggest issue is that I don't actually have a column yet. I'll need to get one before I can do any chromatography, obviously. I'm thinking I'll buy a second-hand C18 (octadecylsilane/ODS) column but honestly I don't really know what I'm doing.

In the meantime I have been testing all of the modules individually and together as a system. I've been surprised by how little work it took to get it going.

I've taken all of the modules to bits, cleaned them out, and fixed various minor problems. Some of the modules had clearly been cannibalised for spare parts (and I took even more off of them) but with a small amount of effort I now have a full working set:

• 1x quaternary pump
  
• 1x variable wavelength UV absorbance detector
  
• 1x autosampler
  
• 1x solvent cabinet w/ column oven
  

The second quaternary pump and second autosampler have been broken down into the useful spare parts (which I'll keep and/or sell on eBay - but I'm going to throw out the bulky cases to save space).

The second variable wavelength UV detector complains with a 'low intensity' message, so I'm guessing that the deuterium lamp is near the end of its life, or perhaps there's an alignment issue in the optical block. Superficially it does appear to work though, but I haven't spent any time looking at it since the other one works perfectly.

The online vacuum degasser is not currently working. It runs for a bit but shuts off with an error light - apparently this means that the pressure didn't reach the threshold before a timeout, and it gave up. However the vacuum pump definitely works, so I think there is a small leak somewhere.

Quote: Originally posted by stamasd

I won't do it again.

My biggest problem so far has been water quality. Turns out distilled water really isn't good enough - at least, for gradient runs.

With gradient runs (starting with a weak eluent and gradually increasing the eluent strength) contaminants in the weak eluent (which for reverse phase is water) are retained strongly by the column and then elute later with the stronger eluent, creating ghost peaks.


Here's a blank run (i.e. no sample was injected) using distilled water:

Scale: 1 horizontal division = 5 mins, 1 vertical division = 40 mAU
Detection: UV absorbance at 254nm
Column: C18/ODS 50mmx4.6mm
Eluent: Hold at 20:80 (vol) MeOH:dH₂O for 2 minutes, then linear gradient to 100% MeOH over 10 minutes, then hold for 3 minutes




Isocratic runs (i.e. runs with constant eluent composition) are much less sensitive to water quality issues because strongly retained contaminants are, well, strongly retained (may never elute at all or elute much later than all sample components). Of course, the accumulation of contaminants will gradually change the chromatographic properties of the column, but washing with strong eluent periodically will sort that.

Here's an isocratic blank run using distilled water:

Scale: 1 horizontal division = 5 mins, 1 vertical division = 40 mAU
Detection: UV absorbance at 254nm
Column: C18/ODS 50mmx4.6mm
Eluent: Isocratic, 66:34 (vol) MeOH:dH₂O

Quote: Originally posted by DavidJR

Some updates: So: I have constructed a simple data acquisition device with an ADS1232 ADC (24 bits, however since I'm feeding a single-ended signal into a differential input, I only use half of the dynamic range). It can record to an SD card or directly to the PC in real time. It is also connected to the HPSystem remote bus to automatically start acquisition at the correct time etc. [Edited on 17-7-2018 by DavidJR]

Quote: Originally posted by wg48temp

I recognised that chip. 23.5 usable bits for about £5. Did you use the evaluation board for it or did you design your own? How do you get the info in to the PC. Are you going to write up the details? Please do. Great project by the way. It must have been a huge task getting it going and understanding it. Any particular plans on what you will do with it?

Main ongoing issue was enormous ghost peaks from mobile phase impurities. My own attempts at purifying water and methanol to a sufficient degree were not all that successful. To deal with this I have:

1. Bought an old Milli-Q Synthesis A10 ultrapure water machine, along with a set of new old-stock consumables for it (they are very pricey).
   
   This takes in purified water (I'm feeding mine from a reservoir filled manually from an el-cheapo ebay reverse osmosis / mixed-bed DI system) and puts out Type I ultrapure water (resistivity = 18.2 megaohm centimetres at 25C, total organic carbon around 7 ppb).
   
   
   
2. Bought HPLC grade solvents. These have been purified to remove UV-absorbing trace organic contaminants.
   
   I purchased HPLC grade methanol and HPLC Sp. grade acetonitrile from a local company, Rathburn Chemicals.
   

Together, these result in a dramatically cleaner baseline in gradient runs.

Here's two overlaid chromatograms, both no-injection blank runs of the same gradient method:



Guess which one used AR grade methanol + triple-distilled water, and which one used HPLC-grade methanol + Milli-Q water...

Quote: Originally posted by unionised

I always wondered what would happen if you tried vodka as a mobile phase...

Quote: Originally posted by phlogiston

Before you had the HP software, how were you programming the pump? Does it let you program gradients manually?

Yes, you can program all of the functions via the front panel buttons. It appears that the GPIB card inside each module which is used for comms with the PC was actually an optional extra - so these were all designed to be usable without Chemstation.

--

Here's my current HPLC workbench (as usual, please excuse the mess):


[Edited on 13-10-2018 by DavidJR]