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Topic: Colour Measurement  (Read 2547 times)

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Offline sci0x

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Colour Measurement
« on: April 03, 2017, 09:09:29 PM »
Hello,
I'm measuring the colour on a highly viscous liquid.
I dilute x5 and then x100 and measure the OD on a spectrophotometer.
Because I have to do such a big dilution I want to be accurate so I've started using a micro-pipette instead of a glass pipette and the results can vary.
What experiment can I carry out to ensure which pipette is giving me the correct result?
Cheers
:)

Offline Corribus

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Re: Colour Measurement
« Reply #1 on: April 03, 2017, 09:37:06 PM »
Can you do it gravimetrically?
What men are poets who can speak of Jupiter if he were like a man, but if he is an immense spinning sphere of methane and ammonia must be silent?  - Richard P. Feynman

Offline sci0x

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Re: Colour Measurement
« Reply #2 on: April 03, 2017, 10:26:26 PM »
My background isn't in Chemistry so you'll have to bare with me.
Would this involve instead of using a pipette for measuring, using a balance for dilution?

Offline Corribus

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Re: Colour Measurement
« Reply #3 on: April 03, 2017, 11:36:17 PM »
Yes.

Unless they are specifically designed for it, many micropipettes are not really good for measuring volumes of non-aqueous liquids.  Your opening post isn't very descriptive about what you're trying to do (what are you diluting your oil with?), and what kinds of volumes you are working with. You can do your dilutions volumetrically, but this assumes you have confidence in your ability to measure out volumes. A pipette will work if it's designed for your solvent. If it's not, a high quality burette or volumetric flask (or both) is a good way to go.

If you don't have a great way of making accurate volumetric dilutions, you can make your dilution using an (analytical!) balance. The only caveat is that your dilutions will be mass based rather than volume based. If you know the density of the liquids you are working with, you can easily convert to liquid based dilutions. Actually, I do most of my dilutions using a balance because I find it to be easier.

Note that the choice will often come down not just to convenience, but the accuracy and precision needs of your application. If all you are interested in is how your various pipettes are doing, measure out 1 mL, weigh it, and see which one comes closest. This again requires that you know the density, of course. And that your balance is properly calibrated. :)

(And just to be annoyingly pedantic about it - sorry but I can't help it; I'm a spectroscopist by training - a spectrophotometer measures absorption, not color.)
What men are poets who can speak of Jupiter if he were like a man, but if he is an immense spinning sphere of methane and ammonia must be silent?  - Richard P. Feynman

Offline sci0x

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Re: Colour Measurement
« Reply #4 on: April 03, 2017, 11:59:45 PM »
Thank you for your detailed response.
I'm diluting my viscous liquid with deionised water. I dilute 100ml of my liquid measured by volumetric and dispense into 500ml volumetric and make up to the mark water dH20. I filter the sample, and then measure 5ml of this diluted sample into 500ml volumetric flask and make up to the mark with water. I then blank the spec, and then measure the adsorption at 430nm, which I convert to EBC, and multiplying up my dilutions to get the colour of the original liquid.
I get variation in colour if I'm using a 5ml glass pipette for or 5ml measured by 1ml micropipette.
I have an alcolyser, I can get the density of the liquid when I do my 1 in 5 dilution. Its too viscous to get the sample without first diluting.
Whats the best way to perform this dilution if I use an accurate balance and use the density of the liquid?
Thanks!
« Last Edit: April 04, 2017, 12:10:46 AM by sci0x »

Offline Corribus

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Re: Colour Measurement
« Reply #5 on: April 04, 2017, 10:15:39 AM »
If you have good micropipettes and they have been calibrated, then these will be perfectly acceptable for making accurate dilutions. I'm not sure what you mean by a "glass pipette". If you mean a good quality serological pipette, then these should also be acceptable. If you are getting different absorption readings by making your dilutions in these two different ways, then either your technique is poor for one or both methods, or your micropipette isn't calibrated. You can easily check which method is more accurate by dispensing 1 mL of water onto a properly calibrated analytical balance. You should measure out 1 gram (or whatever the density is of your water).

Is your error consistent in the same direction? For example, is the UV-Vis reading when using the micropipettes consistently less than when using the serological pipette? If you notice a consistent error like this, it can help to tell you about what the problem is. But the first thing to do is (1) ensure your pipettes and balances have been calibrated, and (2) verify that your technique is good. 99/100 times, issues with dilutions come down to one of these two categories.
What men are poets who can speak of Jupiter if he were like a man, but if he is an immense spinning sphere of methane and ammonia must be silent?  - Richard P. Feynman

Offline sci0x

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Re: Colour Measurement
« Reply #6 on: April 04, 2017, 08:38:55 PM »
Thank you for your help, i'll try this and also try doing the dilutions gravimetrically and let you know how I get on :)

Offline the inquisitor

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Re: Colour Measurement
« Reply #7 on: April 06, 2017, 07:14:43 AM »
sci0x,

If you haven't done so already, check that the pipette tips that you are using with your micropipette are suitable for your application.

Is there any liquid left in the pipette tip after you dispense your aliquot? If not, then I would rule this one out.

Kind regards.

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