Example of using Amount in Empower?

Empower2018

I have seen so many ways to use Amount in Empower but cant seen to grasp the fundamentals. I want to calculate the amount (in %w/w) in my unknown samples. My samplweight is 250mg dissolved in 25mls for sample 1 and my sample 2 is a 2 in 20 dilution of sample 1. I don't have the purity tab (Its Empower 2) so do I put the concentration value (in mcg/mL) for the component editor and leave my Sampleweight field and dilution blank for standards but populated for samples? What would my dilution be for Samples 1 and 2- 25 and 0.5?? I use two bracketing standards for calibration by the way. 

Sorry if this seems really basic but I still cant get it. Thanks in advance for your help.

MJS

You touch on a few different issues within this, so I'll do my best to answer/explain:

Standard Purity:
In Empower 2, you could just enter a µg/mL concentration (or mg/mL or whatever you want) and leave the sample weight/dilution each as 1.  To get sample results in anything other than these original units, you have a bit of work to calculate the correct dilution values, but it is certainly possible and common enough to do this. 

Alternately, you can work around the lack of a purity tab by utilizing the sample weight field.  You enter a standard's mass in mg for the standard (or µg or whatever mass value you want), purity as a factor (so 0.99 for 99% rather than 99) in the sample weight field, and the applicable dilution in the dilution field.  Empower would then generate the cal curve in mg/mL (or µg/mL, etc. based on your dilution's unit conversion). The basic formula Empower uses on the standards is "CalibrationFactor=Areastd/(Value*SampleWeight/Dilution)" when generating the cal curve (note that "value" is the field name if you ever need to use the standard's entered amount in a custom field or show it on a report which is something I do in order to show on a report what amount an analyst entered for the stds).

Sample Dilution/Units:
For samples, it reverses the above, effectively.  "Amount=Concentrationsample*dilution/sampleweight" where  "Areasample/CalibrationFactor=Concentrationsample."  So, if you are looking for a %w/w, the key is that you need to enter the appropriate values in for the sample weight and dilution to get the units correct.  Remember that this is then dependent upon what you are entering in for the standards for any possible unit conversion.

So, if I assume you measured something like 100 mg into 10 mL for the standard to get a 10 mg/mL standard solution.  I'd enter the 100 mg in for the standard in the component editor, whatever purity I have as a factor in the sampleweight field, and 10 for the dilution field.  Empower would then generate a cal curve in mg/mL vs area response.

For sample 1, you say 250 mg sample dissolved in 25 mL.  So, that would mean 250 goes into the sample weight, so what goes into the dilution?  Empower would take the area count and calculate a "concentrationsample" in mg/mL per the standard curve.  If we follow through the rest of the calculation...concentrationsample*dilution=amount in mg, amount in mg/sample weight=amount in mg per mg of original sample, so w/w.  If you want to go the step further to convert this to a %w/w rather than just the w/w, then you'd need the dilution field to account for the percentage conversion and use a value of 2500 instead of 25.

Whenever calculating dilution factors for samples for Empower, the trick is to simply invert the normal mathematical formula...for example:
If sample is diluted in x (mL) then a secondary dilution is performed where y (mL) of the dilution was further diluted in z (mL) to get a final sample, you would try to calculate the concentration as mass/x*y/z.  For a sample dilution field, you invert this so the dilution=z/y*x to get the mass of the sample out of Empower as the amount field.

Hopefully this helps.  It's a bit tricky to write any formula so feel free to follow up if it seems like I have any typos.

HeatherLongden

Brilliant detailed response... but I know that its hard to write that out in text. You might find online a presentation called.... How to get from Area to Amount. Waters created it for an Inform in 2004 I think.

• As you didn't mention Standard Purity, I don't think you need the purity field at this point. This adjusts for less ( or more) than 100% pure reference standard.
• Use Amount rather than Concentration if your injection volumes are the same
• The unit you enter for the Std is the SAME unit you will get your answer out in until you adjust it with Dilution and Sample Weight as MJS suggests.

To be sure you are using this right, you could take a training course, have Waters send in an expert consultant to walk you though it or, take a set of data and calculate by hand or spreadsheet, to help guide you to use the fields in the right combination. It is unlikely that you will need any custom fields to get this done.

Empower2018

Thank you so much for your detailed and thorough response MJS, that is the clearest answer to using Amount I have yet to read. I have printed out your response and intend to use it when I get a chance in the lab over the next week or so. Thanks also to Heather for the reply. Both answers are very welcome.