[nightmare00000]

Hello,

I would like to ask you something about FTIR spectroscopy.
I've had this in my head... - why do the ATR is not suitable for quantity measurements, and we need to use transmission (KBr pellets). In quality measurements there were no problems.


Regards

[Corribus]

Mostly because it's difficult to control the absorption path-length in the ATR configuration.

[nightmare00000]

Thank you Corribus. Could you please write me more about it? What you mean writing - hard to control the absorption path-length?

[Corribus]

Do you know how you determine concentration using a spectrometer?
Now compare the way spectra are acquired via a conventional transmission expt in FTIR and how the sample is prepared in ATR. What are the differences?

[nightmare00000]

In ATR generally there is no sample preparation - we just put the sample onto crystal and that's it. In transmission we need to use specific amount of analyze material and mix it with specific amount of for example KBr or in case of liquid we can put the drop between two KBr pellets.

We determine the concentration of the sample basing on the calibration curve made of few sample, where we are sure of their concentration. We need to state the peak on the FTIR spectra which is nice separate from others and basing on it's absorption change of measured samples they are "fit" to curve and the concentration of measured sample is calculated.

[Corribus]

Ok, for the most part nothing is wrong with anything you wrote, but it's also a very applied version of what's going on. To answer your question we'll have to consider what actually is responsible for the measured signal and investigate the assumptions involved in the experiment. Thankfully we can do this without really touching upon the physics of absorption events.

In a transmission expt, you're not actually measuring concentration directly. The spectrometer essentially measures the intensity of light after it has passed through the sample (as a function of light wavelength) and compares that value to a amount of light that strikes the detector when the sample isn't there. The ratio, known as the transmittance, can be converted into an absorption value, which is related by a logarithmic function. The absorption value, A, to a first approximation, is linearly proportional to three parameters, the path length for absorption, the concentration of absorbers, and a molecular parameter called the extinction coefficient. (The extinction coefficient can be thought of as the probability that a molecule will absorb a photon when a photon and the molecule collide.) This is known as the Beer Law or Beer-Lambert Law.

https://en.wikipedia.org/wiki/Beer%E2%80%93Lambert_law

If there are more absorbers in the way of the light, the amount of light that will be absorbed increases, which translates into lower transmittance as measured by the instrument. The amount of absorbers in the beam is related to the density of absorbers (how many absorbers per unit of volume), and how much of the sample the light has to pass through before it gets to the detector. The latter parameter is the path length. To remove this confounding factor, most absorption experiments utilized a standard path length value so that changes in measured absorbance only depend on changes in absorber density (concentration). This assumes you are in a situation where the Beer-Lambert law holds, and where the molecular extinction coefficient isn't changing.

So - in the transmission experiment this allows you to build up your concentration calibration line by measuring standards of known concentration, all using the same sample design (path length) because the signal is proportional to the absorbance in this case.

In the ATR configuration, however, it is hard to control how much sample the light passes through, and so the signal is no longer strictly proportional to the measured concentration. There are a lot of potential causes, many of them related to the physics of the ATR experiment. To some extent the difficult is dependent on the type of sample (thin film, powder, etc.). But briefly - it's not just a simple matter of putting the sample on the crystal and that's it. The evanescent field passes out of the crystal and into the sample only by a length scale of (at most) a few microns. So the absorption is really sensitive to the contact between the sample and the crystal. Contact is often attenuated by applying a top-down force to your sample, but defects or varying degrees of roughness on the sample surface (for a solid) can significantly impact this efficiency of contact. For a powder, how much pressure you apply can impact the amount of air spaces between powder particles, which again impacts the effective path length. For a solution, where the solution is dropped onto the crystal, contact is usually very good, but varying rates of evaporation and interactions of solute molecules with the crystal can create a dynamic process that may be hard to duplicate from one sample to the next.

Another effect that can be considered is that the amount of pressure applied can actually change the extinction coefficient of the absorber at the measured wavelength. This is kind of equivalent to the same kind of breakdown of the Beer-Lambert law you might observe in a transmittance experiment when your absorber is at too high of a concentration. (Basically, when molecules get really close together, this can modulate the molecular energy states that give rise to the absorption events.)

In the end it's not impossible to get quantitative information from the ATR experiment, but it's very difficult. So the technique is usually thought of as semi-quantitative as a means of measuring concentration.

[wildfyr]

Corribus, I'd just like to congratulate you for an absolutely superb answer.

[nightmare00000]

Many thanks to you for your time and comprehensive answer.
I'm the beginner user of FTIR spectrometer and I'm trying to get knowledge about it. I read about the basics of FTIR, but I still have some question.

If we are in topic of quantitative analysis could you tell me some advices how to select good peak to calibration curve and what's better - peak high or peak area?

[Corribus]

An intense, sharp peak is probably better for calibration, as it will be easy to tell if there is an energy shift or other aberrations due to concentration. Also, you will get a better detection limit using a strong peak - although possibly you may also approach a saturation limit at lower concentrations.

[nightmare00000]

I'm back with the question, again connected with quantity measurements.
I have problem to solve connected with mixture analysis.

For now theoretically I know for example - how to quanitfy isopropanol in water (take few samples with known concentration build calibration curve, basing on the nice peak area/high, and when I take sample to measure, in which I don't know the concentration -> measure it and the FT-IR software will predict the concentration basing on calibration)


I will explain it on some theoretical "example":

Mixture is prepared from substance X (solid state), substance Y (liquid), and substance Z (liquid) -> it results in syrup state mixture.


Questions:
1) Sample preparation -> could be this syrup state placed between two KBr pellets to mesurements? (as I know it now - ATR can't be used in quantity measurements).
2) I would like to know how much of each substance is in the mixture, so do I need to build three calibration curves? What about choosing the good peak area/high? Do I need to observe which peak is changing when we have other concentration of each substance?


I wrote this intricately, but I think you will know what's my problem to solve out...

Thank you.

[Corribus]

1) Sample preparation -> could be this syrup state placed between two KBr pellets to mesurements? (as I know it now - ATR can't be used in quantity measurements).
2) I would like to know how much of each substance is in the mixture, so do I need to build three calibration curves? What about choosing the good peak area/high? Do I need to observe which peak is changing when we have other concentration of each substance?

1. In theory you can take two KBr (or other transparent material) windows and sandwich a sample between them. We used to have a small device that held two windows and then you inserted a teflon spacer (so you know the pathlength) to create an assembled optical cell. I say in theory because in practice it was messy - hard to get a liquid to stay sandwiched between them without dripping all over the place. Also your path length will be very short, which may be ok for some samples but may not give you enough absorptivity for others. What will be important is to make sure your path length is well controlled, otherwise you run into the same issue as before for quantification. A better solution TBH would be to just buy some FTIR cells. Just make sure you get ones that are transparent in the IR region of interest.

http://www.specac.com/products/liquid-transmission-cell/quartz-cells-and-cuvettes/666

Depending on viscosity a syrup may or may not be hard to use with one of these. Note that it's possible that a syrup may give you reasonable results on an ATR accessory because contact should be good and reproducible from sample to sample. It may be worth trying out.

2. Assuming each substance has an absorbance feature that does not overlap with features from other substances in the mixtures, building your calibration curves should be pretty straightforward. It would be best to do this with solutions of each substance independently, although bear in mind that the nature of the solution can impact where absorption features show up. You will need to verify that the absorption feature doesn't change frequencies over the concentration ranges of interest.

Being honest, absorption frequencies of molecules in FTIR can be extremely sensitive to the composition of a substance, particularly because many FTIR absorptions are very narrow. In my experience this makes FTIR often a less-than-ideal technique for measuring concentrations. Compare this to UV-VIs spectroscopy, in which electronic transitions tend to be very broad, so small changes in absorption energies are unlikely to interfere with the measurement. Not to say it can't be done, but FTIR generally far more useful as an identification technique than a quantification technique.

[nightmare00000]

Thank you Corribus.

Good idea with the ATR. It could be the fastest and the easiest way.
But do we have some limitations in ATR - I mean the concentration level which could be measured?
And what about practical aspect - do we need to put always the same drop of sample or it is not important due to nice contact of syrup to diamond crystal (it only measure penetrating in few μm deep)?

[Corribus]

Your sensitivity will depend on what kind of ATR you have. The number of bounces, the incidence angle, etc, all affect the effective path length.
As long as the liquid covers the entire sample, the volume shouldn't matter, but you need to take care about evaporation if that's an issue. Also, because the ATR only measures a few microns at the crystal/environment interface, any preferential adsorption of solutes onto the crystal surface can, I imagine, skew your results. (This would also happen in a transmission experiment, but because transmission assays the entire sample volume, the effect is negligible.) I.e., imagine if your diamond surface is nonpolar but your solution is polar, and your analyte is somewhat nonpolar. It could adhere to the diamond surface giving an artificially high concentration (and altered FTIR spectrum), especially during long experiments. This is just something you'd have to investigate.

Bear in mind, I've never used FTIR-ATR to measure concentrations. So, I'm just ruminating on what all the problems might be...