I recommend not to extract and fractionate as you will alter what was previously there. Instead, directly run 13C CPMAS NMR on air-dried finely ground samples. When the signal-to-noise ratio is poor, treatment with hydrogenfluoride to reduce mineral content is needed.

You need to ask an expert on 13C NMR. I think it is nescessary in order to avoid iron from interfeering and to get a higher concentration of 13C. Usually HF is used (don't use this dangerous compound onless you know what you are doing). Maybe you can ask Heike Knicker from germany or somebody else that have done this. Sander

Why do you want to extract the acid? Ypu can perform the 13CP MAS NMR directly on the soil, I recomend you to use a 600 MHz Magnet to imprve the sensitivity

Actually you have two different possibilities to analyze your samples by CPMAS 13C NMR spectroscopy. The first one is that you already have read: extraction and purification. In this case you will get very nice spectra which are quite understandable and easily to interpret. Just you have to check carefully all your NMR parameters in order to be sure about the quantitativeness of your spectra. The second way to proceed is to run NMR experiments directly on the soil sample. In this case the quality of your results depend strongly on the soil properties. I mean that NMR sensitivity is strongly influenced by the paramagnetic content (either organic or inorganic). Paramagnetic content strongly reduces NMR sensitivity. In addition you have to be sure that the carbon content of your soil system is at least >2%, if paramagnetism can be neglected. In this latter case, when paramagnetism is neglectable, you may increase carbon conentration by "washing" your sample with a 10% HF solution. However, the same question on what you destroy still stay. In other words you base your experiments on the assumption that your organic material is kept undisturbed by the treatment This is the same assumption you use when you analyze you extracted humic systems. In my opinion you can analyse your organic material either in soils or extracted depending on the amount of machine time you have and the quality of the spectra you want get.

HF demineralization prior to 13C NMR work can have very significant impact on your result. You need to be very careful when using this chemical first of all and secondly in some instances it has been shown that this procedure can lead to loss of upwards of 50% of the C in your sample ... only use HF if there is no way you can get well resolved spectra with your sample (if your %C is too low, concentration of paramagnetic species is high, and if long run times, higher spinning rate, low field magnets, etc can't help with your samples.

I think you should consult people with experience on application of NMR in soils.

Its better to use untreated samples for 13C NMR analysis. Any chemical treatment alter the quality of soil organic matter. Therefore, chemically treated soil give you a partial result. However, HF treatment is common for 3C NMR study might be to increase the OC content of soil (>2%). HF treatment extracts mineral bound organic carbon from soil. The HF extraction residue mostly represents the most stable recalcitrant organic matter fraction accounting nearly 50% of soil organic carbon. In few cases (Mahieu et al, 2000, Schmidt-Rohr et al, 2004) 13C NMR analysis was done on SOC fractions (humic and fulvic acid and humin) isolated by wet chemical fractionation method. However, it depends on your objective.

I have my own experinces with 1HNMR study on the HS (HA, FA) from bottom sediments with prior extraction and purification of extracts. After them I have obtained very nice data . I suggest to HS extraction before detail analysis and after them data will comparable to earlier publishes data

No

I agree with Dr. Conte. The "appropriate" method depends on your research question. Humic and fulvic acids are operationally defined based on solubility. If I were doing a study where I'm only interested in acid or base-soluble organic matter, or dissolved organic matter, I would likely analyze them using solution-state (likely 1H because of abundance, NMR sensitivity, and resolution) rather than solid-state NMR. If I wanted to analyze total OM, I would do solid-state 13C NMR, with the minimum treatment that my sample would allow (depends on C content, amount of paramagnetic species - as described by other responses). We published a paper in Environmental Chemistry (Clemente et al. 2012) that compares data from solid-state 13C NMR and solution-state 1H NMR. Although data from solid-state 13C NMR of untreated (ideal) samples is considered more representative, the amount of structural information that you get from these experiments is also limited by severe signal overlap. On the other hand, you may have more NMR experiments available; and anisotropic interactions (which leads to line-broadening in solid-state) are averaged by free rotation of molecules in solution-state. However, you are limited to analyzing extractable components.

I suggest you to read the following two articles which gives an in depth answer to the question.

First of all you may have a clear background on the necessity of analyzing soil organic matter through solid state 13 C NMR.

Please note, soil organic matter is not a single entity and it consists of thousands of compounds and each one is different with different structure and biochemical nature. We can not isolate each one and estimate and can be correlated to the soil properties. Instead we group them to different categories and then try to correlate with different soil properties. The most popular such groups are labile and recalcitrant pools - the labile group consists of carbohydrate or sugar like compounds having O-alkyl groups as identified by NMR while the recalcitrant groups consists mainly of lignin, tannin or chitin type compounds having long chain alkyl skeleton again identifiable through NMR. The ratio of these groups broadly give us the relative abundnace of labile or recalcitrant pool of C compounds, hence we can know about the soil organic matter quality.

Essentially what is the relevance of labile or recalcitrant groups ? Hopefully you know that these compounds behaves differently to microbes. labile ones are easily attacked by microbes, hence the mineralization rate would be much more or soil biological activity will be very high. Vice versa in the case of recalcitrant pool.

As far as NMR probing is concerned, I strongly belive that the soil as such should be used and no chemical modification to be made. When using an alkali to separate the so called humic or fulvic acid what is resulted- a modified or chemically altered group of compounds from soil. What would be the relevance of estimating such modified or chemically and structurally altered compounds with respect to the to the soil properties ?

Of course you may find difficulty in mineral soils where paramagnetics like Fe content is more. You may have to go for a powerful NMR tool such as 600 or more MHz.

If time allows read the following articles. One is mine and the other is a review giving valuable info.

Joshua Abraham and J.A.Chudek 2008, Studies on litter characterization using 13 C NMR and assessment of microbial activity in natural forest and plantation crops' (teak and rubber) soil ecosystems of Kerala, India. Plant and Soil. 303(1&2) :265-273

Kleber, M. AND M. G. JOHNSON. Advances in understanding the molecular structure of soil organic matter: Implications for interactions in the environment. Advances in Agronomy. ELSEVIER, AMSTERDAM, Holland, 106:77-142, (2010).

you can do whole soil analysis; you may see some of my papers or Robert Cook's work http://www.collectionscanada.gc.ca/obj/s4/f2/dsk3/ftp04/nq24531.pdf