Name: improving infrared spectroscopy characterization of soil organic matter with spectral subtractions
Uploaded: 2019-01-10T12:30:00
Description: Watch this Scientific Journal Video about Improving Infrared Spectroscopy Characterization of Soil Organic Matter with Spectral Subtractions at JoVE.com

We appreciate the guidance from Dr. Randy Southard on NaOCl oxidation and various discussions of spectral subtractions with Dr. Fungai F.N.D. Mukome.

1. Prepare Soil for Non-treated DRIFT Spectroscopy and SOM Removal
<ol>
	<li>Sieve the soil to &#60; 2 mm using a stainless-steel mesh (the &#39;fine-earth fraction&#39;). 
	NOTE: This demonstration employs two soils of similar texture but a nearly 3-fold difference in total SOM content (Table 1).</li>
</ol>
2. SOM Removal by Chemical Oxidation: Example of NaOCl
<ol>
	<li>Adjust the pH of 6% w/v NaOCl to pH 9.5 by adding 1 M HCl dropwise to the solution while mixing and m...

The method of removing SOM carries two considerations: 1) the amount of SOM removed, and 2) absorbance artifacts in the resulting mineral reference spectrum. It is fortunately possible&#8212; and arguably necessary&#8212; to identify and quantity these issues in order to avoid biased interpretations of SOM composition from the resulting subtraction spectrum. Ideally, spectral subtractions would employ a mineral-only reference spectrum to yield a spectrum of &#39;pure&#39; SOM. In reality, the resulting subtraction spectr...

Soil organic matter (SOM) is a minor constituent by mass in most soil samples but is implicated in multiple properties and processes underlying soil functions, such as nutrient cycling and carbon sequestration1. Characterizing the composition of SOM is one of several approaches to link SOM formation and turnover with its role(s) in soil functions2,3. One method of characterizing SOM composition is Fourier transform infrared (FTIR) spectroscopy, which offers detection of functional groups that constitute organic matter in soils and other environmental samples (e.g., carboxyl C-O, aliphatic C-H)4. However, the utility of FTIR spectroscopy for revealing SOM functional group composition is challenged by the dominant mineral component for the majority of soils (typically &#62; 95% mass) due to strong inorganic absorbances that challenge or severely limit detection and interpretation of organic absorbances.
Spectral subtractions offer a way to improve FTIR spectroscopic characterization of organic matter in soil samples. Subtracting mineral absorbances from the soil spectrum can be used to enhance absorbances of organic functional groups of interest in the analysis of SOM composition
(Figure 1).
Advantages of spectral subtractions over standard FTIR spectroscopy (i.e., soil spectra) include:
(i) Improved resolution and interpretation of organic absorbance bands compared to normal soil spectra. Though interpretation of organic bands in soil spectra can be performed by assuming that the relative differences in absorbance are due to differences in organic functional groups, this limits comparisons to samples with the same mineralogy and relatively high SOM content, and may be less sensitive to changes in organic bands, even those considered to be relatively mineral-free (e.g. aliphatic C-H stretch)5
(ii) Analysis of soils beyond high SOM samples or organic matter-enriched extracts or fractions
(iii) Highlighting changes induced by experimental treatments from mesocosm to field scales6
Additional applications of spectral subtractions in FTIR analysis of SOM include complementing structural and molecular characterizations (e.g., NMR spectroscopy, mass spectrometry)5,7, identifying the composition of SOM removed by an extraction or destructive fractionation8, and fingerprinting SOM composition for forensic purposes9. This method is applicable to a wide variety of mineral-organic mixtures beyond soils, including sediment10, peat11, and coal12,13.
The potential of spectral subtractions to improve FTIR spectroscopic characterization of SOM is demonstrated using examples of organic matter removal to obtain mineral reference spectra, and then, using these mineral reference spectra, performing and evaluating ideal and non-ideal spectral subtractions. This demonstration focuses on diffuse reflectance infrared Fourier transform (DRIFT) spectra collected in the mid-infrared region (MIR, 4,000 - 400 cm-1), as this is a widespread approach for the analysis of soil samples4.
The two example methods of SOM removal for obtaining a mineral-enriched reference spectrum are (i) high-temperature combustion (&#39;ashing&#39;) and (ii) chemical oxidation, using dilute sodium hypochlorite (NaOCl). It should be noted that these are examples of commonly employed SOM removal methods, rather than prescriptive recommendations. Other methods of SOM removal may offer reduced mineral artifacts and/or enhanced removal rates (e.g., low-temperature ashing)14. High-temperature ashing was one of the first methods used to obtain mineral-enriched reference spectra for performing subtractions, initially for OM-enriched samples derived from soils (e.g., dissolved organic matter, litter)15,16 followed by its application to bulk soil samples17,18. The example chemical oxidation used to remove SOM is based on the method of NaOCl oxidation described by Anderson19. This was originally developed as a pretreatment for removing organic matter in soil samples prior to X-ray diffraction (XRD) analysis, and has been investigated as a potential chemical fractionation sensitive to SOM stabilization20,21. Both high-temperature removal and chemical oxidation using NaOCl can entail soil-specific artifacts and have limitations on spectral interpretation that should be considered when selecting a method of SOM removal14,22.

<table border="0" cellpadding="0" cellspacing="0" width="1093">
	<tbody>
		<tr height="19">
			<td height="19" style="height:19px;width:263px;">Nicolet iS50 spectrometer</td>
			<td style="width:180px;">Thermo Fisher Scientific</td>
			<td style="width:128px;">912A0760</td>
			<td style="width:523px;">infrared spectrometer used to collect spectra</td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:263px;">EasiDiff</td>
			<td style="width:180px;">Pike Technologies</td>
			<td style="width:128px;">042-1040</td>
			<td>high throughput sample holder</td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:263px;">OMNIC</td>
			<td style="width:180px;">Thermo Fisher Scientific</td>
			<td style="width:128px;">INQSOF018</td>
			<td>software used to perform subtractions</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">6% v/v sodium hypochlorite</td>
			<td style="width:180px;">Clorox</td>
			<td style="width:128px;">n/a</td>
			<td>generic store-bought bleach for oxidative removal of soil organic matter</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">Type 47900 Furnace</td>
			<td style="width:180px;">VWR International</td>
			<td style="width:128px;">30609-748</td>
			<td>muffle furnace for ashing soils to removal soil organic matter</td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:263px;">VWR Gooch Crucibles, Porcelain&#160;</td>
			<td style="width:180px;">VWR International</td>
			<td style="width:128px;">89038-038</td>
			<td>crucibles for ashing</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">VWR Tube 50 mL Sterile CS500&#160;</td>
			<td style="width:180px;">VWR International</td>
			<td style="width:128px;">89004-364</td>
			<td>for sodium hypochlorite</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">Forced air oven</td>
			<td style="width:180px;">VWR International</td>
			<td>89511-414</td>
			<td>for drying soils after oxidation and water washes</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">VersaStar pH meter</td>
			<td style="width:180px;">Fisher Scientific</td>
			<td>13 645 573</td>
			<td>for measuring pH of oxidation solution</td>
		</tr>
	</tbody>
</table>

improving infrared spectroscopy characterization of soil organic matter with spectral subtractions

Soil organic matter (SOM) underlies numerous soil processes and functions. Fourier transform infrared (FTIR) spectroscopy detects infrared-active organic bonds that constitute the organic component of soils. However, the relatively low organic matter content of soils (commonly &#60; 5% by mass) and absorbance overlap of mineral and organic functional groups in the mid-infrared (MIR) region (4,000-400 cm-1) engenders substantial interference by dominant mineral absorbances, challenging or even preventing interpretation of spectra for SOM characterization. Spectral subtractions, a post-hoc mathematical treatment of spectra, can reduce mineral interference and enhance resolution of spectral regions corresponding to organic functional groups by mathematically removing mineral absorbances. This requires a mineral-enriched reference spectrum, which can be empirically obtained for a given soil sample by removing SOM. The mineral-enriched reference spectrum is subtracted from the original (untreated) spectrum of the soil sample to produce a spectrum representing SOM absorbances. Common SOM removal methods include high-temperature combustion (&#39;ashing&#39;) and chemical oxidation. Selection of the SOM removal method carries two considerations: (1) the amount of SOM removed, and (2) absorbance artifacts in the mineral reference spectrum and thus the resulting subtraction spectrum. These potential issues can, and should, be identified and quantified in order to avoid fallacious or biased interpretations of spectra for organic functional group composition of SOM. Following SOM removal, the resulting mineral-enriched sample is used to collect a mineral reference spectrum. Several strategies exist to perform subtractions depending on the experimental goals and sample characteristics, most notably the determination of the subtraction factor. The resulting subtraction spectrum requires careful interpretation based on the aforementioned methodology. For many soil and other environmental samples containing substantial mineral components, subtractions have strong potential to improve FTIR spectroscopic characterization of organic matter composition.

The method of SOM removal has practical as well as theoretical implications for the interpretation of subtraction spectra. For example, mineral alterations from high temperature ashing can manifest as losses or appearances of peaks and/or as shifted or broadened peaks in the mineral reference spectrum. These spectral artifacts are prone to occur in regions of overlap with organic bands at 1,600-900 cm-1,22 compromising interpretation of organic bands. Co...

Watch this Scientific Journal Video about Improving Infrared Spectroscopy Characterization of Soil Organic Matter with Spectral Subtractions at JoVE.com

Improving Infrared Spectroscopy Characterization of Soil Organic Matter with Spectral Subtractions

SOM underlies many soil functions and processes, but its characterization by FTIR spectroscopy is often challenged by mineral interferences. The described method can increase the utility of SOM analysis by FTIR spectroscopy by subtracting mineral interferences in soil spectra using empirically obtained mineral reference spectra.

Soil organic matter (SOM) underlies numerous soil processes and functions. Fourier transform infrared (FTIR) spectroscopy detects infrared-active organic ...

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