In order to test soil potential denitrification rate (acetylene inhibition, and analyse the N2O at intervals of 1, 2, 3, and 4 h) in lab. Do you add the same volume of gas into the closed system to keep the pressure constant when gas sample is taken out at each interval?
Yeah, I think it is better you add same amount of nobele gas like argon, to compensate the volume lossed.
Hi, Shu Xin, as stated, I think it is better you add same amount of nobele gas like argon, to compensate the volume lossed.
If you are collecting small volumes (>10 mL) you might consider over pressurizing the vial with sufficient gas at the start of the incubation that you do not need to add gas during the incubation. This makes the calculations much simpler. If you add gas during the incubation you must calculate the dilution of the headspace for each gas addition.
Whether you pressurize system or add inert gas or do NOT compensate withdrawn gas volumes, you change the pressure in your incubation flask. It does affect the calculated denitrification rate. Thus one of the solutions is to recall gas law equations (pV/T=const) and refine your calculations by taking into account sacrificed gas volumes. It could be useful to keep pressure record.
Another solution is to have very large volume of incubation flask (e.g. 1 L) with cumulative sampling size small (e.g. 10 ml or less). Then you can neglect pressure reduction caused by sampling.
The best solution is to use continuous system, when inert gas (N2, Ar, or He) is continuously flowing through soil column together with inhibitor (acetylene) and you measure the gaseous product (nitrous oxide) in the outflow gas stream. It is so called dynamic method that is not sensitive to withdrawn samples and is very accurate.
You can do single term sampling, no doubts. But you will miss all the elements of transient response (possible lag phase, acceleration, decline, etc). All depends on objectives of your particular study.
The easiest way in this case is addition of similar volume of noble gas to the volume of sample extracted and considering the dilution factor as you know the total volume of your incubation bottle
You know the volume of the headspace. After adding a specific volume of noble gas you can include this in the following sampling event. so the N2O gas dilution with noble gas should be = (headspace vilume + noble gas volume)/headspace volume. For the next sampling event you can add up noble gas volume in your calculation. I hope this will give a reasonable measurement of denitrification rate
If you measuring gas accumulation over multiple sampling times don't forget to subtract the volume of gas you removed and keep track its N2O content. This becomes a bit complicated as you account for the amount of N2O dissolved in the water at each time step (in which case it is only the amount of N2O in the headspace at the time of sampling). This ends up being an exercise in both accounting and chemistry.
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Thank you, @ Nicolai S Panikov. That is to say, if I only collect sample gases at one time, the result is still reasonable to present soil denitrification rate?
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It is hard to say without knowledge on your experimental design. If you make survey with plenty of diverse samples -- perhaps one-time sampling should be OK. In a more mechanistic study (if you want to decipher underlying mechanisms) it would be NOT good. My advice is just to increase the ratio 'total headspace volume : sampled volume' to 100 or higher by using larger containers and more soil and smaller gas samples. If the cumulative samples withdrawal will be within 5% of the total headspace volume -- no problems.
Correction discussed by David Burton and Jahangir are fundamentally correct, no doubts. I used Excel to implement such corrections at each sampling step (be careful -- it is easy to underestimate products). However it is impossible to account all sources and sinks for N2O, such as gas solubility in soil water (you can't apply equilibrium formula based on Henri law).
An important aspect of measuring denitrification in soil is to account for nitrous oxide entrapped in soil (besides the gas dissolved in solution phase. This is because, especially in closed systems all nitrous oxide produced within soil does not come out in the headspace. This error can be avoided by thoroughly shaking the incubation vessel just before sampling...it is easy when denitrification potential is measured in the presence of ample amount of water. However, in case of soil core, after sampling the headspace, entire core is removed from the incubation jar, disrupted and water is added to make slurry in an other gas tight jar. After shaking this slurry for few minutes, the entrapped nitrous oxide comes out in headspace. Therefore, measuring denitrification in a closed system needs to account for nitrous oxide in the headspace PLUS that dissolved in the solution phase PLUS that entrapped within soil matrix. There are only few reports on this important aspect of quantifying denitrification.
Hi Xin,
I have two papers on field measurement of actual denitrification rates (not potential denitrification rate) which also include measurements of N2O and CO2 entrapped in soil. Besides, you can also find some more related papers referred in these two publications.
1. Mahmood T., Ali R., Azam F., Malik K.A. (1999) Comparison of two versions of the acetylene inhibition/soil core method for measuring denitrification loss from an irrigated wheat field. Biology & Fertility of Soils 29: 328-331.
2. Mahmood T., Ali R., Sajjad M.I., Chaudhri M.B., Tahir G.R., Azam F. (2000) Denitrification and total fertilizer N losses from an irrigated cotton field. Biology & Fertility of Soils 31:270-278.
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