Does Temperature Affect the Rate of Nitrification and Dentrification ?

Jamie White

9th grade

Byng Jr. High

INTRODUCTION

The Earth’s atmosphere is seventy-eight percent nitrogen gas, but plants and animals cannot use nitrogen directly from the air as they do carbon dioxide and oxygen. (3) Plants depend on the nitrogen cycle to covert nitrogen, an essential constituent of protein, which is a building block of all living materials, into a useable form. The processes involved in the nitrogen cycle are fixation, ammonification, nitrification, and denitrification.

            Fixation is the conversion of nitrogen gas to ammonia or nitrate. Primarily symbiotic bacteria, members of the genus Rhizobium, living in root nodules of legumes and some other plants, accomplish this. Some species of free-living bacteria and blue-green algae also nitrogen fixers. In this step of the nitrogen cycle, molecular nitrogen (N2) is split. The free N atoms then can combine with hydrogen to form ammonia (NH3). Another process, high-energy fixation of nitrogen by lightning in the atmosphere, results in nitrate (NO3) as the product. This process accounts for very small proportion on the total nitrogen fixed. (4)

            Ammonification is the process by which amino acids are broken down by decomposers to produce ammonia, Ammonium, or ammonia ion, is absorbed directly by plant roots. The plants use the ammonium to make amino acids, which are the basic units of proteins. The NH4 that is not absorbed by plants continues though the cycle.

            Nitrification is the process in which ammonia is oxidized to nitrate and nitrite. Nitromonas bacteria convert ammonia in soil to the nitrite ion (NO2), which is generally considered to be an intermediary nitrogen product. Nitrobacter bacteria quickly convert NO2 to nitrate (NO3). Oxygen is required for nitrification. Warm, moist, and well-aerated soil provides ideal conditions. (4)

            NO3 is either taken in by the soil microorganisms or plants, lost to leaching, or undergoes another microbial conversion-denitrification. The denitrifiers, fungi and the bacteria Pseudomonas, prefer an aerobic environment. However, if oxygen is limited, they use NO3 Instead of O2 and release N2 gas as a product. (8)

            Nitrification and denitrification both require a sufficient supply of organic matter, a limited supply of oxygen, a pH range of six to seven, and an optimum temperature of 60°C. (5)

            Human activity has greatly increased availability of this essential element of nature. This excess in nitrogen is disrupting ecosystems around the world. Nitrates from fertilizers seep into the ground and pollute the air. “Because industrial fertilizer is the biggest human contribution to the nitrogen cycle it is important that it be used as efficiently as possible. While economically more developing countries have become more efficient, applications in developing countries, are rising dramatically,” says Vitousek. (3)

            Temperature is one of the most important factors affecting rates of microbial growth. Temperatures, at which specific enzymes and cellular structures function, varies from one microbial species to another, depending on the specific chemical compositions specified by the genome on the organism. Within the growth range there will be an optimal growth temperature at which the highest rate of reproduction occurs. The minimum and maximum temperatures at which a microorganism can grow define the temperature growth range. Some microorganisms grow best at low temperatures. (1)

            The purpose of this project was to determine the effect of temperature on the processes of nitrification and denitrification. The research hypothesis was that as the temperature increases the rates of nitrification and denitrification increases. The null hypothesis was that the rate of nitrification and denitrification are not affected by changing temperatures.

PROCEDURE AND MATERIALS

            The electronic balance was used to mass 0.4 grams of ammonium chloride (NH4Cl), which was placed in a two-liter bottle and shaken to dissolve the NH4Cl. 700 grams of sand was massed and poured into a glass quart (946.97mL) jar. Then, 0.5 grams of garden soil was added to the jar to introduce bacteria. A graduated cylinder was used to measure 500 mL of the ammonium chloride solution and it was poured into the glass jar containing the sand and soil. A lid was placed on the jar and it was shaken to mix. The lid was removed after shaking. Eight more jars were prepared in the same way.

            Three jars were placed in a refrigerator, three jars were placed in an area incubated with a heat lamp, and three were left at room temperature. The outsides of the jars were covered to control the amount of light. The temperature of the contents in each jar was recorded daily. After the contents settled, a syringe was used to withdraw 0.5 mL of the solution from each jar, which was tested daily for the presence of nitrates using a nitrate test kit. The test was performed at approximately the same time each day. After testing the liquids, the lids were placed on the jars and they were shaken again to aerate. The lids were removed after shaking to provide an aerobic environment for nitrification.

            When the nitrate concentration reached 50 ppm (mg/L), five grams of sugar were added to the jars at room temperature and under the heat lamp to provide a ready source of energy for the denitrifying bacteria. The nitrate levels continued to be tested daily. The jars were not shaken and lids were placed on the jars to provide an anaerobic environment. The experiment ended when the nitrate concentration read less than 5 mg/L. The rates of nitrification and denitrification at the different temperatures were compared.

RESULTS AND DISCUSSION

            The results of the experimental setup showed the differences in nitrification and denitrification at different temperatures (see graph 1). The jars under the heat lamp (mean temperature of 28.94°C) took an average of 18 days to reach 50 mg/L of nitrate. The jars at room temperature (mean temperature of 19.65°C) took an average of 20 days to reach to reach the same nitrate concentration. Since the jars in the refrigerator (mean temperature 2.5°C) reached only 41 mg/L over the entire experiment period of 32 days, a comparison of the number of days to reach this level at the three temperatures was made (see graph 2). The jars at room temperature took an average of 15 days and the heat lamp setup needed an average of 16 days.

            When sugar was added and denitrification began the jars at room temperature reached 0-mg/L nitrate in an average of 13 days, while the jars at heat lamp needed 10 days (see graph 3). Sugar was not added to the refrigerated jars, as the nitrification rate was still being measured.

            An analysis of variance for replicated treatments was calculated to determine the significance of the variation in number of days to reach 41 mg/L of nitrate in the three jars of the three treatments. The variation was found to be significant at the p= 0.5 confidence level (see table 3)

CONCLUSION

            It was concluded there was a significant difference in the nitrification and denitrification at the different temperatures. The research hypothesis was accepted and the null hypothesis was reject. Further studies may involve varying the amount of sugar, or adding grass clippings or leaves to the jars to determine the effects on nitrification and denitrification.