Effect Of Organic Carbon On Nitrification Efficiency And Community Composition

Biological nitrification is the key process in the removal ofammonium from wastewater. The process of nitrification is sensitive toenvironmental factors such as pH, temperature, dissolved oxygen (DO),ammonium and nitrite concentrations, organic loading, and hydraulicloading rate. The impact of organic loading is significant because inthe presence of organic substrates nitrifying bacteria are usuallyoutcompeted by heterotrophic bacteria. This paper researches theshort-term and long-term effects of organic substrate on thenitrification.In the short-term effects of organic carbons on nitrificationexperiments, we inoculated 10% enriched nitrifying bacteria to 500mlmedium which NH₄⁺-N was 200mg/L with different C/N (0,0.5,1,2,4,8,16)at 30℃and 6mg/L DO. And then we monitor the nitrification by analyzingNH₄⁺-N, NO₂^--N, NO₃^--N and total N and research the shifts of communitystructure with increasing C/N by DGGE (Denaturing gradient gelelectrophoresis). The profile similarity and biodiversity werecalculatedbased on DGGE profiles. And the DGGE bands were recovery andsequenced to identify specific bacteria.In the long-term effects of organic carbons on nitrification experiments, the enriched nitrifying bacteria were formed a biofilm. Andthen they were cultivated in flasks with 150ml medium which NH₄⁺-N was200mg/L with different C/N at 30℃and 6mg/L DO. We analyzd NH₄⁺-N, NO₂^--N,NO₃^--N and total N and refreshed medium every 24h. After 3 months thenitrifying activity was analyzed. When the medium was refreshed sampleswere taken from the flasks periodically (2h) for 24h batch nitrificationkinetic analyses. And also we researchthe shifts of community structurewith increasing C/N by DGGE. The profile similarity and biodiversity werecalculated based on DGGE profiles. And the DGGE bands also were recoveryand sequenced to identify specific bacteria.In the short-term effects of organic carbons on nitrificationexperiments, the NH₄⁺-N was exhausted after 64h at C/N=0. But it is needmore and more time with theincrease of C/N. The NH₄⁺-N was not exhaustedafter 144h at C/N=8 and 16. It need for 32h to reach the maximumaccumulation of NO₂^--N at C/N=0, and the time increase with C/N increasing.It need for more than 144h to reach the maximum accumulation of NO₂^--Nat C/N=8. And there wasn't any NO₂^--N at C/N=16. rhe concentration of NO₃^--Nreach the maximum after 64h at C/N=0. It need for more time to completeit with the increasing C/N. In 144h the concentration of NO₃^--N didn'treach the maximum at C/N=4 and 8. Also there wasn't any NO₃^--N at C/N=16.Total N tests show that no significant nitrogen loss. The dominantbacteria were Nitrosomonas sp., Nitrobacter sp. and Flavobacteriaceae.But the community structure shift to Pseudomonas sp., Acinetobacterjunii and Pseudoxanthomonas byssovorax with increasing C/N. The clusteranalysis of total bacteria patterns indicating that the correlationcoefficient decreased with the increasing C/N. The total bacteriadiversity at C/N=1 was higher than at other C/N. No other significantdifferences were found at different C/N. The AOB and NOB diversity waslow and no significant differences among different C/N.In the long-term effects of organic carbons on nitrification experiments, the influentNH₄⁺-Nwas 200mg/L. The NH₄⁺-N removal rate wasdecreased 10%,20%,25% and 50% when organic carbons were added fromC/N=0.5 to 4. And it gradually recover as C/N=0, but it need more daysto complete it with increasing C/N. The NH₄⁺-N removal rate was decreased50% at C/N=8 and 16, and they weren't recovered. 24h kinetic comparisonshows that it is need more 0h, 4h, 4h, 6h, 8h, 8h to completed NH₄⁺-Noxidization from C/N=0.5 to 16. We detected that effluent NO₂^--N was 10% and 20% of influent NH₄⁺-N at C/N=1 and 2. But it recover as C/N=0after 20 days. 24h kinetic comparison indicates that a maximum nitriteaccumulation was reached at C/N=0 in 12h with a concentrationapproximately 80% of the initial ammonium concentration. It isnoteworthy that the appearance of maximum nitrite concentration atC/N=0.5, 1, 2 and 4 needed more and more time, and the maximum nitriteconcentration was lower than that at the C/N=0. There wasn't detectedany nitrite in effluent and NO₂^--N 24 kinetic analysis at C/N=8 and 16. Theeffluent NO₃^--N reduce to only about 60%,80%,70% and 20% of influentNH₄⁺-N at C/N=0.5,1,2,4. The 24h kinetic comparison shows that theappearance of maximum nitrate concentration needed more time than C/N=0.There was no nitrate in effluent and NO₃^--N 24 kinetic analysis at C/N=8and 16. The loss of nitrogen was about 35%, 20%, 25%, 60%, 60% and60% from C/N=0.5 to 16. It is indicated the denitrification appeared.Sequence analysis of 16S rRNA gene fragments revealed that the dominantpopulations shifted from nitrifying bacteria to denitrifying bacteriawith increase of C/N ratios. The dominant bacteria were Nitrosomonaseuropaea, Pseudoxanthomonas byssovorax and Nitrobacter sp. which mostlyare nitrifying bacteria at C/N=0 and the dominant bacteria werePseudomonas sp., Acinetobacter junii, Acidovorax sp. and Comamonas sp.which mostly are denitrifying bacteria. Although at high C/N ratios thedenitrifying bacteria was the dominant populations, nitrifying bacteriacan simultaneously grow with them. The nitrification process was changed to the simultaneous occurrence of two processes, nitrification anddenitrification. The total bacteria diversityat C/N=2 was higher thanat other C/N. No other significant differences were found at differentC/N. The AOB and NOB diversity was low and no significant differencesamong different C/N.