Nitrogen and carbon removal thanks to bacterial processes in a single stadium system ( PRiBioAC 3S ) in wastewaters with high organic and nitrose load

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Biologic procedure for nitrogen and carbon removal in a single step system (PriBioAC 3S) from zootechnical sewage with an high organic and nitrogen level.

A solution for nitrate problem

Paolo Broglio ( - Ecologia Applicata S.r.l.
Services for Ecology
Scientific Organization of Environmental Research

In agreement with the University of Milan

The nitrate directive enacted by EU highlighted the lack of a specific structure of the zootechnical sector in Italy, making clear to farmers and breeders the need to decrease the nitrogen level on open fields, following the parameters contained in the directive.

The main obstacle faced in this case, comes from the fact that many farmers don’t have specific fields for the purpose.

In this research is highlighted the upstanding efficiency of a new biologic process (PriBioAC 3S) which is able to drastically reduce more than the 80% of the overall nitrogen contained in cattle and swine sewage.

Moreover, this process also reduces around 80% the COD level, it can be registered as a patent and can be easily realized.


1.2  Analysis on the environment of the sector

The approval of a specific regulation enacted by the executive cabinet of the Lombardia region regarding action plan for agricultural nitrogen parameters established by law is worrying the whole zootechnic sector, especially in some specific areas (romagnolo/lombarda).

The executive cabinet of the Italian Confederation of Farmers (see CIA) of the Lombardia region, in joint session of15 November 2006,put a lot of pressure on all institutions located in the area in order to shaping a new prospective for the sector, avoiding the creation of a chain process that will lead inevitably to permanent closure.

Following the new territorial divisions, over 59% of the Lombardian plain is to be considered as “vulnerable” to the nitrogen problem, due to the fact that we are talking about 62% of the agricultural fields where are produced top quality foodstuff. (Grana Padana and Prosciutto di Parma)

The CIA emphasized that the nitrate problem needs to be solved overcoming the burocratic obstacles, widening competition and innovation of Lombardia agriculture and giving a dominant role to the sustainable development, that underline this new regional production system.

The actual rigid application of provisions and “collaterals” contained in the EU directive would mean a sure deep crisis of the agro-alimentary system as a whole.

The legislative decree issued by the Ministry of Agricultural Policies on 7th April 2006 (the so called “Nitrate Directive”) set out technical norms and principles of guidance for both vulnerable areas and not vulnerable ones.

This norm establishes a maximum nitrogen load on field of 170 Kg/ha for the former and 340 kg/ha for the latter, following also the specific loads based on the kind of breeding animals.

In the fatten up swine case, the limit goes from 112 to 110 Kg/t live weight, that corresponding respectively to 17 and 17,3 heads/ha, causing a reduction of heads/ha of around 3%, while in the fatten up bovines case the reduction scores 10% and 30% for milk producing bovines.

This decree represents the latest legislative act aimed at safeguarding the Lombardia plain from a nitrate pollution that is getting worse and worse every day.

Needless to say, Lombardia region has a national and European relevant role in this sector and the new regulation put in jeopardy its development and position, since top priority[1] seems to be reduction the nitrate loads on fields.

1.2. Description of the knowledge status

Until the discover of the high nitrate level in the water table of our plain, the nitrogen removal from zootechnical sewage was considered of scarce relevance; we know that the main cause for this type of pollution comes from the diffused practice of nitrogen fertilizing and fertigation.

The typical high N/C ratio of animal dejections made always troublesome inventing a simple and cheap treatment.

In several year different research Institutes, among which CNR and CRPA, published scientific literature[2], [3], [4], [5] in which were suggested many system to reduce nitrogen presence in the surface water bodies.

The disequilibria of nutrients makes harder the use of a biological depuration process, called “active mud” in an efficient way, (either it being single or double stadium) thus obliging farmers and breeder to dilute wastewaters using a lot of water, buying expensive treatments and fields not yet saturated or leaving the activity.

The high level of ammonia and organic nitrogen slows down the biological process, limiting the plant performance and the removal of nitrogen as gas N2 .

The mixing of civil with zootechnic wastewaters, strongly suggested, however represents an exceptional case with respect to normal practice, because breeding farms being far from cities and urban areas (an obvious reason for that: the bad smell that often comes from a farm of this kind)

Although, the path of a biological treatment and transformation of nitrogen had always being the recommended one, opposed to other processes considered expensive and scarcely monitor (combined treatments, chemical or physical treatments).

The nitrogen removal from zootechnic wastewaters follows a metabolic way that transforms organic nitrogen in ammonia (usually for hydrolysis or enzimically), then into nitrates (thanks to Nitrobacter and Nitrosomonas Spp.) and finally in N2gas.

The whole chemical reaction is displayed by the following scheme:

(N org.)  NH +4 +  O2 a NO -2 + H2O + H+  aNO - 3 + H2O + H+  a N2 + H2O

It appears clear the energetic waste needed to oxidize ammonia in nitrite and then nitrate in an oxidative environment; then reduced into gas nitrogen molecules, in an anaerobic environment.

Every step plans specific pools and suitable monitoring processes in order to obtain the best results, that might be in contrast with a good management of breeding farms.

The complexity of the nitro-denitro plant made quit many breeders from wastewater treatments, choosing the monitored spreading on fields: a system established by Regional regulation that cannot be enforce anymore because of the high level of nitrates in the water table, from where comes drinking water.

1.3. Other related researches

The excessive level of nitrogen in wastewaters isn’t a distinguishing feature of the zootechnic sector only: we also find it in the percolates, produced in the urban waste disposal site, and in the supernatant of the thickening sludges, produced in the biological depuration plant.

Already at the end of the 90’s, in order to reduce the size of the growing and growing problems, some researchers[6]  created a process for the treatment of high ammonia-load wastewaters, called SHARON.

This process, given particular conditions of Ph, oxygen and temperature, is able to produce only nitrites (around 50% of the ammonia input, avoiding the transformation of it in nitrates and saving 20%-30% of energy), then reduced to nitrogen gas in a second phase, as resumed in the chemical equation scheme:

NH4+ + HCO + 0.75 O2 → 0.5 NH4+ +0.5 NO2- + CO2 + 1.5 H2O.

This important innovation wasn’t able to propose a definitive solution, since it was based on two different states: aerobic and anaerobic. In 2002, a research team[7] built a process called CANON that was able to transform in a single stadium process ( single reactor) the ammonia into nitrogen gas, planning alternate anaerobic periods. Anyway, this process was relatively unstable and applications on larger scale were and are till now really a few, regarding for the most part the treatment of supernatants of thickening sludges. 

Those two last innovation we resumed up to now for nitrogen treatments through biological process have been verified in other researches[8], that already in 1997 highlighted the possibility to use particular bacterial stems to eliminate directly the ammonia in wastewaters as nitrogen gas, using a reactor completely anaerobic. This process was called Annamox and is characterized by the following equation:

NH4+ + NO2- → N2 + 2H2O.

Un efficient engineering plan activate the SHARON process before (weakly aerobic) and the Annomox after, (anaerobic) obtaining a complete removal of the wastewaters nitrogen and achieving energy saving for 25-30% with respect to the traditional nitro-denitro system. Needless to say, the reactors needed are two anyway.

The combination of both reactions can be easily represented as displayed by the scheme:      

95% N2                               

100 % N-NH4 → SHARON → 50% N-NH4 + 50% N-NO2 → ANAMMOX  → 5% N-NO3

The possibility to realize a single reactor able to ammonificate the organic nitrogen, to partially nitrificate and denitrificate in the case of heavy load, in terms both of carbon (20-40.000 mg/l of COD) and of overall nitrogen ( 3500-4000 mg/l Ntot), it’s extremely concrete after the publication of SNAP project[9] (Single-stage Nitrogen removal using Anammox and Partial nitritation) which demonstrate the possibility to have 60-80% nitrogen removal (with respect to the ammonic nitrogen input).

Anyway, in scientific literature were reported data about the relatively low level of ammonic nitrogen (500-600 mg/l maximum) and very low level for the SNAP process case: lower than 150 mg/l. For its intrinsic nature, the zootechnical wastewaters presents particular characteristics: an high COD level, an important nitrogen and ammonia load, and high level of solids in suspension.

[1]    Intervista con l’Assessore Regionale Viviana Beccalossi- Gestione deiezioni.

     L'Informatore Agrario. 1/2007

[2]    Piccinini, S. ,( 1990 )

     Olanda: Il Trattamento dei liquami zootecnici in impianti consortili.

      Rivista di Suinicoltura Ed. Edagricole

[3]     Bortone, G., Piccinini, S., ( 1991)

     Nitrification and denitrification in activated sludge plants for pigslurry and wastewater from cheese

      Bioresource Technology

[4]     Bonazzi, G.,Piccinini, S.,( 1997 )

     Come gestire i liquami suinicoli.

      CRPA. Rivista di Suinicoltura Ed. Edagricole       

[5]     Bonazzi G. ( 2007)

      Sempre più difficile gestire e smaltire le deiezioni

      L'Informatore Agrario. 1/2007

[6]    Hellinga,C., Schellen, A.A.J.c., Mulder,J.W., van Loosdrecht, M.C.M. and Heijnen,J.J. ( 1998)

     The SHARON process : an innovative method for nitrogen removal from ammonium-rich waste water.

      Water Science Technology, 37 (9), 135-142

[7]      Sliekers ,A.O., Derwort,N., Gomez,J.L.C., Strous, M., Kuenen, J.G. and Jetten, M.S.M. ( 2002)

     Completly autotrophic nitrogen removal over nitrite in one single reactor.

      Water Research, 36, 2475-2482.


       Strous M, Van Gerven E, Kuenen JG& Jetten M (1997)

      Effects of aerobic and microaerobic conditions on anaerobic ammonium-oxidizing (Anammox) sludge. Appl.Environ.Microbiol. 63: 2446-2448.


[9]    Furukava,K.,Lieu, P.K., Tokitoh,H. and Fujii,T. ( 2006 )

      Development of single-stage nitrogen removal using anammox and partial nitritation ( SNAP ) and its treatment performance. Water & Science Technology. 53 (6), 83-90.


The inevitable breaking of fixed standard levels of nitrogen established by law in water coming from the water table located in agricultural and breeding areas of the Italian territory, with particular reference to the Lombardia plain lead to the approval of a ministerial decree on April 7th 2006 ( called “Nitrates directive”).

This regulation establishes criteria and general technical norms for breeding water streams, to be respected by the regions that host either vulnerable or non vulnerable areas.

Fist of all, it fixed maximum level for nitrogen on field (170 Kg/ha in a vulnerable area and 340 Kg/ha in a ordinary one) and, consistently with the zootechnical species, the specific loads (living weight).

The procedure we presents here achieve a reduction of nitrogen and its subproducts on field, leading to an improvement of the drinking water that comes from the underneath water table.

It’s then almost trivial the need to prevent wastewaters dumps from zootechnical installations.

The concrete possibility of building a process on large scale, cheap and easily managed, able to drastically reduce the overall nitrogen up to 80-90%, it’s the perfect solution to increase wastewater quality and then, indirectly, the pollution of the water table underneath (that gave the jump-start to the “nitrates chapter”).

2.1 Objectives of applied research

The research target it’s the improvement of the knowledge related to the process, that at its actual status it is not supported by enough scientific references to be replicated on larger scale, even in combined forms(different nozzles, different size of reactor, different diffusion system, ecc). Then, our objective is to verify the possible applications of the SNAP process, modified in a such way to obtain zootechnical wastewater that has the following characteristics ( schedule 1):

Input T.Q.          Separated input (filtered)       

COD (mg/l)               63.891                60.930                                        

N Tot. (mg/l)               4.952                   4.702                                          

N- NH4 (mg/l)             2.911                   2.900                                          

N-NO3 (mg/l)           61                            57

N-NO2 (mg/l)             n.r.                         n.r.     

The process undergoing an adequate period of seeding (30÷60 days) should assure the removal of total Nitrogen around 80%.

The main purpose of the applied research is focused on the Codification of Biological Nitrogen Removal for 80-90% of the zootechnical waste water.

It should be realized over a true scale or at least over a semi-true scale and consequently demonstrate its technological reliability and its reproducibility.

The best practical result of this project is shown in the possibility to overpass the obstacles, hidden in the technical law on Nitrates Pollution Control Directive (91/676/EEC of 12 December 1991).

Consequently it should help to by-pass the Directive limitations, which could force the farmers in peculiar situations to close the cattle-breeding farm.

2.2 Results

The possibility to apply the modified SNAP process has successfully been tested both in laboratory and partially also in the practice.
The experiments have been executed in anoxic alternate condition for sufficient period and the process resulted considerably stable.
Analyses have shown a total Nitrogen Removal of about 80%, measuring a minimum nitrite concentration (never recorded before and indicating a bad functioning in the traditional plant) and obtaining a substantial Nitrate absence[1].

On the basis of the obtained results, an applied research took form, already ratified in main lines as process, which brought to codify on industrial or real scale a new type of process called :PRiBioAC 3S (Biological Removal Process over Nitrogen and Carbon through a Single Stage System)

The proposed project for an applied research could be realized by setting up a pilot plant having capacity of few net cubic meters or by revamping an existing sewage tank (around 450 liters volume), transforming it in a biologic reactor, placed in a cattle-breeding farm that supports the project.

Some air baffle diffusers should be settled at the bottom of the tank enabling to regulate the blowing air from minimum to maximum, as it was done in the biologic reactor OXANOX.

There should also exist the possibility to stop at fixed interval the airflow and somehow to keep the sewage in mixing and suspension.

The system will also be provided with a partial regulation of pH, using CO2 as acidifying agent; moreover, oxymeters and pHmeters will collect additional data during tests.

[1]           Stefano Franco, Dario Pagani – GB. Odobez Srl, Varese; Paolo Broglio – Ecologia Applicata, Milano

               “Applicazioni al trattamento di reflui industriali e di reflui zootecnici del processo di Bioflottazione®:

                  abbattimento di tensioattivi, coloranti diretti ed alti carichi di azoto”. Ecomondo. Rimini- 9 novembre 2006

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