substrate and product inhibition of nitrification
TRANSCRIPT
Substrate and Product Inhibition of Nitrification
J. BUDAY, M. DRTIL, M. HUTŇAN, and J. DERCO
Department of Environmental Science, Faculty of Chemical Technology, Slovak University of Technology, SK-812 37 Bratislava
Received 14 June 1999
Great effort is paid to the development of new technologies, that enable the short-cut of the nitrification-denitrification cycle avoiding the oxidation of NO J to N0^" (nitrite route). The nitrification of NH4" only to NO2 and its reduction to gaseous N2 offers several advantages: lower oxygen demand for nitrification, lower demand of organic matter for the denitrification of N0^", higher denitrification rates of N0^
The aim of this work was to obtain more detailed information about the inhibitive forms (dissociated or nondissociated) of the substrate/product as well as about the concentrations that cause inhibition. The substrate—product inhibition was tested in batch tests. Each experiment at a certain pH was carried out using a set of six reactors. One of them served as a reference, while the five others contained different concentrations of the tested compounds. Five sludges from different municipal wastewater treatment plants were used in the experiments. The results obtained from batch inhibition tests carried out on different sludges are briefly summarized in this work. The very similar behaviour of different sludges is worth to note.
One of the most common technologies to remove nitrogen from wastewaters is biological nitrification-denitrification. Nitrification is a process, in which NH4" is gradually oxidized to N0^" (nitritation) and subsequently to N0^~ (nitratation) by nitrifying microorganisms. The produced N0^" are then, under anoxic conditions, reduced to gaseous N2 by heterotrophic microorganisms, utilizing the organic matter content of the wastewater. Regardless of some inhibitory influences, that can make difficult the use of these biological processes, even the lack of organic matter for denitrification can often have a negative impact on the overall nitrogen removal efficiency. In most of the cases, additional dosage of biodegradable organic matter (methanol, acetic acid, organic wastes, etc.) is used to resolve these problems. However, this additional dosage can increase the costs of the treatment process. Thus great effort is paid to the development of new technologies, that enable the short-cut of the nitrification-denitrification cycle avoiding the oxidation of N0^~ to N 0 ^ (the so-called nitrite route). The nitrification of N H j only to N0^" and its reduction to gaseous N 2 offers several advantages
- lower oxygen demand for nitrification (only 75 % of the amount necessary for complete nitrification to NO3-),
- lower demand of organic matter for the denitrification of N 0 ^ (only 60 % of the demand necessary to the denitrification of N 0 ^ ) ,
- 40 % higher denitrification rates of N0^~ [1],
- lower biomass yield during anoxic growth [2], - possible adaptation of heterotrophic microorgan
isms to high concentrations of N0^" (up to p(N0^"— N) = 2000 mg d m " 3 at pH 8—8.5) [1].
An interesting possibility to achieve only partial nitrification of N H j to N0^", is the SHARON process [3]. It is based on the kinetic selection of ammonium oxidizers in a system without sludge retention. Another possibility to perform the nitrite route is based on the different sensibility of nitrifiers to the substrate—product inhibition [4], where nitrite oxidizers should be selectively more inhibited at lower NH3 concentrations than ammonium oxidizers. Also, temperatures higher than about 25 °C are reported to favour nitrite accumulation [5].
The aim of this work was to obtain more detailed information about the inhibitive forms (dissociated or nondissociated) of the substrate/product as well as about the concentrations that cause inhibition. The obtained results should be utilized during the following research directed towards the short-cut of nitrification-denitrification. Despite of possible troubles caused by adaptation of the nitrite oxidizers to the substrate—product inhibition [2, 6], there could be cases, where specific conditions (high NH4" and N 0 ^ content of the wastewater connected with high or low pH, high salinity, specific inhibitors strengthening the substrate—product inhibition) can enable the long-term performance of the nitrite route.
Chem. Papers 53 (6) 379—383 (1999) 379
J. BUDAY, M. DRTIL, M. HUTNAN, J. DERCO
E X P E R I M E N T A L
The substrate—product inhibition (in sequel as inhibition) was tested in batch tests lasting for 1—1.5 h. Each experiment at a certain pH was carried out using a set of six reactors (V— 200 cm3). One of them served as a reference, while the five others contained different concentrations of the tested compounds. The required concentrations of PO^", NHj , NO J , and N 0 ^ were achieved by addition of K 2 HP0 4 , (NH4)2S04, NaNCb, and NaNOß solutions. The pH was continuously controlled and maintained by addition of KOH and H2SO4 solutions. Five different sludges were used in the experiments:
- sludge A: a sludge coming from a nitrifying municipal wastewater treatment plant (MWWTP) with pure oxygen aeration (ca. 500 000 inhabitants),
- sludge B: a sludge coming from a MWWTP with nitrification, denitrification, and phosphorus removal in side stream (ca. 200 000 inhabitants),
- sludge C: a sludge coming from a nitrifying-denitrifying lab-scale sequencing batch reactor,
- sludge D: a sludge coming from a nitrifying MWWTP (ca. 150 000 inhabitants),
- in Fig. 2 even some results from [7] are plotted for comparison. These results were obtained with a sludge (sludge E) coming from a nitrifying MWWTP
(ca. 350 000 inhabitants). The concentrations of PO^"—P and dissolved oxy
gen in the reactors were 3—5 mg dm"3 and 4—7 mg dm - 3 , respectively. The concentrations of the activated sludge were in the range of 1—5 g dm"3 .
All analyses were carried out according to the Standard Methods [8], except the determination of NO^~, which was carried out using the Zambelli method [9] (a colorimetric method with sulfanilic acid and phenol).
RESULTS A N D DISCUSSION
The concentrations of nondissociated NH3, HNO2, and HNO3 were calculated on the basis of acid-base balances [4, 10]. The inhibition was evaluated using the equations of noncompetitive inhibition
1 =
/ =
P + Ki
P-Po P-P0 + Щ
(1)
(2)
where: J - inhibition, Ki, K\ - inhibition constant (concentration causing an inhibition of 50 %), po -concentration over which inhibition starts to demonstrate itself, p - concentration.
100
I 80 p В 60 Б £ 40 о
I 20
-20
100
^ 80 о S 60 S £ 40 о о 20
1 ° -20
-
:
ГГ 1 1—' i i i i • |
coef.ofcor=0.4 П
D
% * • • • Li 1 1 __I 1 1 1 1 1 1 1 I I
-
10 100 ^(NH4
+-N)/(mgdm-3) 1000
coef. of cor =0.45
•í jŕ 4^ô*
uJ 1 1 I I I M i l 1—I I I I l l l l 1 I M i l l
0.1 1 10 100 p(NH4
+-N)/(mgdm-3) 1000
1 10 100 p(NH3)/(mgdm-3)
T 1—i i I
coef. of cor =0.91
0.01 0.1 1 10 píNHaVímgdm"3)
100
с о '£3
ÍS в с о с о •-3 Ъ
о
S
с о
Fig. 1. Inhibition of nitrification by NHj—N/NH3 (inhibition of nitritation measured using sludge D, • pH = 8.9, • pH A pH = 7.5, • pH = 6.8; inhibition of nitratation measured using sludges • A, A B, I C; / = p/(p + K{ I = (p-po)/(p-po + KD).
= 8.2,
380 Chem. Papers 53(6)379—383 (1999)
SUBSTRATE AND PRODUCT INHIBITION OF NITRIFICATION
100
í 80 с p Jä 60 i •s 4 0
c o
.c c
20
0.1
i 1 — Г - Г Т
-
-
-Т-ГТТТ 1—1 1 1 M I H r
coef. of cor =0.41
•
' ' i i ' 111A • '—-"Л
" * * ! • • A •
A A A A
i и nl i i i i mil—Aj._T.t_i i iul i i i i n u
1 10 100
p(N02*-N)/(mgdm'3)
coef.of cor =0.77
1000 0.000001 0.0001 0.01 p(HN02)/(mgdm-3)
c o
J5 1c c
100
10 100 /HNO^-N^mg dm'3)
coef. ofcor.=0.55
•
"
4 A
A A
. T^I i ^ . i i c • i - i - t i i u — i
A •
A
•
•
A •
A
1 M I H
•
•
A
A
1000 0.0001 0.001 0.01 0.1
p(HN0 2 )/(mgdm" 3 )
100
80
60
40
20
0
1
c o
s 2 c •s c g 3 c
Fig . 2. Inhibition of nitrification by N O ^ — N / H N 0 2 (A sludge A, • sludge В, О sludge E; J = p/(p + Ki), (p-po)/(p-po + K<)).
I =
Eqn (2) was used in cases, where there was clearly found that up to a certain concentration no inhibition occurred (inhibition occurred only at concentrations exceeding p 0 - see Fig. 1). The inhibition was calculated using the following equation
Tret - r
1 = Tref
100% (3)
where: J - inhibition, r - nitritation/nitratation rate, rref - nitritation/nitratation rate in the reference reactor.
Correlation coefficients were calculated according to [11] as
coef. of cor. = • пТ,хУ~Т,хТ,У
y/[nZ*2 - (£*) 2]>Ľ2/ 2 - Q » 2 ] (4)
where: n - number of measurements, x - concentration of the inhibitive substance, and у - inhibition.
Inhibit ion of Nitrification by N H + / N H 3
The tested ranges of pH and NH4"—N concentrations were 6.8—9 and 0—250 mg d m - 3 , respectively. The reference reactors contained at the beginning of the experiments píNH^"—N) = 1 mg dm"3
and p(NO^~—N) = 15 mg d m - 3 (reference reactor for the nitratation), and p(NH|—N) = 13 mg d m - 3 (reference reactor for the nitritation). The average concentration of the examined substance in the reference reactor is signed with a dashed line (in all of the figures). It is obvious from Fig. 1 (the horizontal axes of all of the figures are in logarithmic scale) that the inhibition of nitritation and nitratation was caused by the nondissociated form, the NH3.
This becomes evident, as soon as the NH4"—N concentrations are converted to the NH3 ones (see the coefficient of correlation). As the inhibition started only over a certain concentration (So), both equations of noncompetitive inhibition were utilized to describe it (both equations were utilized even for the HNO2 and HNO3 inhibition). The very similar behaviour of three different sludges is worth to note.
Inhibi t ion of Nitrification by N O 2 / H N O 2
The tested ranges of pH and NO-7—N concentrations were 6.8—8.9 and 0—415 mg dm"3 , respectively. The reference reactors contained at the beginning of the experiments ^(NHj—N) = 15 mg dm"3
and p(NO^~—N) = 15 mg dm"3 (reference reactor for the nitratation), and ^(NHj—N) = 15 mg dm"3 (ref-
Chem. Papers 53 (6) 379—383 (1999) 381
J. BUDAY, M. DRTIL, M. HUTNAN, J. DERCO
10 100
p(N03--N)/(mgdm-
3)
-
-
coef. ofcor.=0.98
A
У
• * * 1 l l t l l l l L J__l 1 1 MIH 1 1 1 1 H i l l 1
-
1000 1-10"* 1-10'7
1-10"* 1-10"
p(HN0
3)/(mgdnT
3)
60
50
40
30
20
10
0
-10
e: c o
4 3
B 'Б c
o c o
43 ja
л c
1-10̂
80
60
£ 40 o c o a 20
10 100
p(N03--N)/(mgdm-3)
1000 MO-1 1-10'7 1-1 o 4
p(HN03)/(mgdm-3)
-
-
• " •
•
•
11 111. .
• coef. ofcor.=0.40
á,
Á
Á
Á
80
60
- 40
20
1-10"£
c o '43
B
o c o 43
Fig . 3 . Inhibition of nitrification by N 0 3 —N/HNO3 (inhibition measured using sludge D, • pH = 8.5, • pH = 7.7, • pH / = p/(p + Äi), / = (p - Po)/(p - Po + K<)).
6.8;
Tab le 1. Forms and Concentrations of NHJ /NH3 , NO^"/HN0 2 , and N O ^ / H N 0 3 Causing Inhibition of Nitrification
Form
NH+ NH3
Nitritation no yes Nitratation no yes
causing inhibition
NO" no yes
Concentration
Nitritation beginning of the inhibition (po/(mg d m - 3 ) ) inhibition of 50 % (Ki - eqn ( I ) ) / (mg d m " 3 ) inhibition of 50 % {K[ - eqn (2))/(mg d m " 3 )
Nitratation beginning of the inhibition (po/{mg d m - 3 ) ) inhibition of 50 % {Ki - eqn (I)) / (mg d m " 3 ) inhibition of 50 % (К*. - eqn (2))/(mg d m " 3 )
causing
N H 3
1 46 30
1 20 16
HNO2 yes no
inhibition
N O " — N
198
N O " no yes
HNO2
1 x Ю - 4
0.0109 0.0105
N O ~ — N
205
H N 0 3
yes no
H N 0 3
8 x 1 0 " 8
7 x 1 0 " 6
6 x I Q " 6
Note: In cases of inhibition by dissociated forms it was not possible to determine po from the obtained results.
erence reactor for the nitritation). As it is obvious from Fig. 2, the inhibition of nitritation was caused by the nondissociated HNO2. The behaviour of different sludges, like previously in the case of NH3 inhibition, was again quite uniform. Contrary to the commonly
accepted theory of HNO2 inhibition of nitratation, the dissociated NO^" was found to cause the inhibition of nitrite oxidizers. The inhibitive effect of a certain concentration of NO^~ was not significantly strengthened by lowering the pH.
382 Chem. Papers 53 (6) 379—383 (1999)
SUBSTRATE AND PRODUCT INHIBITION OF NITRIFICATION
I n h i b i t i o n of N i t r i f i c a t i o n b y N O 3 / H N O 3 R E F E R E N C E S
The tested ranges of pH and N0^~—N concentrations were 6.8—8.5 and 0—400 mg d m - 3 , respectively. The reference reactors contained at the beginning of the experiments p ( N H j — N ) = 1 mg d m - 3
and p(NO^~—N) = 15 mg d m - 3 (reference reactor for the ni t ratat ion), and ^ ( N H j — N ) = 15 mg d m - 3 (reference reactor for the nitr i tat ion). The nondissociated HNO3 was found to cause the inhibition of nitritation (see Fig. 3), while the ni tratat ion was inhibited by the dissociated NO^~
C O N C L U S I O N
The nondissociated N H 3 , H N 0 2 , and H N 0 3 were found as inhibitive forms for the nitritation. The nondissociated HNO2 and HNO3 were expected to inhibit the ni t ratat ion [4], but on the basis of our results the nondissociated NH3 and the dissociated NO^~ and N0^" were found t o inhibit the nitratat ion. Inhibitive concentrations of substrate/product, as well as the inhibition constants are summarized in Table 1. Comparing the inhibition constants, nitratat ion seems to be more sensitive to NH3 inhibition than nitritation. All these experiments were carried out as batch tests, so the behaviour of the nitrifying microorganisms in long-term experiments can differ due to acclimatization. The subst ra te—product inhibition will be studied further in long-term experiments.
1. Abeling, U. and Seyfried, C. F., Water Sei. TechnoL 26, 1007 (1992).
2. Turk, O. and Mavinic, D. S., J.W.P.C.F. 61, 1440 (1989).
3. Jetten, M. S. M., Horn, S. J., and van Loosdrecht, M. C. M., Water Sei. TechnoL 35, 171 (1992).
4. Anthonisen, A. C , Loehr, R. C , Prakasam, T. B. S., and Srinath, E. G., J.W.P.C.F. 48, 835 (1976).
5. Beier, M., Hippen, A., Seyfried, K. H., Rosenwinkel, K. H., and Johansson, P., Eur. Water Manag. 2, 61 (1999).
6. Turk, O. and Mavinic, D. S., Water Res. 23, 1383 (1989).
7. Ondrejčeková, M., Diploma Thesis. Slovak University of Technology, Bratislava, 1997.
8. Standard Methods for the Examination of Water and Wastewater, 17th Edition. APHA, Washington D.C., 1989.
9. Metodi analitici per le acque, 1—2. Consiglio Nazionale delle Ricerche, Istituto di Ricerca sulle Acque, Roma, 1972.
10. Pitter, P., Hydrochemical Tables. Nakladatelství technické literatury (Publishers of Technical Literature), Prague, 1987.
11. Tuček, F., Holata, L, and Eckschlager, К., Výpočetní technika pro technologii vody a prostředí. (Computer Technique for the Technology of Water and Environment.) Nakladatelství technické literatury (Publishers of Technical Literature), Prague, 1983.
Chem. Papers 53 (6) 379—383 (1999) 383