Diffusional transport mechanisms and biofilm nitrification characteristics influencing nitrite levels in nitrifying trickling filter effluents

~

Wat. Res. Vol. 29, No. 10, pp. 2287-2292. 1995

Pergamon

0043-1354(95)00061-5

Copyright ( 1995 ElsevierScience Ltd Printed in Great Britain. All rights reserved 0043-1354/95 $9.50 + 0.00

D I F F U S I O N A L T R A N S P O R T M E C H A N I S M S A N D BIOFILM NITRIFICATION CHARACTERISTICS INFLUENCING N I T R I T E LEVELS IN N I T R I F Y I N G T R I C K L I N G F I L T E R EFFLUENTS M. N I J H O F ~ and A. K L A P W I J K 2@ ~Netherlands Institute of Fisheries Research (RIVO-DLO), Haringkade 1, P.O. Box 68, NL 1970 AB IJmuiden, The Netherlands (correspondence address), Department of Fish Culture and Fisheries, Agricultural University Wageningen (LUW), P.O. Box 338, 6700 AH Wageningen, The Netherlands and 2Department of Environmental Technology, Agricultural University Wageningen (LUW), P.O. Box 8129, 6700 EV Wageningen, The Netherlands (First received January 1994; accepted in reHsed form Februa O' 1995)

Abstract--The widely reported accumulation of nitrite during nitrification is studied in recirculating fish culture systems. Nitrite oxidation capacity was homogeneously distributed over the trickling filter, as opposed to the ammonia oxidation capacity, which decreased at an increasing filter depth. Under all circumstances tested, nitrite accumulation in the system was restricted to an elevated but stable nitrite concentration, indicating complete oxidation of the produced nitrite. For a specific trickling filter, a fixed ratio between nitrite and ammonia concentrations in the recirculating water was found. Nitrifying biofilms maintaining relatively low nitrite concentrations were characterized by a relatively high nitrite oxidizing capacity compared to ammonia oxidizing capacity, whereas biofilms with a relative low nitrite oxidation capacity induced high nitrite concentrations. The occurrence of high nitrite concentrations in trickling filter effluents can entirely be explained by diffusional transport mechanisms in combination with the characteristics of the biofilm. There is no argument that high levels of nitrite are caused by insufficient activity or inhibition of the nitrite oxidation process. Key words--nitrification, nitrite accumulation, biofilms, fish culture, recirculating systems

INTRODUCTION Nitrifying biofilters are often applied in recirculating fish culture systems in order to minimize the high water requirements. In these systems, toxic ammonia is converted into far less harmful nitrate, of which relatively high concentrations can be maintained in the water, drastically diminishing the need of water renewal. However, concentrations of the intermediate product nitrite, which is toxic for fish, are often reported to exceed the ammonia concentrations (Otte and Rosenthal, 1979; Poxton et al., 1981: Bovendeur et al., 1987; van Rijn and Rivera, 1990), which can even impose restrictions on the application of nitrification in recirculating fish culture systems. The phenomenon of nitrite accumulation in nitrifying systems is often attributed to inhibiting factors, demonstrated by the temporary nitrite build-up in batch wise nitrification experiments (Balmelle et al., 1992; Yang and Alleman, 1992) or by the nitrite accumulation in continuous experiments (Anthonisen et al., 1976). However, circumstances in recirculating systems are far from the reported conditions in which inhibition can be expected. Moreover, considerable confusion arises from the use

of terms such as '~accumulation" or "build-up", without indicating whether a continuing concentration rise due to a net nitrite production is meant, or just a high equilibrium concentration. High concentrations of nitrite in recirculating systems might also be explained by temporal and/or spatial distribution of the ammonia oxidation (nitritification) and nitrite oxidation (nitratification), in combination with aspects regarding diffusional transport mechanisms (van Rijn and Rivera, 1990). Characterization of nitrifying biofilms by comparison of both nitritification and nitratification is introduced by Bovendeur (1989) (Fig. 1). In that study, it is demonstrated that the nitratification capacity develops with some delay after the development of nitritification, but will eventually exceed the nitritification capacity. It was correctly emphasized that the measured nitratification rates in those studies refer to removal rates from bulk fluid only, whereas during nitrification, nitrite is produced inside the biofilm itself. Nevertheless, regarding the balance between oxidation capacities of both nitrification steps, it was believed that a net nitrite production during nitrification would never occur when applying biofilms that have reached constant

2287

2288

M. Nijhof and A. Klapwijk

capacity ("full grown biofilms"). This was demonstrated by conducting biofilm monitoring experiments following Nijhof and Bovendeur (1990) in which the biofilm was continuously loaded with ammonia at several fixed loading rates, and a constant low nitrite concentration was observed (Fig. 2). Even at the highest ammonia loading rate of 0.66 g NH4-N m -' d t, where m a x i m u m nitritification was achieved judging by the net increase in ammonia concentration, the nitrite concentration remained low throughout the experiment. The reported nitrite "accumulations" in recirculating fish culture systems are probably elevated but nitrite concentrations are constant, which indicates equal rates of nitritification (Rym) and nitratification (R~o:). R~,~ = R~o~

(1)

It has been demonstrated that the half-order model for substrate elimination in biofilms (Harremoes, 1982) can be applied for trickling filters in fish culture recirculating systems at low substrate concentrations and air saturated dissolved oxygen contents (Bovendeur et al., 1987). The ammonia and nitrite removal rates can be described, as a function of the substrate concentrations, respectively as:

R~,., -- a~/Snm

(g N H 4 - N

m :d

1)

(2)

R~o_, = b./S~,o:

(g N O ~ - N

m -' d ')

(3)

If Rxm = Ryo,, then:

and:

SnH~ =

b

(4)

From equation {4) it follows that a fixed ratio in ammonia and nitrite concentration can be expected, depending on the ratio of the half-order removal constants. The characteristics of the biofilm in combination with the biofilm loading are reflected in the steady-state ammonia and nitrite concentrations in the recirculating water, which also implies that the nitrite concentration does not necessarily have to be low. The objective of this study is to assess whether the nitritification and nitratification rates are indeed equal under conditions found in recirculating fish culture systems. Subsequently, it is studied whether the level of the nitrite concentration can be explained by the biofilm characteristics of the nitrifying trickling filter. MATERIALS AND METHODS

MateriaL~ The study was conducted with a pilot scale recirculating fish culture system for the culture of eel (Anguilla anguilla) and with a system for the culture of turbot (Scophthahnus maximus). The eel culture system and its trickling filter are

Developing biofilm

F u l l - g r o w n biofilm ~

NO2 -N /

'= E ~ z ~

~ Z

Concentration (g N m -3)

NH4+-N

Concentration (g N m 3)

Fig. 1. Schematic representation of the development of nitritification (solid line) and nitratification (dashed line) in nitrifying biofilms (redrawn from Bovendeur, 1989)

described in detail by Nijhof (1995a). The basically similar fish culture recirculating system for the culture of turbot is described in Nijhof (1995b). Characteristics of nitritification and nitratification were measured in biofilm monitoring experiments as described in Nijhof and Bovendeur (1990). Elevated ammonia influent concentrations were obtained by adding an ammonia solution (domestic use grade, 70 x 10~g m ~ NH4-N) to the system by a peristaltic dosage pump. Total ammonia concentration (NH~ + NH~, throughout this paper expressed in nitrogen, indicated as NH4-N) was measured by the indophenol-blue method (S61orzano, 1969). Nitrite was measured by a method from Bendschneider and Robinson (1952). Dissolved oxygen and pH were measured with probes (WTW. Germany).

Evperimental set-up and methocLs" The nitritification and nitratification characteristics as a function of filter depth were measured for the trickling filter from the pilot scale eel culture system. From four equidistant depths in the 2.5 m high trickling filter, special biofilm samples were taken out from the filter for measurement. From the turbot culture system, only biofilm material from the top of the filter bed was tested, as this system was not equiped with a filter suitable for stratification measurements. In the first experiment with the complete tilter of the eel culture system, effects of varying hydraulic loading rate and influent ammonia concentration were studied. Three hydraulic loading rates were applied of, respectively, 75. 150 and 300 m ' m : d L Within each hydraulic loading rate, the effect of ammonia influent concentration was studied in the range of approx. 0.5-5 g NH~-N m ~. Water samples were S

E

6

0.6(~ g m-2 d 1 .

}

2

-

Z

•~.

4

--

NHa-N

0.52 g m-2d I

~) Z

NH4-N ~ 1 0.38 g m - d0 0

I 30

NO2-N , 60

I 90

I 120

Time (min) Fig. 2. Ammonia and nitrite concentrations of a full-grown biofilm sample during nitrification at three rates of ammonia supply (Nijhof, unpublished)

Nitrite accumulation during nitrification ,.-., ~'

¢7

..-.

1.2-

1.2

zi.n. 1.0 --

A

?

O

O

E 0.15 m

O

:o/5 oo

z

7 i

0.8

1.0 I

Z

0.8

0.88 m

7 z

0.6

./z%-=7"

0.4

IIzO A A

1.62 m

.-w;-Tz-.~ "-

z

~ •

•

0.6

d

2.35 m

"~

2289

~

AA

0,4

> o

E

'~ 0.2 Y_ E E

E

0 0

<

1

1

I

I

2

4

6

8

I 10

Ammonia concentration (g NH4-N m -3)

0.2

0

z

II

I

I

I

I

I

2

4

6

8

10

Nitrite concentration (g NO2--N m -3)

Fig. 3. Biofilm nitrification characteristics of biofilm samples taken from four depths from the trickling filter in the eel culture system. Left: nitritification rate capacity; values represent depth (in m) measured from the top of the filter bed. Right: nitratification rates of same biofilm samples. O: 0.15 m; O: 0.88 m; A: 1.62 m; A: 2.35 m.

taken at six sampling outlets at various filter depths, and were analysed for both ammonia and nitrite concentration. Dissolved oxygen and pH were monitored at all sample outlets to assure that these conditions where not limiting the nitrification. In order to study some aspects found in the first experiment in more detail, a second experiment was conducted in which ammonia supply to the system was increased daily after obtaining steady-state ammonia and nitrite concentrations, checked by regular water sampling. In this experiment, the hydraulic loading rate was 150 m 3 m - -' d t. This experiment was performed in both the eel culture system and the turbot culture system.

RESULTS

In c o n t r a s t to the p r o n o u n c e d vertical stratification f o u n d in a m m o n i a oxidation capacity (Nijhof, 1995a), the nitrite oxidation capacity appeared to be very h o m o g e n e o u s l y distributed over the filter c o l u m n (Fig. 3). In addition, the nitrite oxidation capacity 1.4 - o

"

12-

NO2-N ~

-

7 z

1.0

-

o.8 .~ 0.6

-/

.,/,of - ' -

;

0.4 E

4

P 0.2 z 0 i~"

I

I

I

2

4

6

Concentration (g

N

I 8

m -3)

Fig. 4. Nitritification and nitratification characteristics of the biofilm from the upper part of the trickling flter in the turbot culture system.

a p p e a r e d to be lower t h a n the a m m o n i a oxidation capacity. Figure 4 shows the nitrification characteristics o f the biofilm in the u p p e r part of the filter from the t u r b o t recirculating system. The characteristics differ from that o f the eel culture system (Fig. 3) by having a nitrite oxidation capacity exceeding the a m m o n i a oxidation capacity. D u r i n g the first experiment with the eel culture system, nitrite c o n c e n t r a t i o n s fluctuated slightly during the passage t r o u g h the filter bed, but no systematic increase or decrease, caused by c h a n g i n g inlet a m m o n i a c o n c e n t r a t i o n or hydraulic loading rate, could be detected (Fig. 5). In addition, nitrite c o n c e n t r a t i o n s where higher as the inlet a m m o n i a c o n c e n t r a t i o n s were higher. In the second experiment, the daily increase in a m m o n i a supply caused a gradual increase in b o t h a m m o n i a a n d nitrite c o n c e n t r a t i o n s over the l0 day period (Fig. 6). D u r i n g the day, however, the c o n c e n t r a t i o n s o f b o t h a m m o n i a a n d nitrite decreased slightly due to the decrease in a m m o n i a excretion by the fish in the morning. The increased a m m o n i a concentration, and hence increased nitrification rate, never caused a steady increase in nitrite c o n c e n t r a t i o n during the day due to the lack in nitratification capacity. It appears, therefore, that the " a c c u m u l a t i o n " of nitrite is only manifested as a n elevated nitrite concentration, and not as continuing rise due to a relatively insufficient nitratification rate. The nitrite c o n c e n t r a t i o n again appeared to be p r o p o r t i o n a l to the a m m o n i a concentration. Figure 7 shows the relation between the inlet a m m o n i a c o n c e n t r a t i o n a n d the inlet nitrite c o n c e n t r a t i o n of the eel culture system a n d the t u r b o t culture system during the second experiments.

M. Nijhof and A. Klapwijk

2290 H=75 m d -I

7

z i

6

ff z

--

..._:

:--:--:--1

6.64

5

cm "-" 4

g

'~

H=150 m d -z

--

6.51

--

7 o [ - - / o ~ o , . ~ o ~ O ~ o

6

--

6

5 --

-

4'

3

.

1 t 50

"~ 0 z

t 100

I 150

I 200

-o~o

-'~'"~ • ~ ° ~ ° , ~.----,-'- • - - o - - o ~ o

3 ,~J

~ 2 /o..~°~O~o~o

H=300 m d -r

7

• ~ • ~ o /

2.24 •

4.10

•

3.08

4.17

54~ _ / • - ' - - - - , ~ o . . _ . _ . o ~ o

3.17

? . ~

3[

•

- /

o........0~• / ~o

2.23

2.32

~o.-~o/

5.45

2 --

2 ~-

0.88

l ~---.-•~o----~°~, 0.73 ~ - - - - - " " ' - - " - o ~ • . - " " - "'"~iJ 0.70 0 ~ ~ - - " ~ ~ 0.61 0 50 100 150 200 250

l ~-'~•"---..o~• ~ ~o 2.05 " r 07" ? ~ o - - ¶ ~ ? ~ o 0.61 0 50 100 150 200 250

I 250

~ ,,-,.--.,.,.. o ~

•

Depth in filter (cm) Fig. 5. Nitrite concentrations of the percolating water in the nitrifying biofilter at three hydraulic loading rates. Values indicate the influent ammonia concentration (g NH4-N m- 3) at the moment of sampling. In both cases, a linear relationship is found, which indicates a fixed ratio between ammonia and nitrite at all nitrification rates encountered. However, the ratio differs between the two systems by a pronounced high nitrite-ammonia ratio in the eel culture system, whereas the turbot culture system is characterized by a very low ratio. Estimations of the values a and b in equations (1) and (2) are made by evaluating the characteristics of the biofilm from the upper part of both filters (Fig. 8). The estimations of a and b are presented in Table 1, together with the ratios of nitrite-N/ammonia-N as observed in the fish culture systems. DISCUSSION

In this study it is demonstrated that in the recirculating fish culture systems studied, nitrite concentrations always stabilized at an equilibrium concentration, in agreement with the results of the biofilm experiments with continuous loading with ammonia (Fig. 2). This argument was not completely

valid as results with a biofilm sample were extrapolated to a whole trickling filter disregarding the observed effects of bacterial stratification (Fig. 3) and hydraulic loading effects in such filters (Nijhof, 1995a). Therefore, different nitritification and nitratification rates over the whole trickling filter could have occurred, resulting in a steadily rising nitrite concentration under some conditions. Although it might seem from Fig. 3 that, especially in the upper parts of the filter bed, nitritification capacity exceeds the nitratification capacity, a situation with net nitrite production during passage was never observed. At any N-loading rate, ammonia and nitrite concentrations remained constant, indicating equal rates of both nitrification steps for the whole filter. Exceptional occasions will be the transient situations during an increase in N q o a d i n g rate, in which a net nitrite production should indeed occur. These results are in close agreement with findings of van Rijn and Rivera (1990), where biofilter nitrite 12 --

10 --

NO2--N

o~/Eel

culture system

10 --

z leq © 8 -z

4' ,/,.

Z

too

6

6

Z .-

.~

¢o

4

2

•

o .Me(/-

r.)

0

4 --

O

~k Z

I

I

I

L

J

48

96

144

192

240

Time (h) Fig. 6. Ammonia and nitrite influent concentrations of the eel culture system during the period with daily increased ammonia supply.

o[-.--~ 0

"1

I o

I

k

I

I

I

2

3

4

5

6

7

Ammonia concentration (g NH4-N m 3) Fig. 7. Nitrite concentrations in both the eel culture system and the turbot culture system plotted against the ammonia concentration during the period with daily increased ammonia supply.

Nitrite accumulation during nitrification 1.5 -

/

4,

,R2=097,

'7

//

1.5 -

/

NH4-N 05 k=-0"I5+0"56SNH4

_.--,

2291

/

k = 0'11 + 0-42 SOO5~ -

~4,

y

/

/

E

Z 1.0

.= o

• "/"

-

--

•

O

o 0.5 --

~

O / 0

d

~ .. oO

NO2--N

^.

(R2 = 0.74)

O

,...~_~" e

N.,N

t,,

k---0A 6 + 0.51 SNH4 (R2 = 0.98)

/ ~

"~ O 0.5 --

E Z

/,4

~'o,.¢o"

Z ,..¢

Io

I

I

I

2

3

J

0

4

0

~ / N concentration (g m-3)°'5

I

2

3

4

X~N concentration (g m-3) °'5

Fig. 8. Ammonia-N and nitrite-N removal rates as a function of ~/SNM,and x/Sr~o:, respectively, for the biofilms from the trickling filters of the eel culture system (left) and the turbot culture system (right). production also increased with increasing N-loading rate. However, they reported that at a m m o n i a influent concentrations exceeding 1 g NH4-N m -3, a production of nitrite during passage through the filter occurred, leading to "nitrite accumulation", but again, no information is given as to what exactly was meant. The level of nitrite concentration can be explained by diffusional transport mechanisms and differences in biofilm characteristics. There is no argument that high nitrite concentrations are caused by nitratification inhibition. According to equations (1)-(4), a fixed ratio between a m m o n i a and nitrite concentrations can be expected in recirculating water as was observed. The quantitative estimation presented in Table 1 shows that the nitrification characteristics are indicative for the observed concentration ratios. The rise in nitrite concentration indicates that the nitratification rate needed for complete removal of the produced nitrite can only be achieved at higher bulk fluid nitrite concentrations. This demonstrates a substrate limited diffusional process, not different from, for example, the biofilm a m m o n i a removal process. Nitratification differs from nitritification by having the substrate produced within the biofilm layer itself. Nevertheless, the reason diffusional mechanisms can explain the observed phenomena is schematically depicted in Fig. 9. The relation between bulk fluid nitrite concentration and nitritification is likely to be found in a partial outward diffusion of nitrite (Bovendeur, 1989). As nitrite is produced at the biofilm surface, a high Table 1. Estimated values of a and b for the eel culture system and the turbot culture system, together with the observed ratio between nitrite-N and ammonia-N concentrations in both systems Eel culture system Turbot culture system

a

b

(a/b) 2

0.56 0.42

0.24 0.51

5.44 0.68

SNo2/Ssr~4

4.0 0.4

concentration gradient with an initially low nitrite concentration in the bulk fluid is induced during nitritification. An equilibrium situation, with no net outward diffusion of nitrite and thus complete nitrification into nitrate, is achieved at a bulk fluid nitrite concentration equal to the nitrite concentration of the upper biofilm layer. At higher nitrification rates, which occur at higher bulk fluid a m m o n i a concentrations, enhanced nitrite concentrations are induced in the intercellular biofilm fluid, which again would cause an outward diffusion of nitrite. The equilibrium will be readjusted at a higher nitrite bulk fluid concentration that prevents the nitrite outward diffusion. Thus, a nitrifying trickling filter in a single pass configuration, as opposed to the recirculating systems used in this study, might indeed display an apparent deficiency in nitratification capacity, judged by the increased nitrite concentration in the effluent compared to the influent.

N concentration

Biofilm Biofilm [Bulk fluid suppoy

/ / / / / / / / /

~

--NH4 +

I

I

"" \ ~ +

Nitritification ~

Nitratification NO3~_I

NO2-

NO2-

]

Depth in filter

°, ~ NO3NO3"-""1

Fig. 9. Tentative scheme of biofilm nitrification, incorporating a net outward diffusion of nitrite, depending on bulk fluid nitrite concentration. Left: hypothetical concentration profile of a single pass trickling filter with an influent containing only ammonia.

M. N~hofand A. KlapwOk

2292

The observed n i t r i t e - N / a m m o n i a - N ratio in the recycled water, i n d e p e n d e n t of nitrification rate, appears to vary between biofilms. Biofilms m a i n t a i n i n g a relatively low nitrite c o n c e n t r a t i o n are characterized by a relatively high nitrite oxidation capacity. O n the other h a n d , low nitrite c o n c e n t r a t i o n s are f o u n d in systems characterized by a relative high nitrite oxidation capacity. The variation e n c o u n t e r e d in the relative capacities o fnitritification and nitratification o fvarious biofilms has been reported as a p a r a m e t e r to distinguish between developing biofilms a n d full-grown biofilms (Bovendeur, 1989). Regarding the fact t h a t all biofilms in this study were several years old and continuously operative, it should be n o t e d that terms such as " d e v e l o p i n g " and "full g r o w n " should be interpreted carefully; a biofilm develops a specific nitrification characteristic, but a high relative nitratification capacity is not necessarily the final developmental stage encountered in all nitrifying biofilms. CONCLUSIONS The nitrite c o n c e n t r a t i o n in recirculating fish culture systems stabilizes at a level at which all the produced nitrite is oxidized. The ratio between a m m o n i a and nitrite concentration in the recirculating water appears c o n s t a n t for a specific fish culture system. This ratio is well explained by the half-order nitritification a n d nitratification rates o f the specific biofilter. The ratio of nitritification a n d nitratification rates in biofilters varies, a n d a relative high nitratification rate is neither i n h e r e n t to a long-term operating biofilter n o r to the salinity. REFERENCES

Anthonisen A. C., Loehr R. C., Prakasam T. B. S. and Srinath E. G. (1976) Inhibition of nitrification by ammonia and nitrous acid. J. Wat Pollut. Control Fed. 48, 835-852. Balmelle B., Nguyen K. M., Capdeville B., Cornier J. C. and

Deguin A. (1992) Study of factors controlling nitrite build-up in biological processes for water nitrification. Wat. Sci. Technol. 26, 1017 1025. Bendschneider K. and Robinson R. J. (1952) A new spectrophometric method for the determination of nitrite in sea water. J. Mar. Res. 11, 87. Bovendeur L, Eding E. H. and Henken A. M. (1987) Design and performance of a water recirculation system for high-density culture of African catfish, Clarias gariepinus (Burchell 1822). Aquaculture 63, 329-353. Bovendeur J. (1989) Fixed-biofilm reactors applied to waste water treatment and aquacultural water recirculating systems. Ph.D. thesis, Agricultural University Wageningem The Netherlands. Harremo~s P. (1982) Criteria for nitrification in fixed film reactors. Wat. Sci. Technol. 14, 167-187. Nijhof M. and Bovendeur J. (1990) Fixed film nitrification characteristics in sea water recirculation fish culture systems. Aquaculture 87, 133-143. Nijhof M. (1995a) The application of commercially exploited recirculating system concepts in the culture of turbot Scophthalmus maximus (L.). In Turbot Culture; Problems and Prospects (Edited by Lavens P. and Remmerswaal R. A. M.), pp. 272 282. European Aquaculture Society, Special Publication No. 22, Gent, Belgium. Nijhof M. (1995b) Bacterial stratification and hydraulic loading effects in a plug-flow model for nitrifying trickling filters applied in recirculating fish culture systems. Aquaculture. In press. Otte G. and Rosenthal H. (1979) Management of a closed brackish water system for high-density fish culture by biological and chemical water treatment. Aquaculture 18, 169 181 Poxton M. G., Murray K. R., Linfoot B. T. and Pooley A. B. W. (1981) The design and performance of biological filters in an experimental mariculture facility. Proc. World Symp. on A quaculture in Heated Effluents and Recirculation Systems, Stavanger, 28 30 May 1980. Sol6rzano L. (1969) Determination of ammonia in natural waters by the phenolhypochlorite method. Limnol. Oceanogr. 14, 799 801. van Rijn J. and Rivera G. (1990) Aerobic and anaerobic biofiltration in an aquaculture unit--nitrite accumulation as a result of nitrification and denitrification. Aquacult. Engng 9, 217 234. Yang L. and Alleman J. E. (1992) investigation of batchwise nitrite build-up by an enriched nitrification culture. War. Sci. Technol. 26, 997-1005.