Environment International 31 (2005) 1 – 13

www.elsevier.com/locate/envint

Phytoremediation of aquaculture wastewater for water recycling and

production of fish feed
A.E. Ghaly *, M. Kamal, N.S. Mahmoud
Biological Engineering Department, Dalhousie University, P.O. Box 1000, Halifax, Nova Scotia, Canada B3J 2X4

Received 8 May 2004; accepted 28 May 2004

Available online 20 July 2004

Abstract

Five plants were examined for their ability to remove nutrients from aquaculture wastewater and suitability as fish feed: alfalfa, white
clover, oat, fall rye, barley. The seeds were first germinated in water in a hydroponic system, and the plants were fed wastewater from Tilapia
production facility. Clover and alfalfa seeds were infected with fungus shortly after germination, and their roots were completely destroyed by
day 14. Oat, rye and barley had the fastest growth and showed greater tolerance to fungal disease compared with alfalfa and clover. Although
substantial amounts of soluble and insoluble substances were released by the seeds during the germination period, the plants were able to
remove all the pollutants in wastewater and significant portions of those released substances. The total reductions in total solids, COD, NO3 –
N, NO2 – N, phosphate and potassium ranged from 54.7% to 91.0%, 56.0% to 91.5%, 82.9% to 98.1%, 95.9% to 99.5%, 54.5% to 93.6% and
99.6% to 99.8%, respectively. Oat, barley and rye grow well in this type of hydroponic system and can be used as a fish feed after being
supplemented with fat, Ca, Na, Mn and Fe. Oil seeds and the chlorides of these elements could be added to these plants when formulating the
fish feed. For a continuous operation, a two-unit system could be configured to allow for one week germination and one week cleaning and
startup in one unit while the other unit is in operation.
D 2004 Elsevier Ltd. All rights reserved.

Keywords: Aquaculture; Wastewater; Phytoremediation; Hydroponic; Fish feed; Alfalfa; White clover; Oat; Fall rye; Barley

1. Introduction waters as organic matter loading reduces dissolved oxygen

Until recently, fish farming was limited to areas where
water resources were almost unlimited. Techniques involv-
ing the recirculation of the wastewater after biological
treatment and/or filtration can make aquaculture almost
independent of available water. However, the content of
nutrients in the waste stream from these systems will
probably set the limits to future fish farm size. Aquaculture
systems produce large quantities of organic matter and
nutrients (nitrogen, phosphorus and other elements) that
require treatment and/or disposal. The production of 1 ton
(1000 kg) of live channel catfish releases 1190 kg of dry
matter, 60 kg of nitrogen and 12 kg of phosphorus to the
culture water as metabolic wastes (Boyd, 1985). Aquacul-
ture wastewaters exert adverse environmental impacts when
the effluent from these systems is discharged to receiving
* Corresponding author. Tel.: +1-902-494-3275.
E-mail address: abdel.ghaly@dal.ca (A.E. Ghaly).
levels and contributes to the buildup of bottom sediments
and high nutrient loading impairs water quality by stimu-
lating excessive phytoplankton production (Joyner, 1992).
Hydroponic production systems have potential for the
treatment and reuse of wastewater in intensive aquaculture
systems. Combining aquaculture with the hydroculture
technique may serve the double purpose of reducing the
pollution caused by fish farming and the demand for
commercial fertilizers, thereby helping to preserve surface
and ground water quality (Pettersen, 1987). Aquaculture
systems integrated with plant culture involve the use of
nutrient-rich effluent from aquaculture production tanks to
provide water and nutrients to plants (Mathieu and Wang,
1995).
Recirculating systems that incorporate hydroponics are
referred to as aquaponic systems. Substantial research has
been done on the use of aquatic plants for wastewater
treatment. Algae culture, floating emerged or submerged
plant culture, raft or suspended culture, nutrient film tech-
nique (NFT), aeroponics and static culture (combination of

0160-4120/$ - see front matter D 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.envint.2004.05.011
2 A.E. Ghaly et al. / Environment International 31 (2005) 1–13
inert medium and water culture) represent different water
culture designs (Mathieu and Wang, 1995). Although some
of these systems are effective, the production of large
quantities of aquatic plants with low economic value poses
another waste problem (Rackocy, 1995). Thus, crops can be
cultured hydroponically in recirculating systems to produce
a valuable by-product, while improving the water quality is
highly desirable. High-value vegetable crops, such as toma-
toes, lettuce, cucumbers and sweet basil, have been cultured
in a variety of hydroponic media (Rackocy and Hargreaves,
1993). However, the production of crops that can be used as
fish feed may have some economic and technical advan-
tages over these crops.
2. Objectives
In this study, the possibility of using agriculture plants in
reducing the pollution potential of aquaculture wastewater
and their suitability as a fish feed were investigated. The
objectives were to evaluate (a) the effectiveness of five
plants (barley, white clover, fall rye, oat and alfalfa) in
reducing the pollution potential of aquaculture wastewater,
as measured by TS, COD, nitrogen compounds, P and K
and (b) the nutritional content and suitability of the plants as
fish feed.
3. Experimental apparatus
The hydroponic system (Fig. 1) consisted of a frame,
growth troughs and aeration, lighting, cooling, waste appli-
cation, supernatant collection and control systems.
The frame (Fig. 2) was constructed of angle iron with a
width of 244 cm, a depth of 41 cm and a height of 283 cm.
The back and the top were covered with 0.6-cm-thick
plywood sheets. The frame consisted of three shelves (76
cm apart), each shelf was divided vertically into two cells by
dividers made of 1.2 cm thick plywood sheets. This pro-
vided a better control of the light and feeding.
The hydroponic system has six growth troughs; the
dimensions of the each trough and plant supporting tray
are shown in Fig. 2. Each trough was made of galvanized
steel and was divided into three compartments. Each com-
partment held a tray that acted as the plant support medium
and was made with a wire-mesh base (16 openings/cm2) and
5-cm-high metal sides. The trays were positioned in the
troughs so that the plant roots were in contact with the
liquid waste. This was done by means of supports welded
into the corners of each compartment 5 cm below the top
edge of the trough. An outlet from each compartment
allowed the supernatant from that compartment to be
collected separately.
An aeration unit was installed in each compartment to
provide oxygen to the immersed roots of the growing crops.
The air flow from the main supply to the manifold on each
shelf was controlled by a pressure regulator (Model 129121-
510, Aro, Brayn, OH). Six aeration units were connected to
a manifold on each shelf using plastic tubing of 0.75 cm
outside diameter. Small screw valves mounted on each
manifold were used to control the airflow to each aeration
unit, which consisted of a main tube with three perforated
stainless steel laterals coming off it at right angles to the
main. The tubing had an outside diameter of 0.6 cm. Each
lateral was approximately 30 cm long, whereas the main
was 26.5 cm long.
The lighting system was designed to provide approxi-
mately 430 hlx of illumination (19280 lumen per trough).
This was achieved by a mixture of fluorescent and incan-
descent lamps. Six 40-W cool white fluorescent lamps (122
cm in length) and two 100-W incandescent bulbs were
fastened above each trough. The lights were controlled by
an electronic circuit, which was set to provide the required
number of hours of light per trough per day.
A cooling system was designed to continuously remove
the heat produced by the lamps to avoid heating of the
wastewater on the upper and middle shelves. For each of
these two shelves, a 5-cm-diameter PVC pipe, having 6-
mm-diameter holes spaced 6 cm apart and facing out, was
placed under the backside of the troughs. Two metal blocks
supported the front side of the trough. This provided a 5-cm
space between the trough and the lighting system of the
shelf below it. A 5-cm-diameter PVC pipe acting as a
manifold was attached vertically to the left side of the
frame, through which air was blown by means of motor-
driven fan (Model AK4L143A type 821, Franklin Electric,
Bluffton, IN).
The waste application system consisted of a control
system, a feeding tank and a pump. The feeding tank had
a capacity of approximately 100 l. A mixing shaft, with a
40-cm-diameter impeller, was installed through the center of
the cover of the tank to agitate the wastewater in the tank.
Four 2.5-cm baffles were installed vertically along the inside
wall of the tank to promote complete mixing. A 1 hp motor
(Model NSI-10RS3, Bodine Electric, Chicago, IL) with
speed reducer was mounted on the tank cover to drive the
mixing shaft and impeller. A variable speed pump (Model
110-23E, TAT Pumps, Logan, OH) with a capacity of 138
cm3/rev was used to transfer the wastewater from the feed
tank to the applicators through a manifold (PVC pipe of 2.5-
cm outside diameter) connecting a system of feed tubes to
six solenoid valves. The wastewater was pumped from the
feed tank to a manifold, which had six outlets connected to
six wastewater applicators through the solenoid valves that
controlled the amount of wastewater fed to each cell. Each
wastewater applicator was fabricated from stainless steel
pipe with holes punched along the lower edge to allow the
wastewater to flow out. The wastewater entered the appli-
cator at the center of the top edge. To overcome the problem
of clogging, a water line with six solenoid valves attached to
the applicator was used to flush out the applicator after
feeding periods. The wastewater application system was
A.E. Ghaly et al. / Environment International 31 (2005) 1–13 3

Fig. 1. Experimental setup.

4 A.E. Ghaly et al. / Environment International 31 (2005) 1–13
Fig. 2. The frame, plant support trays, aeration unit and lighting system.
fully automated and consisted of a motor-driven pulley
arrangement on each shelf to which the applicator tubes
were attached. The motors (Sigma Model 20-3424SG-
24007, Faber Industrial Technologies, Clifton, NJ) ran at 6
rpm and were controlled by an electronic circuit. The system
was set up so that each applicator traveled 122 cm (three-
tray length) when a guide on an applicator hit a microswitch
located at each end of the shelf, the motor stopped, waited a
few seconds and then reversed and the applicator traveled in
the opposite direction. This process continued for the
designated feeding time, which was controlled by computer.
The supernatant from each tray was collected in separate
containers (2.7 l each) located at the bottom of the system
by means of tubes attached to the outlet pipes of each trough
compartment. The outlets were connected to plastic tubes of
1 cm outside diameter, which passed through a solenoid
valve. Prior to feeding, the outlet tubes were closed using
solenoid valves so that the fresh wastewater was retained in
the trough compartment to allow time for the plants to have
assimilate the fresh nutrients being added.
An IBM computer was used to operate and control the
various components of the hydroponic system. A basic
computer program allowed the user to configure the
operating frequency and duration of lighting system,
wastewater application system and supernatant collection
system. The computer was connected to a data coordina-
tor, which had 24 digital output ports and 24 digital input
ports. The digital output ports were connected to elec-
tronic circuits, which were responsible for lighting system,
the cooling system, the wastewater application and super-
natant collection systems (solenoid valves, pump and
applicator).
4. Experimental procedure
4.1. Plant seeds
Alfalfa, white clover, fall rye, barley and oat were
used in this study. The white clover, fall rye and alfalfa
 A.E. Ghaly et al. / Environment International 31 (2005) 1–13 5

seeds were purchased from Halifax Seed, Halifax, Nova Table 2

Nutrient requirements for various crops (Atlantic Provinces, 1985; Bruce,
Scotia. The barley and oat seeds were purchased from
1995)
Walker’s Livestock, Halifax. The amount required to
Amount of nutrient required (kg/ha) NPK ratio
cover the surface of a tray was approximately 125, 125,
225, 250 and 250 g for alfalfa, white clover, fall rye, Nitrogen Phosphorus Potassium
barley and oat, respectively. Three different amounts of Alfalfa 113 59 180 10: 5.2: 15.9
seeds (Table 1) were tried to determine the effect of seed White clover 170 40 110 10: 2.3: 6.5
Fall rye 50 22 90 10: 4.4: 18
over- (or less) crowding on plant growth and nutrient
Barley 56 28 17 10: 5: 3
uptake. Oat 56 22 22 10: 3.9: 3.9

4.2. Light duration

4.5.2. Days 2– 7
The selected crops are usually grown during the summer
Germination of seeds was observed and recorded daily.
months when the days are longer. The average length of day
More water was added to the compartments as required to
during the crop-growing period in Canada is approximately
compensate for loss of water by evaporation. The seedling
16 h. The system light intensity was therefore maintained
height was measured in each tray and recorded on days 3
constant at 43,000 lx for 16 h/day.
and 5. A sample of the wastewater was taken from each
compartment and stored in a fridge (at 4 jC) in a labeled
4.3. Nutrient requirements
bottle for chemical analyses.
The nitrogen, phosphorus and potassium requirements
4.5.3. Day 8
for the plants used in this study are summarized in Table 2.
The seedling height was measured in each tray and
The requirements (kg/ha) are for soil plants and were
recorded. A sample of the effluent from each compartment
adapted for hydroponic plants.
was taken and stored in a labeled bottle for chemical
analyses. The required amount of wastewater was applied
4.4. Wastewater application
to each compartment.
The aquaculture wastewater was obtained from Aqua
Farm located in Chezzacook, Halifax County, Nova Scotia. Table 3
The chemical analyses for the aquaculture wastewater are Chemical analyses of influent aquaculture wastewater
presented in Table 3. The calculated NPK requirements for Parameter Concentration (mg/l)
the plants (mg/tray/day) are presented in Table 4. Because Total solids (mg/l) 1000
phosphorus is the growth-limiting nutrient for all plants, the Organic solids 290
application rate of wastewater was based on the phosphorus Fixed solids 690
content. The amount of nutrients provided for each plant is Suspended solids (mg/l) 100
presented in Table 4. Organic solids 80
Fixed solids 18
Chemical oxygen demand (mg/l)
4.5. Experimental protocol Total COD 240
Soluble COD 150
4.5.1. Day 1 Total Kjeldahl nitrogen (mg/l) 60
The required amounts of seed were weighed and Ammonia-Nitrogen 7
Organic-Nitrogen 53
placed on the trays in the growth troughs. The solenoid Nitrate-Nitrogen (mg/l) 110
valves controlling the outlet tubes were turned off, and Nitrite-Nitrogen (mg/l) 5
each compartment was filled with water to a level such Elements
that the seeds were in contact with the water but not Total phosphorous 50.40
submerged. Orthophosphate 6.60
Potassium 230.00
Calcium 50.70
Table 1 Sodium 30.90
Quantities of seeds used in the experiment Sulfur 46.00
Amount of seeds (g) Chloride 57.00
Magnesium 15.90
Less crowded Normal Overcrowded
Manganese 0.04
Alfalfa 75 125 175 Iron 0.37
White clover 75 125 175 Copper 0.03
Fall rye 175 225 275 Zinc 0.20
Barley 200 250 300 Boron 0.16
Oat 200 250 300 pH 7.3
6 A.E. Ghaly et al. / Environment International 31 (2005) 1–13

Table 4
Nutrient required for each crop, amount of wastewater applied and amount of nutrients provided in the waste
Amount of nutrients required (mg/tray/day) Amount of waste Amount of nutrients provided (mg/tray/day)
applied (ml/day/tray)
Nitrogen Phosphorus Potassium Nitrogen Phosphorus Potassium
Alfalfa 120.8 63.0 192.4 1105 134.8 (E 14.0) 63.0 258.6 (E 66.2)
White clover 181.7 42.8 107.6 750 91.5 (D 90.2) 42.8 175.5 (E 67.9)
Fall rye 53.5 23.5 96.2 412 50.3 (D 3.2) 23.5 96.4 (E 0.2)
Barley 59.9 29.9 18.2 525 64.1 (E 4.2) 29.9 122.6 (E 104.4)
Oat 59.9 23.5 23.5 412 50.3 (D 9.6) 23.5 96.4 (E 72.9)
E: amount in excess of requirement.
D: amount deficient below requirement.

4.5.4. Days 9– 12 4.6. Chemical analyses

Wastewater application continued on daily basis. A
sample of the effluent was taken from each compartment
before waste application and stored in the fridge (at 4 jC) in
a labeled bottle for chemical analyses. The solenoid valves
were then opened to allow the effluent to exit through the
outlet tubes. The seedling height in each tray was measured
and recorded on days 10 and 12.
4.5.5. Day 14
The crop height was measured and recorded. A sample of
the effluent from each compartment was taken and stored in
the fridge (at 4 jC) in a labeled bottle for chemical analyses.
The experiment was terminated for alfalfa and clover
because of fungus infection. Wastewater application contin-
ued on daily basis for the remaining plants.
Chemical analyses [potassium, orthophosphate, nitrite–
nitrogen, nitrate – nitrogen, chemical oxygen demand (COD)
and total solids] were carried out within 24 h on the effluent
from the system in triplicates. The analyses were performed
according to the procedures described in the Standard
Methods (APHA, 1980). The carbohydrates, digestible
energy, crude fat and protein were performed on the samples
at Philip Analytical Services, Dartmouth, Nova Scotia. The
elemental composition of the plants was performed at the
Mineral Laboratory of Dalhousie University, Halifax, Nova
Scotia.
5. Results

4.5.6. Day 21 5.1. Plant morphology

A sample of the effluent was taken from each compart-
ment and stored in a labeled bottle for chemical analyses. Within 24 h of placing the seeds onto the germination
Samples were also taken from the plants (rye, barley and oat) trays, they started to swell and began to germinate. After 2
for nutritional analyses. The experiment was terminated. days, the radicle and plumule had broken through the seed

Table 5
Total solids
Plant type Seed Input (mg/l) Effluent Total reduction Reduction from released substances
quantity (mg/l)
Wastewater Released Total (mg/l) (%) (mg/l) (%)
substances
Alfalfa 75 1000 990 1990 750 1240 62.0 240 24.2
125 1000 1660 2660 710 1950 64.3 950 57.2
175 1000 2320 3320 690 2630 79.0 1630 70.3
Clover 75 1000 920 1920 870 1050 54.7 50.0 5.4
125 1000 1540 2540 850 1670 66.0 670 43.5
175 1000 2150 3150 820 2330 74.0 1330 61.2
Rye 175 1000 2010 3010 570 2440 81.0 1440 71.6
225 1000 2590 3590 410 3180 89.0 2180 84.2
275 1000 3160 4160 380 3780 91.0 2780 88.0
Barley 200 1000 990 1990 300 1690 85.0 690 69.7
250 1000 1240 2240 260 1980 88.5 980 79.0
300 1000 1490 2490 230 2260 91.0 1260 84.6
Oat 200 1000 710 1710 420 1290 75.5 290 40.8
250 1000 880 1880 380 1500 80.0 500 56.8
300 1000 1060 2060 320 1740 84.5 740 69.8
CV = 2.3 – 9.5%.
 A.E. Ghaly et al. / Environment International 31 (2005) 1–13 7

Table 6
Chemical oxygen demand
Plant type Seed Input (mg/l) Effluent Total reduction Reduction from released substances
quantity (mg/l)
Wastewater Released Total (mg/l) (%) (mg/l) (%)
substances
Alfalfa 75 240 480 720 230 490 68.0 250 52.1
125 240 760 1000 200 800 80.0 560 73.7
175 240 1070 1310 190 1120 85.5 880 82.2
Clover 75 240 460 700 310 390 56.0 150 32.6
125 240 705 945 270 675 71.5 455 61.8
175 240 990 1230 260 970 78.0 730 73.4
Rye 175 240 425 665 180 485 73.0 245 57.6
225 240 620 860 130 730 85.0 490 79.0
275 240 755 955 120 875 88.0 635 84.1
Barley 200 240 235 475 70 405 85.0 165 70.2
250 240 295 535 60 475 89.0 235 79.7
300 240 355 595 50 545 91.5 305 85.9
Oat 200 240 365 605 130 475 78.5 235 64.4
250 240 455 695 120 575 82.5 335 73.6
300 240 545 785 110 675 86.0 435 79.8
CV = 1.0 – 11.7%.

coat and were visible on most of the oats, barley and rye
seeds while alfalfa and clover seeds started to swell and
germinate after day 3. By day 5, the root mat was starting to
develop, and the seedlings of alfalfa, clover, oat, rye and
barley were grown approximately 4, 5, 10, 12 and 8 cm in
height, respectively. By day 8, the seedlings of alfalfa,
clover, oat, rye and barley were 6, 7, 14, 14 and 17 cm in
light, respectively.
Generally, the plant height increased with time. Oat,
barley and rye grew the fastest, while alfalfa and clover grew
the slowest. The seed density did not seem to affect plant
height or the growth rate. The three cereal crops (oat, rye and
barley) grew very well in the hydroponic system, whereas
alfalfa and clover were suitable for hydroponic system, as
they were attacked by fungus shortly after germination (day
5), and by day 10, the trays were completely infected with
fungus that destroyed most of their roots.
5.2. Effluent characteristics
The seeds were first germinated in tab water for 7 days,
and the wastewater application started on day 8 and
continued for 2 weeks. The total solids, COD, nitrate –
nitrogen, nitrite –nitrogen, orthophosphate and potassium
concentrations in the effluent measured on day 14 are
shown in Tables 5 –10. The values are the average of three

Table 7
Nitrite-nitrogen
Plant type Seed Input (mg/l) Effluent Total reduction Reduction from released substances
quantity Wastewater Released Total (mg/l) (mg/l) (%) (mg/l) (%)
substances
Alfalfa 75 5 1.68 6.68 0.60 6.48 91.1 1.48 88.1
125 5 2.79 7.79 0.40 7.59 94.9 2.59 92.8
175 5 3.92 8.92 0.20 8.72 97.8 3.72 94.9
Clover 75 5 0.84 5.84 1.00 5.64 82.9 0.64 76.2
125 5 1.39 6.39 0.60 6.19 90.6 1.19 85.6
175 5 1.95 6.95 0.40 6.75 94.2 1.75 89.7
Rye 175 5 0.23 5.23 0.10 5.13 96.2 0.13 56.5
225 5 0.29 5.29 0.10 5.19 98.1 0.19 65.5
275 5 0.37 5.37 0.10 5.27 98.2 0.27 73.0
Barley 200 5 0.21 5.21 0.10 5.11 96.1 0.11 52.4
250 5 0.26 5.26 0.10 5.16 96.2 0.16 61.5
300 5 0.31 5.31 0.10 5.21 98.1 0.21 67.7
Oat 200 5 0.76 5.76 0.20 5.56 96.5 0.56 73.7
250 5 0.95 5.95 0.20 5.75 96.6 0.75 78.9
300 5 1.14 6.14 0.20 5.94 96.7 0.94 87.5
CV = 8.1 – 12.5%.
8 A.E. Ghaly et al. / Environment International 31 (2005) 1–13

Table 8
Nitrate-nitrogen
Plant type Seed Input (mg/l) Effluent Total reduction Reduction from released substances
quantity (mg/l)
Wastewater Released Total (mg/l) (%) (mg/l) (%)
substances
Alfalfa 75 110 10.0 120.0 4.9 115.1 95.9 5.1 51.0
125 110 12.2 122.2 3.9 118.3 96.8 8.3 68.0
175 110 13.5 123.5 3.7 119.8 97.0 9.8 72.6
Clover 75 110 8.0 118.0 4.7 113.3 96.0 3.3 41.2
125 110 9.0 119.0 3.4 115.6 97.1 5.6 62.2
175 110 12.0 122.0 2.6 119.4 97.9 9.4 78.3
Rye 175 110 1.5 111.5 0.5 111.0 99.5 1.0 66.6
225 110 2.3 112.3 0.7 111.6 99.3 1.6 69.6
275 110 2.9 112.9 0.8 112.1 99.3 2.1 72.4
Barley 200 110 1.6 111.6 0.5 111.1 99.5 1.1 68.8
250 110 2.5 112.5 0.7 111.8 99.4 1.8 72.0
300 110 3.0 113.0 0.7 112.3 99.4 2.3 76.7
Oat 200 110 1.9 111.9 0.6 111.3 99.5 1.3 68.4
250 110 2.9 112.9 0.8 112.1 99.8 2.1 72.4
300 110 3.2 113.2 0.8 112.4 99.3 2.4 75.1
CV = 2.3 – 7.1%.

measurements. The coefficients of variation were very sugars (which are important source of respiration during the
small (ranged from 1.0 to 12.5%). germination period) as follows (Mayer and Doljakoff-
Mayber, 1975):
a amylase
6. Discussion Amylose or Amylopectin ! Glucose þ Maltose
ð1Þ
6.1. Effluent quality

Seeds contain protein, fat, carbohydrates and elements a glucosidase

(Table 11). The proteins include albumins, globulins, stut-
lins and prolamins, of which only albumins are water
soluble. The fats include phospholipids, glycolipids and
sterols, all of which are water soluble. Starch is the
carbohydrate most commonly found in seeds, although
hemicellulose, cellulose, raffinose, mucilages, pectins and
series of oligosaccharides may be present. Starch is found in
the forms of amylase and amylopectin and is converted to
Maltose ! Glucose ð2Þ
As seeds start to take up water during germination, there
is a rapid leakage into the medium of solutes, such as
sugars, organic acids, ions, amino acids, proteins and
growth enzymes such as gibberellin and cytokinins, all of
which affect the quality of the growth medium (Bewley and
Black, 1985).

Table 9 Table 10
Ortho-phosphate Potassium
Plant type Seed Input Effluent Reduction Plant type Seed Input Effluent Reduction
quantity (mg/l) (mg/l) quantity (mg/l) (mg/l)
(mg/l) (%) (mg/l) (%)
Alfalfa 75 6.6 2.77 3.83 58.0 Alfalfa 75 230 0.97 229.03 99.6
125 6.6 1.98 4.62 70.0 125 230 0.97 229.03 99.6
175 6.6 0.96 5.64 85.5 175 230 0.97 229.03 99.6
Clover 75 6.6 3.00 3.60 54.5 Clover 75 230 0.44 229.56 99.8
125 6.6 2.12 4.48 67.9 125 230 0.45 229.56 99.8
175 6.6 0.98 5.62 85.2 175 230 0.44 229.56 99.8
Rye 175 6.6 2.13 4.47 67.7 Rye 175 230 0.81 229.19 99.6
225 6.6 1.53 5.07 76.8 225 230 0.66 229.34 99.7
275 6.6 0.60 6.00 90.9 275 230 0.42 229.53 99.8
Barley 200 6.6 0.54 6.06 91.8 Barley 200 230 0.47 229.53 99.8
250 6.6 0.50 6.10 92.4 250 230 0.42 229.58 99.8
300 6.6 0.42 6.18 93.6 300 230 0.19 229.81 99.9
Oat 200 6.6 0.57 6.03 91.4 Oat 200 230 0.85 229.15 99.6
250 6.6 0.56 6.04 91.5 250 230 0.67 229.33 99.7
300 6.6 0.51 6.09 92.3 300 230 0.57 229.43 99.8
CV = 9.0 – 12.1%. CV = 2.2 – 6.9%.
 A.E. Ghaly et al. / Environment International 31 (2005) 1–13
Table 11
The protein, fat, carbohydrates and elements content of seeds
Seed type Protein Fat Carbohydrates Elements (%)
(%) (%) (%)
Total P K
Alfalfa 25 6 52 17 1.13
Clover 24 6 56 13 1.00
Rye 12 2 76 10 0.71
Barley 12 3 76 9 0.43
Oats 13 8 66 13 0.96
6.1.1. Total solids
The solids concentration in each compartment increased
with time during the germination period and was dependent
on both the quantity and the type of seeds (Table 5). The
increase was due to the release of dissolved and suspended
solids from the seeds as a result of the germination process.
Increasing the quantity of seeds increased the amount of
solids released by the plants. The amount of solids released
was the highest for rye followed by alfalfa, barley, clover
and oat.
The solids concentrations declined during the growth
period because the crop root mates were fully developed
and the filtration capacity of the roots increased with the
growth of the crops. The root system was also capable of
absorbing dissolved solids as plant nutrients. The total solid
reduction in the effluent was influenced by the plant type and
the seed density. Despite the fact that the rye had the highest
solids release, it had the highest filtration capacity, with a
total reduction of 81%, 89% and 91% for the compartments
receiving 175, 225 and 275 g, while clover, which has the
second lowest solid release, had the lowest filtration capacity,
with total reduction of 54%, 66% and 74% for the compart-
ments receiving 75, 125 and 175 g. The amount of total
solids in the effluent was much smaller than that released by
the seeds for all plants, indicating the high efficiency of the
root system of removing the solids in wastewater.
Jiang and Xinyuan (1998) achieved 75% removal of
total suspended solids of zoo wastewater using floating
(water hyacinth and mosquito fern), submerged (curly
pondweed, eelgrass and parrotfeather), floating leaf (hindu
lotus) and emerged plants (swamp morning-glory and
alternanthera alligator). Lin et al. (2002) achieved more
than 90% reduction in aquaculture wastewater suspended
solids using subsurface flow wetland planted with reed
(Paspalum vaginatum). Brix (1994, 1997) stated that the
reduction in suspended solids caused by macrophytes is
attributed to the reduction of water current, which en-
hance the sedimentation of suspended solids and the
filtration of solids by the plant tissue. In the present
study, a total solid reduction of 54.7 – 91.0% was
achieved, depending on the type and quantity of seeds
used. It must be emphasized that seeds were used in the
present study, which contributed 710 – 3160 mg/l of solids
(depending on the type and quantity of seeds used) due to
the release of various substances from the seeds during
the germination period, in addition to the 1000 mg/l of
9
solids in the wastewater. Therefore, not only the plants
filtered out all the solids in the wastewater during the
growth period, but also removed between 24.2% and
88.0% of the materials released by the seeds during
2.18 germination. Furthermore, when fungus uninfected plants
1.51 (rye, barley and oat) were left to grow for another week,
1.13 the removal efficiency of the substances released during
0.56
germination increased to 90.2– 95.6%, depending on the
1.10
type of plant used.
6.1.2. COD
The chemical oxygen demand (COD) concentration in
each compartment increased with time during the seed
germination period due to the release of enzymes and other
growth substances and was dependent on both the quantity
and type of seeds (Table 6). Increasing the quantity of seeds
increased the COD in all compartments. The COD caused
by the substances released by plants was the highest for
alfalfa, followed by clover, rye, oat and barley.
The COD decreased significantly during the growth
period because the crop root mates were fully developed
and the filtration capacity of the roots of suspended solids
increased as well as the absorption of dissolved nutrients.
Barley had the highest COD reduction of 85%, 89% and
91% for the compartments receiving 200, 250 and 300 g,
while the clover had the lowest total reduction of 56%,
71.5% and 78% for the compartments receiving 75, 125 and
175 g, respectively.
Jiang and Xinyuan (1998) achieved 44% reduction of
chemical oxygen demand of zoo wastewater using floating
(water hyacinth and mosquito fern), submerged (curly
pondweed, eelgrass and parrotfeather), floating leaf (hindu
lotus) and emerged plants (swamp morning-glory and alter-
nanthera alligator). Gloger et al. (1995) compared the COD
removal rate of hydroponic tanks that had lettuce plants with
aerated tanks that had no plants in treating fish wastewater.
They reported 54% higher COD removal rate for lettuce
tanks compared with tanks with no plants. In the present
study, the substances released during the germination period
contributed 235 – 1070 mg/l COD, depending on the type
and quantity of seeds used, in addition to the 240 mg/l COD
in the wastewater. The plants, however, were able to remove
all the COD in the wastewater during the growth period, and
475 – 1120 mg/l (32.6 – 85.9%) of the COD caused by the
release substances. When the fungus uninfected plants (rye,
barley and oat) were left to grow for another week, the
removal efficiency of the COD caused by the released
substances during the germination period increased to
88.2 – 98.9%.
6.1.3. Nitrogenous compounds
Seeds contained 12– 25% protein and 2 –6% fat on dry
weight basis (Table 11). The proteins and amino acids
released during the germination period are converted into
ammonium. In addition, the solutes released by the seeds
may stimulate the growth of nitrogen fixing organisms, thus
10 A.E. Ghaly et al. / Environment International 31 (2005) 1–13
increasing the concentration of NH4 in the growth media
(Bewley and Black, 1985).
Organic  N ! NHþ
4 ð3Þ
1=2N2 þ 2H2 O ! NHþ
4 þ O2 ð4Þ
The conversion of N2 to NH4 has a large metabolic
energy requirement, and the sugars released from the seeds
provide a ready source of energy for the nitrogen fixing
process (Mayer and Doljakoff-Mayber, 1975).
Organic Matter þ O2 ! CO2 þ H2 O þ DE ð5Þ
Because the system was aerated continuously, the am-
monium is oxidized into nitrite and then into nitrate as
follows:
nitrosomonase
NHþ 
4 þ 1:5O2 ! NO2 þ H2 O þ 2H
þ
ð6Þ
nitrosbacter
NO 
2 þ 1=2O2 ! NO3 ð7Þ
The nitrite– nitrogen and nitrate – nitrogen concentrations
(Tables 7 and 8) in each compartment increased with time
during the seed germination period and were dependent on
the quantity and the type of seeds. The concentrations were
the highest for alfalfa, followed by clover, oat, rye and
barley. This is because both alfalfa and clover had the
highest protein content that was converted to NH4+ and then
to NO2 and NO3. The concentration of the nitrite –nitrogen
in the final effluent was small, as most of the NO2 was
converted to NO3. About 91.1 – 98.2% of NO2N was
converted to NO3N during the 2-week growth period. The
concentration of the nitrate – nitrogen in the final effluent
was also small because the absorption of NO3 by plants
increased with the growth of the crops. About 95.9 – 99.8%
of NO3N was absorbed by plants. Wastewater application
was based on phosphorous content, and as a result, there
was nitrogen deficiency for clover, ray and oat (Table 4).
This may explain the high removal efficiency of NO3.
Plants prefer NO3 –N or NH4 – N for several reasons: (a)
NH4+ interact with other cations where NO3 is highly
soluble, (b) the rate of delivery of NO3 by diffusion is
much higher than that of NH4+, (c) relatively low NH4+
concentration is toxic to plants and (d) the uptake of NO3
avoids the competition that occur for the root enzyme
between NH4+ and other positively charged ions (Schle-
singer, 1996).
Gloger et al. (1995) attributed the reduction in ammoni-
um ions (NH+4) to both direct plant uptake and nitrification.
They reported that lettuce was responsible for the removal
of 9% of nitrogen applied as feed. Jiang and Xinyuan (1998)
achieved 73% removal of total nitrogen of zoo wastewater
(110 kg total nitrogen/year) using floating (water hyacinth
and mosquito fern), submerged (curly pondweed, eelgrass
and parrotfeather), floating leaf (hindu lotus) and emerged
plants (swamp morning-glory and alternanthera alligator).
Naegel (1977) reported a nitrate level reduction in aquacul-
ture wastewater of about 78% (from more than 450 to about
100 mg/l) in 8 weeks when using tomatoes and about 89%
(from more than 450 to about 50 mg/l) in 4 weeks when
using lettuce. Gloger et al. (1995) reported that the accu-
mulation rate of nitrate nitrogen in a recirculating fish
culture system as a result of fish feed addition was lower
by 24% when lettuce was grown in hydroponic tank. Mant
et al. (2003) achieved 57.7% nitrogen removal using Salix
viminalis grown in gravel hydroponic system to treat pri-
mary settled sewage wastewater. The removal efficiency of
nitrogenous compounds in this study was much higher than
those reported in the literature.
6.1.4. Orthophosphate
Phosphorous is considered to be a major growth-limiting
nutrient in aquatic systems (Marschner, 1988). Wastewater
application was therefore based on phosphorous require-
ment by plants. There was no release of orthophosphate
during the seed germination period, and unlike nitrogen,
there is no atmospheric form that can be made biologically
available. Under normal conditions, phosphorous occurs in
oxidized state either as ions of inorganic orthophosphate
(HPO42  or H2PO4) or organic compounds dissolved in
water. Phosphorous is converted from inorganic to organic
and vise versa, with microorganisms playing a key role in
both transformations. Biologically available phosphorous in
aquatic system includes soluble reactive phosphorous and
soluble unreactive phosphorous. The latter is considered to
be available by enzymatic hydrolysis (Holtan et al., 1988).
The orthophosphate concentration in each compartment
decreased with time during the growth period and was
dependent on both the quantity and the type of seeds (Table
9). The concentration of the orthophosphate in the final
effluent was low because the crop root mates were fully
developed and the amount of orthophosphate absorbed by
the roots increased. The orthophosphate concentrations in
the effluent decreased with the increase of the seed quantity.
Barley had the highest total reduction of 91.81 – 93.63%,
while the clover had the lowest total reduction of 54.54 –
86.36%. The lower removal efficiency of orthophosphate
observed with alfalfa and clover could be attributed to the
lack of growth due to fungus infection. On the other hand,
when the fungus uninfected plants (ray, barley and oat) were
left to grow for another week, the phosphorous removal
efficiency increased to 93.2 –99.1%.
Jiang and Xinyuan (1998) achieved 62% removal of total
phosphorous of zoo wastewater using floating (water hya-
cinth and mosquito fern), submerged (curly pondweed,
eelgrass and parrotfeather), floating leaf (hindu lotus) and
emerged plants (swamp morning-glory and alternanthera
alligator). Brown et al. (1999) reported plant – soil system
removal efficiency of 99% and 97% of total and soluble
reactive phosphorous in saline aquaculture wastewater using
salt tolerant plants grown in sand biofilter. However, they
mentioned that the soil was responsible for the removal of
 A.E. Ghaly et al. / Environment International 31 (2005) 1–13 11

most of the phosphorous because it is readily absorbed to Table 12

Nutritional Value for the oat, barley and rye plants
soil particles, and hence, a small fraction was available to
plants. Gloger et al. (1995) reported a decrease of PO4 Oat Barley Rye Fish feed
concentration in fish wastewater by recirculation in a Carbohydrates (g/kg) 270.5 318.8 330.3 300.0 – 330.0
hydroponic lettuce tank (4.9 mg/l) compared with hydro- Crude fiber (g/kg) 64.1 41.5 41.7 45.0 – 80.0
Crude fat (g/kg) 92.0 93.0 90.0 180.0 – 340.0
ponic tank without plants (6.3 mg/l). Mant et al. (2003)
Digestible energy (MJ/kg) 4.0 4.0 4.0 12.2 – 12.3
achieved 90.6% phosphorous removal using S. viminalis Crude protein (g/kg) 262.5 281.3 262.5 40.0 – 80.0
grown in gravel hydroponic system to treat primary settled Macroelements (g/kg)
sewage wastewater. Phosphorus 7.5 7.9 9.0 7.8 – 8.2
Potassium 2600 3500 2300 650 – 1170
Calcium 2.89 2.48 3.40 10.4 – 17.9
6.1.5. Potassium
Sodium 1.07 2.85 3.24 6
There was no potassium released from seeds during the Chloride 400 2100 200 10 – 5700
seed germination period. The potassium concentration in Microelements (mg/kg)
each compartment decreased with time during the growth Magnesium 2610 3060 2870 2500 – 2900
period and was dependent on the quantity and type of Manganese 98 40 37 102 – 287
Iron 144 225 175 252 – 456
seeds (Table 11). About 99.6– 99.8% of the potassium in
Copper 6 6 6 5.91 – 143.09
the wastewater was removed from the system. Dontje and Zinc 1720 1040 299 190 – 483
Clanton (1999) reported 25 – 71% potassium removal in Boron 0.6 0.6 0.6 0.07 – 4.50
recirculating aquacultural systems using cattails, reed ca-
nary grass and tomatoes grown in sand beds. Mant et al.
(2003) achieved 24.9% potassium removal using S. vim- amount of carbohydrates is nevertheless required to achieve
inalis grown in gravel hydroponic system to treat primary proper physical characteristic of the feed (Bureau and Cho,
settled sewage wastewater. The high removal efficiency of 2003). The carbohydrates content in the plants ranged from
potassium observed in this study could be attributed to 270.3 (oat) to 330.8 g/kg (rye) on dry weight basis, which
plant uptake, as well as to the precipitation of potassium in were within the limits for typical carbohydrate content in
the form of K2S. The wastewater was rich in potassium fish feed (300 – 330 g/kg).
and the application rate was based on phosphorous re- Fats are high-energy nutrients that can be utilized to
quirement by plant, which resulted in potassium over partially substitute for protein in aquaculture feeds (Craig
fertilization for all plants (Table 4). Marschner (1988) and Helfrich, 2002). Fats have many roles, including energy
indicated that when potassium supply is abundant, ‘‘luxury supply, structure formation and as precursors to many
consumption’’ of potassium often occurs, which may have reactive substances (Bureau and Cho, 2003). Fats supply
possible interference with physiological availability and about twice the energy as proteins and carbohydrates. The
uptake of magnesium and/or calcium. The elemental anal- digestible energy level in these plants of 4 MJ/kg was lower
yses performed on the plant tissues showed lower calcium than the recommended level of 12.3 MJ/kg. The crude fat
content. Although potassium is highly mobile in the concentrations in the plants ranged from 90 (rye) to 93 g/kg
growth medium and within the plant system, it can interact (barley) on dry weight basis, which is lower than the limits
with sulfur in the wastewater to form K2S, which is an for typical fat content in the fish feed (180 – 340 g/kg). All
insoluble form of potassium (the sulfur content of the fish display reduced growth and poor food conversion
wastewater was 46 mg/l). efficiency when fed diets deficient in fats (Tacon, 1985).
These plants must therefore be supplemented with a fat-
6.2. Nutritional value of the plants as fish feed containing material to bring the digestible energy level to
that recommended in fish feed.
Both alfalfa and clover were infected with fungus and Proteins are composed of 50% carbon, 16% nitrogen,
cannot, therefore, be recommended for use to reduce the 21.5% oxygen, and 6.5% hydrogen (Tacon, 1985). Proteins
pollution potential of aquaculture wastewater and as fish are required in the diet to provide indispensable amino acids
feed. The nutritional values and the elemental composition and nitrogen for synthesis of nonindispensable amino acids
of the fungus uninfected plants (oat, barley and rye) are (Bureau and Cho, 2003). The protein content in the plants
shown in Table 12. Although the results showed significant were in the range of 262.5 (oat and rye) to 281.35 g/kg
reduction in the pollution potential and the potential use of (barley) on dry weight basis, which is higher than the limits
these plants as fish feed, they need to be supplemented with for a typical protein concentration in fish feed (40 –80 g/kg).
some nutrients (Fat, Ca, Na, Mn and Fe). Fish are capable of using a high-protein diet, but most of the
Carbohydrates (starches and sugars) are the most eco- excess protein will be lost to the environment. Most nitro-
nomical and inexpensive sources of energy for fish diets gen is excreted as ammonia (NH3) by the gills of fish, and
(Craig and Helfrich, 2002). Carbohydrates only represent a only 10% is lost as solid wastes (Tacon, 1985).
minor source of energy for fish. Although most of the fish Inorganic elements (minerals) are required by fish for
species have a poor ability to utilize carbohydrates, a certain skeletal formation, maintenance of colloidal systems, regu-
12 A.E. Ghaly et al. / Environment International 31 (2005) 1–13
lation of acid-base equilibrium and for important com-
pounds such as hormones and enzymes (Bureau and Cho,
2003; Watanabe et al., 1997). They can be divided into two
groups (macrominerals and microminerals) based on the
quantity required in the diet and the amount present in fish.
Common macrominerals are phosphorous, potassium,
calcium, sodium and chloride. These minerals regulate
osmotic balance and aid in bone formation and integrity
(Bureau and Cho, 2003). Phosphorous deficiency reduces
growth and causes bone demineralization, skeleton defor-
mity, abnormal calcification of ribs and the soft rays of the
pectoral fin, cranial deformity and increased visceral fish,
whereas potassium deficiency reduces growth and causes
anorexia, convulsion and death (Tacon, 1985). Inadequate
calcium concentration in fish feed results in reduced growth
(Tacon, 1985). Sodium stimulates ATPase activity of gill
microsomes and regulates osmotic balance (NRC, 1993;
Craig and Helfrich, 2002). Chlorine is required to regulate
osmotic balance and acid in bone formation and integrity
(Craig and Helfrich, 2002). The phosphorous, potassium
and chloride concentrations were within the limits for a
typical fish feed. Calcium content was in the range of 2.48–
3.40 g/kg on dry weight basis, which was lower than the
calcium content in the typical fish feed (10.4 –17.9 g/kg).
The lower calcium content in plants was caused by the high
potassium content in wastewater, which made calcium non-
bioavailable (Marschner, 1988). Sodium content in the
plants was in the range of 1.07 – 3.24 mg/kg on dry weight
basis, which was also lower than the limit for a typical fish
feed (6.0 g/kg). Both calcium and sodium could be added
during the formulation of plant material into fish feed in the
forms of chlorides since the chloride content in the plant
tissues was in the lower side.
Microminerals (trace minerals) are magnesium, manga-
nese, iron, copper, zinc, and boron, which are required in
small amounts as components in enzyme and hormone
systems (Craig and Helfrich, 2002). Magnesium deficiency
in fish diets causes caclinosis, an abnormal condition in
which calcium salts are deposited in kidney tissues (Bureau
and Cho, 2003). Manganese is necessary for the normal
functioning of brain and for proper lipid and carbohydrate
metabolism. Manganese has two rules: first, as a cofactor for
enzymes, which form metal –enzyme complexes, and sec-
ond, as an integral part id metalloenzymes. An inadequate
supply of manganese results in reduced intake of food, loss
of equilibrium, depressed growth and increased mortality
rate in the young fish (Watanabe et al., 1997). Iron has an
active part in oxidation/reduction reactions and electron
transport associated with cellular respiration. It is found in
complexes bound to proteins, in enzymes and in nonhaem
compounds. Deficiency of iron induces anemia and affects
the hemoglobin level and the hatching rate of eggs (Tacon,
1985; Bureau and Cho, 2003; Watanabe et al., 1997).
Copper is important for fish, as it is involved in the activity
of enzymes such as cytochrome oxidase, superoxide dis-
nutase and tyrosinase. In addition, copper, proteins and
chelates also have metabolic roles (Watanabe et al., 1997).
Boron is essential for the embryonic development of all fish,
and boron deficiency causes development disruption and
death (Eckhert, 1998; Rowe and Eckhert, 1999). Magne-
sium, copper and boron were within the limits for a typical
fish feed. Zinc content was above the limit for the typical
fish feed. High concentration of zinc in the feed may reduce
fish growth rate (Tacon, 1985). Manganese and iron con-
tents were lower than the limits for a typical fish feed.
Therefore, when formulating the plants into fish feed, the
formula must be supplemented with manganese and iron.
7. Conclusion
Of the five plants used in this study, three (rye, barley
and oat) have the ability to reduce the pollution potential of
aquaculture wastewater and have the potential use as fish
feed. Clover and alfalfa were infected with fungus shortly
after germination, and their roots were completely destroyed
by day 14. Oat, barley and rye had the fastest growth and
showed much better resistance to fungal disease compared
with alfalfa and clover. Although substantial amounts of
soluble and insoluble substances were released by seeds
during germination, the plants were able to remove all
pollutants in wastewater and significant portions of those
released substances. The total reductions in the solids, COD,
NO3 – N, NO2 –N, phosphate and potassium ranged from
54.7% to 91.0%, 56.0% to 91.5%, 82.9% to 98.1%, 95.9%
to 99.5%, 55.5% to 93.6% and 99.6 to 99.8%, respectively.
This study indicated that oat, barley and rye can grow in this
type of hydroponic system and could be used as a fish feed
after being supplemented with fat, Ca, Na, Mn and Fe. Oil
seeds and the chlorides of these elements could be added to
the plants when formulating the fish feed. For a continuous
operation, a two-unit production system could be configured
to allow for one week germination and one week cleaning
and startup in one unit while the other unit is in operation.
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