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Seasonal Variations and Aeration Effectson Water Quality Improvements andPhysiological Responses of NymphaeaTetragona GeorgiXiao-Ming Lu a , Peng-Zhen Lu b , Min-Sheng Huang c & Ling-Peng Daid

a Institute for Eco-environmental Sciences , Wenzhou VocationalCollege of Science & Technology , Wenzhou , Chinab Faculty of Civil Engineering and Architecture , Zhejiang Universityof Technology , Hangzhou , Chinac School of Resources and Environment Sciences , East China NormalUniversity , Shanghai , Chinad College of Life and Environmental Science , Wenzhou University ,Wenzhou , ChinaAccepted author version posted online: 19 Sep 2012.Publishedonline: 04 Dec 2012.

To cite this article: Xiao-Ming Lu , Peng-Zhen Lu , Min-Sheng Huang & Ling-Peng Dai (2013) SeasonalVariations and Aeration Effects on Water Quality Improvements and Physiological Responses ofNymphaea Tetragona Georgi, International Journal of Phytoremediation, 15:6, 522-535, DOI:10.1080/15226514.2012.716103

To link to this article: http://dx.doi.org/10.1080/15226514.2012.716103

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International Journal of Phytoremediation, 15:522–535, 2013Copyright C© Taylor & Francis Group, LLCISSN: 1522-6514 print / 1549-7879 onlineDOI: 10.1080/15226514.2012.716103

SEASONAL VARIATIONS AND AERATION EFFECTS ONWATER QUALITY IMPROVEMENTS AND PHYSIOLOGICALRESPONSES OF NYMPHAEA TETRAGONA GEORGI

Xiao-Ming Lu,1 Peng-Zhen Lu,2 Min-Sheng Huang,3 andLing-Peng Dai41Institute for Eco-environmental Sciences, Wenzhou Vocational College of Science& Technology, Wenzhou, China2Faculty of Civil Engineering and Architecture, Zhejiang University of Technology,Hangzhou, China3School of Resources and Environment Sciences, East China Normal University,Shanghai, China4College of Life and Environmental Science, Wenzhou University, Wenzhou, China

Seasonal variations and aeration effects on water quality improvements and the physiologicalresponses of Nymphaea tetragona Georgi were investigated with mesocosm experiments.Plants were hydroponically cultivated in six purifying tanks (aerated, non-aerated) andthe characteristics of the plants were measured. Water quality improvements in purifyingtanks were evaluated by comparing to the control tanks. The results showed that continuousaeration affected the plant morphology and physiology. The lengths of the roots, petioles andleaf limbs in aeration conditions were shorter than in non-aeration conditions. Chlorophylland soluble protein contents of the leaf limbs in aerated tanks decreased, while peroxidaseand catalase activities of roots tissues increased. In spring and summer, effects of aeration onthe plants were less than in autumn. Total nitrogen (TN) and ammonia nitrogen (NH4

+-N)in aerated tanks were lower than in non-aerated tanks, while total phosphorus (TP) anddissolved phosphorus (DP) increased in spring and summer. In autumn, effects of aerationon the plants became more significant. TN, NH4

+-N, TP and DP became higher in aeratedtanks than in non-aerated tanks in autumn. This work provided evidences for regulatingaeration techniques based on seasonal variations of the plant physiology in restoring pollutedstagnant water.

KEY WORDS: aeration, hydrophytes, plant morphology and physiology, water quality,seasonal variation

INTRODUCTION

Increased industry and urban populations have led to the pollution of most urbanrivers throughout China, especially the urban stagnant flow channels (Gao et al. 2008).The dissolved oxygen (DO) concentrations of the stagnant flow channels have almost

Address correspondence to Xiao-Ming Lu, Institute for Eco-environmental Sciences, Wenzhou VocationalCollege of Science & Technology, Wenzhou 325006, China. E-mail: xm155@sina.com

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SEASONAL VARIATIONS AND AERATION EFFECTS ON WATER QUALITY 523

disappeared and few aquatic animals and plants survive due to the poor self-purificationof the channels as a result. The lack of DO affects the degradation of the pollutants inwastewater (Ouellet-Plamondon et al. 2006). Therefore, the degradation of pollutants instagnant water can be considerably improved by restoring DO levels.

Artificial aeration was developed as an approach for the recovery of DO of severelypolluted stagnant water and the ecological restoration of polluted stagnant flow chan-nels (Zhang et al. 2010; Gabriel et al. 2009). This technique involves pumping atmo-spheric air into polluted water, which encourages the increase and recovery of DO lev-els. Thus, the pollutants were purified and the water quality improved (Cristina et al.2009; Gagnon et al. 2007). Engineering practices have also demonstrated that the aer-ation technique is effective in treating wastewater (Guo et al. 2008; Zhang et al.2010).

However, practical experiences demonstrated that when polluted stagnant water wasaerated, except for the water quality improvement (Ouellet-Plamondon et al. 2006), pres-surized convective airflow from the aerator affected the plant morphology and physiologyunder certain conditions, such as a rate of gas flow that was too rapid, a lack of distancebetween the aerator and the planting site, and a duration of aeration was too long. Thepressurized convective airflow from the aerator is most likely to affect the exposed roots ofhydrophytes, and injure the vulnerable ones in particular. The aeration procedure createsnumerous surfactant bubbles, which cover the plant leaves. Consequently, the plants nearthe aerator can be small. It is not beneficial for the long-term of water quality improvements.

Most of the literature studies on aquatic plant treatments in wastewater report pu-rification efficiencies (Francesco and Luigi 2009; Zurita et al. 2011; Cristina et al. 2012),rather than the correlation between the purification process and the physiological responseof hydrophytes. A few reports exist on the effects of aeration on the morphology and physi-ology of hydrophytes (Zhao et al. 2011). However, knowledge on that aspect is important toeffectively utilize the combined technique of hydrophytes and aeration technique that cantreat and restore severely polluted stagnant channels. Therefore, an analysis and evaluationof the effects and mechanisms of aeration on plant growth and the purification of the riverwater has important theoretical meaning to the combined system and operation regulationfor optimizing the treatment practice.

The physiological status of the aquatic plant is an important factor for nitrogen(N) and phosphorus (P) absorption and the removal of polluted water (Li et al. 2007).Soulwene et al. (2009) analyzed the effects of a hydrophytes life cycle on the performancesof a constructed wetland treating domestic wastewaters, and found that the growth rateof hydrophytes (reeds and cattails) was correlated with purification efficiencies. Enzymeactivities of the plant tissues are associated with the physiological status of the plants.Catalase (CAT) and peroxidase (POD) are major components of the antioxidant enzymesystem in the cells of the plant (Martı et al. 2009). Previous studies have shown that POD isa good proxy for cell stress when cells participating in physiological activities are exposedto adverse conditions (Tan et al. 2006). POD is significantly associated with the resistanceof plants to adverse conditions (Ya et al. 2011).

A few reports exist on the treatment of polluted river water using Nymphaea tetragonaGeorgi (Lu and Chen 2012). N. tetragona is a perennial floating plant that belongs toNymphaea and Nymphaeaceae. The plant grows best in strong sunlight, germinates inMarch, and withers in November. N. tetragona is common and widespread in pollutedchannels. N. tetragona has ornamental value, along with a good purification capacity, buthas not yet been extensively used in environmental practice.

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524 X.-M. LU ET AL.

In this study, N. tetragona was utilized to treat severely polluted stagnant water fromthe Gongye channel by mesocosm experiments. We measured morphological and physio-logical differences between the plants grown under aeration and non-aeration conditions.The morphological metrics included root, petiole, and leaf limb length, root density, andtillers. Physiological metrics included POD and CAT activity, SP, Chl-a and Chl-b, N andP content, and biomass. We hypothesized that under aeration conditions where the waterquality improvement is enhanced, the growth of hydrophytes would also increase due to thedecline of the stress of polluted water. We demonstrate that an extended period of contin-uous aeration exerts a role for the water quality improvement, increasing the water qualityimprovement while not increasing the plant growth, however influencing the plant mor-phology and physiology. The objective was to provide evidence for the use of N. tetragonaand an optimization of the aeration technique based on the seasonal variation of the plantduring the ecological restoration practice of polluted stagnant water.

MATERIALS AND METHODS

River Location and Characterization

Gongye channel in Pu-tuo district of Tao-pu town (latitude 31◦11′N and longitude121◦29′E), Shanghai city has a smooth riverbed. It is stagnant and severely polluted all yearround. Concentrations of pollutants indicators, such as chemical oxygen demand COD),total nitrogen (TN) and ammonia nitrogen (NH4

+-N) are above national limits for irrigationwater. It is of average approximate 6–7 m width and 2–3 m depth. The DO of the channel ismaintained at approximately 0.1 mg L−1 all year round. Pollutants are mainly in the formof midstream domestic sewage discharge, with some upstream industrial effluent.

Device Construction and Plant Cultivation

A test apparatus consisting of eight purifying tanks and one “reservoir” tank was usedas the experimental system. The device (Figure 1) was composed of one “reservoir” tank(upper dimension, length 150 cm × width 100 cm; downside dimension, length 145 cm× width 95 cm; height, 60 cm) and eight plastic purifying tanks (upper dimension, length124 cm × width 62 cm; downside dimension, length 115 cm × width 55 cm; height,76 cm). An overflow tube on the “reservoir” tank kept a constant water level of 60 cmand ensured that each purifying tank received the same volume of water. River water wascontinuously pumped with a 25SFBX-13D water pump (Shanghai Boyu Banye Limited,Shanghai, China) through a plastic pipe (diameter, 4 cm) into the “reservoir” tank directly,through the fixed effluent tube and into the purifying tanks. A wire mesh of 0.2-mm porediameter placed at the water outlet of the “reservoir” tank prevented suspended matter in theriver water entering the purifying tanks. The effluent tube on the purifying tank monitoredthe water level in the tank at 60 cm. Each purifying-tank was separated by a baffle throughthe center, and approximately 20 cm space to the tank bottom to avoid causing water circuitflow. To control the effects of the pollutants released from the sediment on the water qualityimprovements, each purifying tank had a sediment exit on the bottom to discharge thesediment mud daily.

Continuous water flow was kept in each tank. Hydraulic retention time was 8 h. AnACO-004 electromagnetic atmosphere pump (60 L min−1, Zhejiang Sensen Shiye Limited,Hangzhou, China) was employed to aerate four tanks continuously from February 26, 2008to October 16, 2008. The gas flow rate in each of the four tanks was 30 L min−1. Seedlings

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SEASONAL VARIATIONS AND AERATION EFFECTS ON WATER QUALITY 525

Figure 1 The test apparatus consisting of eight purifying tanks and one “reservoir” tank. Notes: CK and CKarepresented the non-aerated tank and aerated tank, NK and NKa represented the non-aerated tank and aeratedtank.

of N. tetragona (uniform size: height = 10 cm) were collected from Shanghai ZelongBiology Engineering Limited (Shanghai, China). The seedlings taken to test were firstwashed carefully with the river water several times to remove the mud on the roots. Therewas no acclimatizing period for the seedlings before starting the experiment. No othergrowth medium but the river water was utilized to cultivate the seedlings directly. Then, theseedlings were hydroponically cultivated in purifying tanks with the same planting densityof 10 plants per tank on February 26, and two additional tanks (aerated, non-aerated) wereset up without plants to act as control (CK). The treatment period was from February 26,2008 to October 16, 2008. The sampling was performed in summer (on July 10, 15, and20 2008), and in spring (on April 16, 21, and 26 2008) and autumn (on October 6, 11, and16 2008). During the long-term treatment, no other medium but the river water was utilizeddirectly and the withered leaf limbs of the plants in the purifying tanks were removed fromthe tanks in time. There were no other human influencing factors and the plants grew inthe tanks. The temperature was not controlled. The field experiment was conducted in ariparian area 10 m away from the Gongye channel.

Sampling and Analysis

On July 10, 2008 (air temperature: 21◦C–29◦C, sunny), five plants (each plant hadtwo or more leaf limbs) were collected at random from the plants in every purifying tank.The root (taproot) lengths, petioles lengths and leaf limb (upper leaf limb) lengths weremeasured and standard deviations were calculated. Healthy leaf limbs (0.40 g) and roots(0.40 g) were collected from each one of the selected plants for use. Chemical composition

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526 X.-M. LU ET AL.

of the water in the tanks varied due to the seasonal effect of temperature, but it did notaffect the comparative analysis of the water quality improvements and the plant physiologyresponses in different purifying tanks, since each tank received a same seasonal effect oftemperature in the long-term treatment. Therefore, the seasonal effect of the temperatureon the water chemical features should not be particularly evaluated. The previous testindicated that in the non-aerated control tank, the effluent water quality was almost the sameas that of the inlet water, while the inlet water quality of the control tank was the same as thatof each purifying tank. Therefore, water analysis was done in the effluent water and the inletwas not sampled, and the water quality improvement in each purifying tank was evaluated bycomparing to the effluent water quality of the control tank. Corresponding water analysis ofthe effluent from each of the eight tanks was done using standard methods (APHA 1998).Three replicates of water samples were taken from each tank. DP represents dissolvedphosphorus. Total phosphorus (TP) includes the dissolved and particulate phosphorus. Themineral phosphorus was measured. The tiller and root density of the plants in each tankwas measured.

Enzyme Solution Preparation, Soluble Protein Content, and Enzyme

Activities Measurement

Fresh roots (0.40 g) and leaf limbs (0.40 g) were prepared and refined in the freezingphosphate solution (pH 7.8), centrifuged at 13000 r min−1 and in 4◦C for 30 min. Thesupernatant was collected for the steps below.

The soluble protein (SP) content of the leaf limb of the plant was measured withCoomassie Brilliant Blue (Wessel and Flugge 1984). Standard curve was reported withusing bovine serum albumin at 550 nm. Catalase (CAT) activity in the root was performedby UV-spectrometer analysis at the 240 nm (Rao et al. 1996).

Peroxidase (POD) activity measure of the root of the plant was performed withguaiacol at the 460 nm (Zhang 1990). The variation of the light intensity in per minrepresented the enzyme activity.

Chlorophyll Content Analysis

The method reported by Hegedus et al. (2001) was employed to analyze chlorophyll(Chl) content at the 663 nm and 645 nm. It was modified as follows: 0.05 g leaf and 5 mLof 80% acetone solution was soaked for 24 h and the absorbance of the soaked solutionwas measured with a U-1100 spectrophotometer (Hitachi Ltd., Tokyo, Japan).

Plant Biomass, Nitrogen and Phosphorus Contents

Fresh plant samples were placed in a forced-air oven, killed for 1 h at 105◦C, anddried for 48 h at 80◦C until they reached a constant weight. The biomass (dry weight) of theplant tissues, including the roots and the shoots, was measured and the means of individualplants obtained. The dried plant samples were then ground to a size of 80 meshes and storedfor chemical analysis.

Nitrogen (N) contents in the samples of plant tissues including the roots, petiolesand leaf limbs were determined by the Dumas combustion method using an automatedCN analyser (LECOCHN-1000, LECO Company, St Joseph, MI, USA). Phosphorus (P)contents in the corresponding plant tissues were determined by the vanado-molybdatemethod using a U-1100 spectrophotometer (Hitachi Ltd., Tokyo, Japan) after digestingmaterial in a mixture of concentrated nitric and perchloric acids (Vogel and Bassett 1978).

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SEASONAL VARIATIONS AND AERATION EFFECTS ON WATER QUALITY 527

Statistical Analysis

Morphological and physiological parameters (root, petiole, and leaf limb length, rootdensity, and tillers; POD and CAT activity, SP, Chl-a and Chl-b, N and P content, andbiomass) of five plants of each purifying tank were measured. The physical and chemicalparameters of polluted water in every season were obtained. All data presented were themeans of at least three replicates and were reported as mean value ± standard deviation(S.D.). Significance of differences of samples was calculated by independent-samples t-tests of SPSS 15.0 software. Results of testing were considered significant if calculatedp-values were ≤0.05. Pearson correlation coefficient (r) between the TN concentrations ofpolluted water and the N contents of the plants was calculated by SPSS 15.0 software.

RESULTS AND DISCUSSION

Effects of Aeration on Water Quality Improvements of the Tanks in

Various Seasons

As shown in Table 1, chemical oxygen demand (COD), total nitrogen (TN) andammonia nitrogen (NH4

+-N) were lower by decreasing 9.10 mg L−1, 2.67 mg L−1 and2.01 mg L−1 in aeration than in non-aeration tanks in summer, but TP and DP were higherby increasing 0.025 mg L−1 and 0.026 mg L−1 in aerated tanks. However, COD, TN,NH4

+-N, TP and DP were higher by increasing 2.31 mg L−1, 0.81 mg L−1, 0.58 mg L−1,0.043 mg L−1, and 0.018 mg L−1 in the aeration than in the non-aeration tanks in autumn.There was no statistical significance for COD (p = 0.41), TN (p = 0.13), and NH4

+-N (p =0.10) between aeration and non-aeration conditions in autumn. It differed from the reportson Pontederia cordata by Zhao et al. (2011). They reported that the concentration of COD,TN and NH4

+-N in aerated tanks was lower than that in non-aerated tanks during autumn,respectively. In the aerated control tanks the dissolved oxygen (DO) concentration waslower than in the aerated tanks seeded with N. tetragona. The additional DO concentrationin aerated tanks was resulted from the O2 release of the plant roots. The plants releasedoxygen during the photosynthesis process and resulted in the DO concentration increasing(Luo et al. 2006).

Water quality improvements between treatments in terms of percentages were dif-ferent. For instance, TN was reduced by 25.6% in aerated tanks and 20.5% in non-aeratedtanks during spring. During the summer sampling TN was reduced by 45.0% in aeratedtanks and 38.1% in non-aerated tanks. In contrast during autumn, a smaller reduction inTN was observed in aerated tanks, than in non-aerated tanks (27.0% compared to 33.1%).The p-value of t-test for TN removal efficiencies between aeration and non-aeration condi-tions was 0.06, 0.05, and 0.07 in spring, summer, and autumn, respectively. Therefore, thecombined role of plants and aeration on water quality improvements was more effective inspring and summer than in autumn.

Effects of Aeration on the Morphology and Physiology of N. tetragona

in Various Seasons

As shown in Table 2, continuous aeration affected the morphological and physiolog-ical characteristics, including root, petiole and leaf limb length, root density, tillers, PODand CAT activity and SP content of N. tetragona. Aeration affected the absorption of theN and P by the plant roots from polluted water, while dissolved N and P are important

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Tabl

e1

Eff

ects

ofae

ratio

non

wat

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ality

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sin

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Mea

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OD

(mg

L−1

)T

N(m

gL

−1)

NH

4+ -

N(m

gL

−1)

TP

(mg

L−1

)D

P(m

gL

−1)

DO

(mg

L−1

)pH

WT

(◦C

)

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49.8

2±

2.50

a21

.79

±1.

08a

13.6

0±

0.67

a0.

206

±0.

012a

0.04

7±

0.00

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35±

0.10

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0.1a

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±0.

1aSp

–54

.74

±2.

71b

23.9

8±

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±0.

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0.18

2±

0.01

1b0.

042

±0.

002b

0.92

±0.

04b

8.2

±0.

1a21

.2±

0.1a

CK

+67

.42a

29.3

0a18

.39a

0.31

5b0.

063b

1.43

b8.

4a23

.3a

CK

–69

.60a

30.1

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0.22

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050a

0.12

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4a23

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46.5

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±0.

82a

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71a

19.3

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233b

0.21

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50b

8.5a

34.6

a

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–92

.30a

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144a

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±0.

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0.1a

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.81a

17.5

9a11

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0.20

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102b

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4a27

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–76

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.

528

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Tabl

e2

Eff

ects

ofae

ratio

non

mor

phol

ogic

al(l

engt

hof

root

,pet

iole

and

leaf

limb,

tille

rsan

dro

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)an

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cal(

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,Chl

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ivity

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Dan

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flim

bT

iller

Roo

tden

sity

Mea

ns(m

gg−

1 )(m

gg−

1 )(m

gg−

1 )(U

/g·m

in)

(mg/

g·min

)(c

m)

(cm

)(c

m)

(/m

2 )(/

m2 )

Sp+

33.2

2±

13.3

0a1.

50±

0.31

a0.

41±

0.08

a0.

26±

0.10

a2.

63±

1.05

a13

.9±

1.5a

11.7

±1.

4a3.

1±

0.3a

51±

3a19

8±

10a

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43.8

3±

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6a1.

80±

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52±

0.10

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15±

0.05

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11±

1.06

a18

.4±

1.9b

15.2

±1.

7b4.

0±

0.4b

66±

4b25

9±

13b

Su+

25.5

1±

9.81

a1.

13±

0.18

a0.

34±

0.07

a0.

31±

0.13

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530 X.-M. LU ET AL.

nutrient sources for the development of hydrophytes. Therefore, the physiology and growthof hydrophytes were influenced by the aeration. In particular, the plant height was smallerand the Chl content of the leaf limbs decreased in aerated tanks, while the activity of PODand CAT of the root tissues increased during all three seasons (spring, summer and autumn).In summer, there was no statistical difference for the activities of POD (p = 0.06) and CAT(p = 0.08) between aeration and non-aeration conditions, respectively. The decrease inplant tillers led to the decline of the root density under aeration conditions, differed fromthe reports on P. cordata by Zhao et al. (2011). They reported that the increase in P. cordatatillers caused the increase of the root density in aeration tanks, indicated that the effects ofaeration conditions on N. tetragona were greater than on P. cordata. Effects of aeration onN. tetragona were correlated with the growth state of the plant. In spring and summer, theplant was at the starting growth stage and strong growth stage, and the aeration affected theplant less.

The physiological variation of the plants between aeration and non-aeration tankswas less in spring and summer than in autumn. For instance, the p-value of t-test for thePOD activity of the plant roots between aeration and non-aeration conditions was 0.02,0.06, and 0.01 in spring, summer and autumn, respectively. The plant was severely affectedby the aeration in autumn, and the physiological variation of the plant between aerationand non-aeration tanks was more significant.

Effects of Aeration on the N and P Contents of N. tetragona in Various

Seasons

As shown in Table 3, continuous aeration affected the biomass and the N and Paccumulations of N. tetragona during three seasons (spring, summer, and autumn). Inparticular, in aerated tanks, the N and P content and biomass of the plant decreased,and both differences were significant (p < 0.05). In spring and summer, the gap of theN and P content and biomass of the plant between aeration and non-aeration tanks wasnarrower. In autumn, however, these gaps between the aeration and non-aeration tanksbecame wider, indicating the greater influence of aeration on the plant. It agreed to thereports on P. cordata by Zhao et al. (2011). These gaps between during spring and summerdid not vary significantly, but the differences were significant in comparison to duringautumn (p < 0.05). But during autumn water N concentration did not decrease greatlyby aeration as that in spring and summer. The decrease of water N concentration byaeration was 8.36 mg L−1, 14.70 mg L−1 and 5.14 mg L−1 in spring, summer and autumn,respectively.

Table 3 Effects of aeration on the N and P contents and the biomass of N. tetragona in various seasons.

Means Nitrogen (g kg−1) Phosphorus (g kg−1) Biomass (g plant−1) Water content ratio (%)

Sp+ 5.67 ± 0.26a 0.82 ± 0.04a 6.62 ± 0.31a 95.9 ± 2.4a

Sp– 7.04 ± 0.31b 1.13 ± 0.06b 9.65 ± 0.47b 94.7 ± 2.2a

Su+ 14.24 ± 0.63a 1.74 ± 0.09a 17.23 ± 0.83a 92.8 ± 2.1a

Su– 16.68 ± 0.82b 2.13 ± 0.11b 22.57 ± 1.12b 92.1 ± 2.0a

Au+ 15.33 ± 0.74a 1.90 ± 0.09a 18.49 ± 0.95a 92.4 ± 1.7a

Au– 19.96 ± 1.01b 2.42 ± 0.13b 25.64 ± 1.26b 91.6 ± 1.6a

Notes: the type of statistical test was an independent samples t-test in SPSS 15.0.

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SEASONAL VARIATIONS AND AERATION EFFECTS ON WATER QUALITY 531

In spring and summer, effects of aeration on the plant morphology and physiologywere comparatively less (Table 2). In aerated tanks, the water N concentration was lowerthan in the non-aerated tanks (Table 1). In autumn, aeration severely affected the plantphysiology (Table 2), in which reduced the water N concentration in non-aerated morethan in aerated tanks (Table 1). The seasonal variation of the water N concentration inthe aerated and non-aerated tanks mainly resulted from the seasonal variation of the surfacearea of the plant roots in the aerated and non-aerated tanks. In spring and summer, thegap of the surface area of the plant roots between the aerated and non-aerated tanks wasnarrower in comparison to that in autumn (Table 2). As the surface area of the plantroots in the non-aerated tanks should largely exceed that in aerated tanks in autumn,this most probably led to the seasonal variation of the rhizospheric microorganisms. Ithas been suggested that plant rhizosphere enhances microbial density by providing rootsurface for microbial growth and a micro aerobic environment via root oxygen release (Brix1997; Gagnon et al. 2007). Andrews and Harris (2000) reported that the plants protectedbacteria from the surrounding medium by providing a surface for attachment as well asnutrients. The seasonal variation of the purification effects of the bacteria on pollutedwater led to a corresponding change in the N concentration in the tanks (Zhang et al.2009).

Analysis of the results reveals that the water quality improvement in the purifyingtanks was correlated with the seasonal variation of the morphology and physiology ofthe plants in the tanks. Soulwene et al. (2009) also reported that the seasonal variation inthe hydrophytes (reeds and cattails) growth rate was associated with the performance ofwastewater treatment. The growth of hydrophytes is usually determined by the physico-chemical characteristics of water quality, sediment properties and hydrological conditions(Fan and Li 2005). This experiment was conducted to investigate hydroponic cultures ofN. tetragona without interference from sediment disturbance while the microhydrologicalconditions of the purifying tanks were changed by continuous pressurization of convectiveairflow from the aerator. Aeration affected the growth of N. tetragona, which altered themorphological characteristics of N. tetragona and induced physiological responses, whichin turn affected water quality improvements.

Water currents resulting from aeration affected the growth of the plant roots, resultingin a decrease in both the root length and the root density. The N and P absorption frompolluted water by the roots were subsequently affected. The employed plant did not makephotosynthesis with underwater organs. The results showed that the N and P contents ofthe plant in aerated tanks were lower than in non-aerated tanks. This indicates that in thesame period of time, the N and P absorption from polluted water by the plant was less inthe aerated tanks than in the non-aerated tanks. Therefore, continuous aeration was mostlikely to have influenced the plant transpiration.

The POD and CAT activity of the plant was affected by the aeration. The pollutedwater adversity caused an increase of the superoxide radical in the plant cells and causedoxidation stress (Ya et al. 2011). The anti-oxidase functions of CAT and POD in the plantcells are able to cooperatively prohibit and eliminate the active oxygen radicals (Martıet al. 2009). In this experiment, water current scoured the plant roots continuously for along period, affected the growth status of the roots (and most likely injured young fibrousroots) and decreased N and P absorption from the water by the roots. The physiologicalactivity of the plant was then affected, leading to an increase in the activity of PODand CAT in the roots. This increase of the antioxidant enzymes activity of the plantresulted from the aeration effects, which differed from the report by Li et al. (2007). They

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suggested that the increase of POD activity of the roots of wetland plants after a long-termof soaking in polluted water resulted from the plant absorbing many soluble inorganicpollutants.

The continuous aeration resulted in corresponding changes in the water quality im-provements in the respective tanks. The purification efficiencies of the aerated and non-aerated tanks were correlated with the absorption of the water pollutants by the plants.For instance, in aerated tanks, Pearson correlation analysis revealed a negative correlation(r = –0.311, p = 0.234) between the water N concentration and the plant N content. Innon-aerated tanks, the water N concentration was negatively correlated (r = –0.303, p =0.272) with the plant N content. The plant was dependent on the roots to absorb N and Pfrom polluted water (Luo et al. 2006; Baldy et al. 2007; Vymazal 2011), which was corre-lated with the surface area of the roots, inner structure of the roots and the characteristicsof water quality (Fan and Li 2005).

The N and P contents in the plant tissue of N. tetragona subjected to aeration con-ditions were lower than those of plant tissue from N. tetragona subjected to non-aerationconditions. In summer, the N content of the plants grown in aerated tanks was 26.74 g, lessthan that of the plants grown in non-aerated tanks by decreasing 25.97 g (Tables 2–3), whilethe N removal efficiency of 44.97% from polluted water in aerated tanks was higher thanthat in non-aerated tanks by increasing 6.90% (Table 1). In autumn, the N accumulation of11.34 g from polluted water by the plants grown in aerated tanks was less than that by theplants grown in non-aerated tanks by decreasing 51.10 g (Tables 2–3), and the N removalefficiency of 27.0% from polluted water in aerated tanks was lower than that in non-aeratedtanks by decreasing 6.10% (Table 1).

The growth of plant roots was affected by the continuous aeration (Table 2). In springand summer, the root length in non-aerated tanks was 1.32 times and 1.30 times of theroot length in aerated tanks, respectively (Table 2). The root density was 1.31 times and1.28 times of that in aerated tanks in spring and summer, respectively (Table 2). Therefore,the surface area of the roots in non-aerated conditions probably exceeded that in aeratedtanks. Additionally, the N and P contents of the plants were higher in non-aerated conditionsthan in aerated tanks. Moreover, the airflow from the aerator influenced the sedimentation ofparticulate phosphorous and the adsorption of rhizosphere microorganisms by N. tetragonain the aerated tanks. Thus, TP and DP concentrations in non-aerated tanks were lower thanin aerated tanks during all three seasons. However, the NH4

+-N of polluted water was stillhigher in non-aerated than aerated tanks in spring and summer, which was because thehigher DO concentration was more suitable for the NH4

+-N oxidation by aerobic microbes(Kirk and Kronzucker 2005; Ouellet-Plamondon et al. 2006). Luo et al. (2006) reportedthat nitrogen absorption by harvested hydrophytes only accounted for approximately 17%of the nitrogen removal of polluted water, and that most nitrogen removal was performedby the denitrification of bacteria (Kowalchuk et al. 1998).

In autumn, aeration made the surface area of the plant roots in non-aerated tanksobviously exceeded those in aerated tanks. In a previous study it was shown that microbialgrowth on plants roots was correlated with the surface area provided by the plant roots(Andrews and Harris 2000). Kyambadde et al. (2004) also reported that microbes werepresent on roots as an attached biofilm and abundance was correlated with root surface.Therefore it is likely in this study that, continuous aeration greatly affected the microbedevelopment on the root zones during autumn. Additionally, in non-aerated tanks, the plantabsorption of water pollutants was more effective, reducing the COD, TN, NH4

+-N, TP,and DP in non-aerated tanks than in aerated tanks, respectively.

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SEASONAL VARIATIONS AND AERATION EFFECTS ON WATER QUALITY 533

Effects of the same aeration technique on the plant physiology were correlated withthe growth status of the plant. In particular, when the plant was at the growth stage in springand summer, effects of aeration on the plant was less. An increase in DO lowered the N inthe aerated tanks in comparison to the non-aerated tanks. In autumn, when the plant was atits final stage, effects of aeration on the plant was greater, which lowered the N and P in thenon-aerated tanks compared to those in the aerated tanks. In all, effects of aeration on plantphysiology were more significant in autumn than in spring and summer. The combinedrole of plants and aeration on water quality improvements was more effective in spring andsummer than in autumn.

CONCLUSIONS

An extended period of continuous aeration affected the growth of the Nymphaeatetragona Georgi plant. Physiological characteristics of the plant were also affected, whichled to increased POD and CAT activity of the plant root tissues. The Chl and SP contentof the leaf limbs declined, and the N and P accumulation of the plant and plant biomassdecreased. Effects of the same aeration technique on the plant were correlated with the plantgrowth status. In spring and summer, effects of aeration on the plant were comparativelyless, as were the variation of the physiological characteristics of the plants in the aeratedand non-aerated tanks. The COD, TN, and NH4

+-N in aerated tanks was lower than thatin non-aerated tanks, while TP and DP was higher in aerated tanks compared to non-aerated tanks. In autumn, effects of aeration on the plant were greater. The variation ofthe physiological characteristics of the plant between aerated and non-aerated tanks wassignificant. In the aerated tanks, the COD, TN, NH4

+-N, TP, and DP was higher than inthe non-aerated tanks. Therefore, in the ecological restoration projects of polluted stagnantwater bodies, it appears to be reasonable to regulate the aeration technique accordinglybased on the seasonal variation of plant physiology.

ACKNOWLEDGMENTS

This work was supported by the National Scientific and Technological Major Programof China (No. 2009ZX07317-006) and the natural science foundation of Zhejiang Province(LY12C03007). We gratefully acknowledge the School of Resource and EnvironmentSciences of East China Normal University for kindly providing facilities for this study.

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