Environmental Pollution 134 (2005) 385–395 www.elsevier.

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Amelioration of Indian urban air pollution phytotoxicity in Beta vulgaris L. by modifying NPK nutrients
Anoop Singha, S.B. Agrawalb,*, Dheeraj Rathorea
a

Laboratory of Air Pollution and Global Climatic Change, Department of Botany, Allahabad Agricultural Institute - Deemed University, Allahabad 211 007, India b Department of Botany, Banaras Hindu University, Varanasi 221 005, India Received 28 November 2003; accepted 17 September 2004

Air pollution caused adverse impact on growth and biomass accumulation of Beta vulgaris L. plants while higher fertility levels showed reduced yield losses.
Abstract Air pollution levels are increasing at an alarming rate in many developing countries, including India and causing a potential threat to crop production. Field experiments were conducted to examine the impact of urban air pollutants on biomass (yield) and some physiological and biochemical parameters of palak (Beta vulgaris L. var. All Green) that grew from germination to maturity at seven periurban sites of Allahabad city having different concentrations of air pollutants under different levels of nutrients. The 6 h daily mean NO2, SO2 and O3 concentrations varied from 2.5 to 42.5, 10.6 to 65 and 3.5 to 30.8 mg mÿ3, respectively at different locations. Levels of air pollution showed significant negative correlations with photosynthetic pigments, protein, ascorbic acid and starch contents and catalase activity of palak leaves. A significant negative correlation was found for total biomass with SO2 (r Z ÿ0.92), NO2 (r Z ÿ0.85) and O3 (r Z ÿ0.91) concentrations. The increased fertilizer application (N, P and K) over the recommended dose resulted in a positive response by reducing losses in photosynthetic pigments and total biomass. This study proved that ambient air pollution of Allahabad city is influencing negatively to the growth and yield of palak plants. Ó 2004 Elsevier Ltd. All rights reserved.
Keywords: Air pollution; Nutrients; Beta vulgaris; Biomass; Yield

1. Introduction The impacts of air pollution have long been recognized as major cause of losses in crop production in several developed countries. However, little attention has been paid in developing countries, including India, on potential impacts of air pollution on growth and productivity. The Indian national ambient quality data indicate that emissions of a range of air pollutants are generally increasing (Agrawal, 1998). The annual average of SO2
* Corresponding author. Tel.: C91 542 2368156; fax: C91 542 2368174. E-mail address: sbagarwaldr@sancharnet.in (S.B. Agrawal). 0269-7491/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.envpol.2004.09.017

concentrations ranged from 10.4 to 39.0 mg mÿ3 ppb in most parts of the country, while NO2 concentrations were found between 43.2 and 60.1 mg mÿ3 in metropolitan cities. Pandey et al. (1992) reported elevated concentrations of O3 in Varanasi city, an adjoining district of Allahabad, where significant negative influence of urban air pollutants was recorded on a variety of plant species growing in periurban areas (Agrawal et al., 2003). Urban air pollution has direct impact on periurban agriculture due to dispersion of pollutants in all directions along the wind. During transportation primary pollutants often form secondary pollutants, causing greater adverse effects on crop production in periurban areas. Effects of air pollutants have been

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A. Singh et al. / Environmental Pollution 134 (2005) 385–395

described in terms of foliar injury (Jacobson and Hill, 1970), reduction in photosynthetic pigments (Agrawal et al., 1982), inhibition of physiological processes (Saxe, 1991), alteration in metabolic functions (Malhotra and Khan, 1984), enzyme activities (Nandi et al., 1986) and nutrient uptake and suppression of growth and yield of agricultural crop plants (Lee, 2000; Verma et al., 2000; Ribas and Penuelas, 2003; Singh et al., 2003). Use of chemical protectants, such as growth regulators, antioxidants and fertilizers is suggested to be a short-term solution to reduce the risk of air pollution damage. Researchers have suggested that application of mineral nutrients promotes growth, and reduce pollutant induced injury to crops (Ormrod et al., 1973). Rajput and Agrawal (1994) have found that soybean plants grown at recommended fertility levels were less injured by SO2 in comparison to unfertilized crop. In view of the above, the present investigation was aimed to suggest an economical and ecofriendly solution to ambient air pollution induced damage, by altering the level of mineral nutrients in palak (Beta vulgaris var. All Green) plants grown at different periurban and urban sites of Allahabad city (Eastern Uttar Pradesh, India).

2. Materials and methods The study was performed in the periurban and urban environment of Allahabad city with a population of 0.85 million located in the eastern Gangetic plains of India between 24  47#N latitude and 82  21#E longitude and 96 m above mean sea level. During the study period mean minimum and mean maximum temperatures ranged between 14.9–24.3  C and 30.8–34.6  C, respectively (Table 1). The average relative humidity varied between 59.8 and 68.4% and wind speed 3.5–6.8 km hÿ1. Total precipitation was 164.2 mm during September and 77 mm during October (Table 1). Prominent wind was westerly. The plant species chosen for this study is a cheap and popular vegetable and consumed mainly as a source of iron in the diet. Periurban area of Allahabad provides 85% of the palak crop consumed in the city. An experiment was conducted from September to November 2001 at seven selected sites (viz. Allahabad Agriculture Institute (AAI), Civil lines (CL), Mehdeori (Mh), Jhunsi (Jh), Bahrana (Bh), Arail (Ar) and
Table 1 Meteorological data during the experimental period Month and year Precipitation Temperature (  C) Relative Wind (mm) humidity speed Max. Min. (%) (km hÿ1) 34.6 33.7 30.8 24.3 20.7 14.9 68.4 64.6 59.8 6.8 4.2 3.5

September, 164.2 2001 October, 2001 77.0 November, 2001 0.0

Rajrooppur (RRP)) on the periphery and within the city of Allahabad. The location of sites and a brief description of their characteristics are given in Fig. 1 and Table 2. The soil was prepared at one place by mixing garden soil and farmyard manure in 3:1 ratio following the normal agronomical practices for uniformity of edaphic conditions. Soil used in the experiment had pH 7.62, organic carbon 1.64%, N 690 mg 100 gÿ1 soil, P 16.4 mg 100 gÿ1 soil and K 136.2 mg 100 gÿ1. Palak var. All Green seeds were sown in pots (30 cm diameter) with four treatments of fertilizers, i.e. without fertilizer (F0), recommended dose (RD) of N, P and K (F1), one and half times of RD of N, P and K (F2) and two times of RD of N, P and K (F3) on September 26, 2001. Recommended doses of NPK were 80, 40, 40 kg haÿ1, respectively. Nitrogen was given in form of urea, phosphorus as single super phosphate and potassium as murate of potash. Half dose of nitrogen and full dose of phosphorus and potassium were given as basal dressing and another half of nitrogen as top dressing. After sowing, 32 pots were transferred to each site. Pots were placed in unshaded open area receiving uniform light. Micrometeorological variations in temperature were 0.1–0.2  C, relative humidity 1–3% between the sites. Light intensity was identical at all sites. The pots were uniformly watered throughout the experiment in order to maintain constant soil moisture. For analysis, triplicate random samples of plants from each treatment of each site were taken at 20 days after sowing (DAS) and then at regular intervals of 15 days. Final harvest was done on November 16, 2001 at 50 DAS. For total biomass determination, plants were oven dried at 80  C until the constant weight was obtained and values were expressed as g plantÿ1. The chlorophyll content was expressed as mg gÿ1 dry leaf and measured by using the method of Machlachlan and Zalik (1963). Carotenoid content was calculated by the method of Duxbury and Yentsch (1956). Protein analysis in fresh leaves was performed by using the method of Lowry et al. (1951). Ascorbic acid in fresh leaves was measured using the 2,6 dichlorophenol indophenol method of Keller and Schwager (1977). Catalase and peroxidase enzyme activities were determined using the methods of Kar and Mishra (1976) and Britton and Mehley (1955), respectively. Determination of reducing and total soluble sugars was performed by reference to glucose standards using the calorimetric copper method of Somogyi (1952) and for starch extraction, the method of McCready et al. (1950) was followed. Air monitoring of gaseous pollutants (SO2, NO2 and O3) was done with the help of gas samplers kept at 30 cm height from ground at each site by using wet chemical methods. SO2, NO2 and O3 were measured by methods of West and Gaeke (1956), Merryman et al. (1973) and Byers and Saltzman (1958), respectively. No continuous advanced gas analyzers were available and

A. Singh et al. / Environmental Pollution 134 (2005) 385–395

387

Fig. 1. Map of Allahabad city showing location of experimental sites.

gas samplers using wet chemical methods were the best possible devices with the available resources. Monitoring of pollutants was conducted for 6 h from 10 A.M. to 4 P.M. at weekly intervals at each site, this was the only option to ensure the safety of the samplers and because of frequent failure of electricity at various sites as samplers have a battery back up of only 6 h. Data were analyzed through three-way and two-way ANOVA using SPSS software (SPSS Inc., version 10.0) for assessing the significance of quantitative changes in different parameters due to ambient air pollution.

3. Results Results of air monitoring showed that RRP was the most polluted sites among all experimental sites, where

SO2, NO2 and O3 were recorded in the range of 38.2– 65.0, 30.8–42.5 and 17.0–30.8 mg mÿ3, respectively. Minimum concentrations of SO2, NO2 and O3 ranged between 10.6–18.3, 2.5–12.5 and 3.5–15.3 mg mÿ3, respectively at site Ar (Table 3). Since all the pollutants showed minimum concentrations at Ar, this site was treated as reference site for comparing the levels of changes in various parameters recorded at other sites with relatively elevated levels of pollutants. Total biomass of palak plants was reduced with increasing pollution load at all sampling intervals (Fig. 2) Significant negative correlations were found between total biomass and SO2 (r Z ÿ0.92, p ! 0.01), NO2 (r Z ÿ0.85, p ! 0.05) and O3 (r Z ÿ0.91, p ! 0.01) (Table 4). F2 treatment showed a positive response against air pollutants by increasing the total biomass. Maximum total biomass (4.7 g plantÿ1) was

388 Table 2 Brief description of experimental sites Site code Experimental Character of site site AAI Allahabad Agricultural Institute

A. Singh et al. / Environmental Pollution 134 (2005) 385–395
0.6

Total biomass (g plant-1)

F0
0.5 0.4 0.3 0.2 0.1 0.0

F1

F2

F3

20 DAS

Distance (km) and direction from city centre 3 km south

Total biomass (g plant-1)

Ar

Arail

Bh

Bahrana

Jh Mh

Jhunsi Mehdeori

CL

Civil lines

RRP

Rajrooppur

Near bank of river Yamuna, and national highway (NH-27); heavy traffic, frequent congestion, heavy vehicles, medium density population. Near bank of river Yamuna, open, small population City centre, near national highway (NH-2), heavy and light motor vehicles, frequent traffic jams, high density population. Near national highway (NH-2) periurban area Near bank of river Ganga, light vehicles, periurban area. Commercial area, near railway station, urban area Near national highway (NH-2), industries, railway track, heavy traffic, high density population, urban area

3 km south east

4.0 3.5 3.0 2.5 2.0 1.5 1.0

35 DAS

0 km

5 km east south 5.5 km north

Total biomass (g plant-1)

4.5 4.0 3.5 3.0 2.5 AAI CL Mh Jh Bh Ar

50 DAS

2.5 km north west 6.5 km west

RRP

Experimental sites

recorded at site Ar in F2 treatment, which was 3.9 g plantÿ1 at F0 treatment. Total biomass was least at site RRP (3.3 g plantÿ1) under F0 treatment and it increased to 4.3 g plantÿ1 due to F2 treatment (Fig. 2). Three-way ANOVA test showed that the variations in total biomass were significant ( p ! 0.001) due to plant age, site, nutrient treatment and their interactions (Table 5). Total chlorophyll and carotenoid contents in palak leaves were lower at sites experiencing higher pollution
Table 3 Levels of SO2, NO2 and O3 at different sites during experiment (mg mÿ3) Gaseous pollutants Sep., 2001 SO2 NO2 O3 Oct., 2001 SO2 NO2 O3 Nov., 2001 SO2 NO2 O3 Experimental sites CL 16.4 18.6 10.2 40.0 24.4 14.3 45.7 27.5 20.4 Ar 10.6 2.5 3.5 15.3 8.0 10.3 18.3 12.5 15.3 RRP 38.2 30.8 17.0 55.7 36.6 27.6 65.0 42.5 30.8 Mh 14.2 7.5 6.5 23.5 14.6 10.0 32.5 18.7 15.9 Jh 16.1 12.5 8.4 30.2 20.5 11.7 38.6 23.6 16.9 Bh 35.4 27.5 16.4 51.2 31.9 22.5 60.3 37.5 26.4 AAI 25.4 22.2 12.5 44.6 26.5 18.6 50.4 31.6 24.0

Fig. 2. Effect of air pollution on total biomass of palak plants grown at different experimental sites with varying fertility levels.

load (Fig. 3). Maximum total chlorophyll (0.95 mg gÿ1 dry leaf) and carotenoid contents (0.44 mg gÿ1 dry leaf) were observed at Ar (reference site) and minimum at RRP, most polluted site (total chlorophyll 0.65 mg gÿ1 dry leaf; carotenoid 0.3 mg gÿ1 dry leaf) 50 DAS under F2 treatment. Total chlorophyll and carotenoid contents showed significant negative correlations with SO2 (r Z ÿ1.0, p ! 0.001 and r Z ÿ0.99, p ! 0.001, respectively), NO2 (r Z ÿ0.96, p ! 0.001 and r Z ÿ0.98, p ! 0.001, respectively) and O3 (r Z ÿ0.98, p ! 0.001 and r Z ÿ0.99, p ! 0.001, respectively) (Table 4). ANOVA test showed that total chlorophyll and carotenoid contents varied significantly due to plant age, site, fertilizer treatment and their interactions except for plant age ! site ! fertilizer treatment interactions for carotenoid (Table 5). Protein and ascorbic acid contents showed significant negative correlations with individual air pollutants (Table 4). Protein content increased with the increase of plant age, while ascorbic acid decreased. Maximum protein and ascorbic acid contents were observed at 50 and 35 DAS, respectively under F2 treatment at Ar (Fig. 4). Variations in for protein and ascorbic acid

A. Singh et al. / Environmental Pollution 134 (2005) 385–395 Reducing sugars

389

Table 4 Correlation matrix of gaseous pollutants and total biomass and different physiological and biochemical characteristics of palak plants

contents were significant due to plant age, site, fertilizer treatments (Table 5). Catalase activity decreased with increasing levels of air pollutants, while peroxidase activity increased (Fig. 5). Catalase activity showed significant negative correlations with SO2 (r Z ÿ0.97, p ! 0.001), NO2 (r Z ÿ0.95, p ! 0.001) and O3 (r Z ÿ0.95, p ! 0.001), while peroxidase activity showed highly significant positive correlation with SO2 (r Z 0.99, p ! 0.001), NO2 (r Z 0.95, p ! 0.001) and O3 (r Z 0.98, p ! 0.001) (Table 4). Three-way ANOVA test showed significant variations in enzyme activities due to plant age, site and fertilizer treatment and their interactions except for plant age ! site, site ! treatment and plant age ! site ! treatment interactions for catalase activity (Table 5). Starch and reducing sugar contents also decreased with increasing levels of air pollutants while soluble sugars increased (Fig. 6). Correlation matrix showed a significant negative correlation between individual pollutants and starch and reducing sugars and positive correlation between soluble sugars and pollutants (Table 4). Starch, reducing sugars and soluble sugars varied significantly due to plant age, site, fertilizer treatment and their interactions except for plant age ! site, site ! treatment and plant age ! site ! treatment for starch and soluble sugars (Table 5).

Soluble sugars Peroxidase activity Catalase activity Ascorbic acid Starch

ÿ0.91** ÿ0.95*** ÿ0.95*** 0.85* 0.94** 0.94** 0.91** 1.00

ÿ0.97*** ÿ0.95*** ÿ0.95*** 0.81* 0.98*** 0.98*** 0.94** 0.88** 1.00

0.99*** 0.95*** 0.98*** ÿ0.93** ÿ0.99*** ÿ0.98*** ÿ0.96*** ÿ0.95*** ÿ0.94*** 1.00

ÿ0.96*** ÿ0.94** ÿ0.96*** 0.85** 0.98*** 0.96*** 0.97*** 0.97*** 0.94* ÿ0.98*** 1.00

0.95*** 0.82* 0.87** ÿ0.88** ÿ0.93** ÿ0.90** ÿ0.95*** ÿ0.80* ÿ0.90** 0.94** ÿ0.90** 1.00

ÿ0.99*** ÿ0.97*** ÿ0.99*** 0.90** 0.99*** 0.99*** 0.96*** 0.94* 0.98*** ÿ0.99*** 0.97*** ÿ0.91** 1.00

4. Discussion In many cities of developing countries, the levels of air pollutants often exceed toxic limits and adversely affect human health, vegetation and built cultural heritage. In urban areas of Allahabad city, high levels of automobile emissions have elevated the levels of pollutants to an extent that inhibited the plant growth and reduced the yield of palak grown in urban and periurban areas. Urban air quality of Varanasi, an adjoining city of Allahabad has also been shown to cause deleterious effects on woody transplants grown in urban areas (Pandey and Agrawal, 1994) and yield losses in crop plants grown in periurban areas (Agrawal et al., 2003). Air monitoring conducted in Varanasi has shown that SO2 concentration varied from 14 to 43 mg mÿ3, NO2 from 16 to 34 mg mÿ3 and O3 from 12 to 42 mg mÿ3 during the rainy season (July–October) in urban areas (Pandey et al., 1992). The levels of SO2 and NO2 observed in the present study are similar to that of Pandey et al. (1992), but O3 levels are low. From the meteorological data it is clear that rains were frequent in September and October, and hence all the pollutants including O3 showed lower values during these months. In November, however, O3 formation increased with a longer sunshine period. The permissible annual safe limits set by CPCB, India for 8 hourly SO2 and NO2

Carotenoid

Protein

ÿ0.99*** ÿ0.98*** ÿ0.99*** 0.89** 0.99*** 1.00

ÿ0.96*** ÿ0.89** ÿ0.92** 0.85* 0.98*** 0.94** 1.00

*p ! 0.05, **p ! 0.01, ***p ! 0.001 and NS Z not significant.

Total chlorophyll Total biomass

SO2 NO2 O3 Total biomass Total chlorophyll Carotenoid Protein Ascorbic acid Catalase activity Peroxidase activity Starch Soluble sugars Reducing sugars

ÿ0.92** ÿ0.85* ÿ0.91** 1.00

ÿ0.99*** ÿ0.96*** ÿ0.98*** 0.88** 1.00

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A. Singh et al. / Environmental Pollution 134 (2005) 385–395

Table 5 Variance ratio for total biomass, photosynthetic pigments, protein, ascorbic acid, enzyme activities and carbohydrate content of palak plants grown with different fertility levels at various experimental sites Parameter Total biomass Total chlorophyll Carotenoid Protein Ascorbic acid Catalase activity Peroxidase activity Starch Soluble sugars Reducing sugars Plant age (A) *** *** *** *** *** *** *** *** *** *** Site (B) *** *** *** *** *** *** *** *** *** *** Treatment (C) *** *** *** *** *** *** *** *** *** *** A!B *** *** *** NS NS NS * NS NS ** A!C *** *** *** *** *** *** *** *** *** *** B!C *** *** *** NS NS NS ** NS NS *** A!B!C *** *** NS NS NS NS ** NS NS ***

*p ! 0.05, **p ! 0.01, ***p ! 0.001 and NS Z not significant.

concentrations in urban areas are 60 mg mÿ3. SO2 concentration crossed this limit at two sites (RRP and Bh) during September. NO2 concentration remained always below the permissible limit. There is no
0.7 F0 0.6 0.5 0.4 F2 F1 F3 20 DAS

permissible safe limit set for O3 in India. Since the monitoring of pollutants was conducted on 6 hourly sample collections, the measurement of peak concentrations in between cannot be provided. In Varanasi, 2 h

F0 F2

F1 F3

20 DAS

0.4

0.3

0.2 0.3 0.1 0.2 0.1 0.8 0.7 0.6 0.5 0.2 0.4 0.3 0.2 1.0 50 DAS 50 DAS 0.1 0.0 0.5 0.4

Total chlorophyll (mg g-1 dry leaf)

35 DAS

35 DAS

0.3

0.0 0.5 0.4

0.8 0.3 0.6 0.2 0.4 0.1 0.0 AAI CL Mh Jh Bh Ar RRP AAI CL Mh Jh Bh Ar RRP

0.2

Experimental sites

Experimental sites

Fig. 3. Effect of air pollution on total chlorophyll and carotenoid contents in leaves of palak plants grown at different experimental sites with varying fertility levels.

Carotenoid (mg g-1 dry leaf)

A. Singh et al. / Environmental Pollution 134 (2005) 385–395
14 Protein (mg g-1 fresh leaf) 13 12 11 10 9 8 7 6 1.6 0.75 1.2 0.60 50 DAS 0.45 AAI CL Mh Jh Bh Ar RRP 35 DAS 50 DAS 10 0.90 12 11 15 Starch (mg g-1 dry leaf) 14 13 Protein (mg g-1 fresh leaf) F0 F2 F1 F3
400

391

F0
375

F1

F2

F3

350

325

Ascorbic acid (mg g-1 fresh leaf)

Ascorbic acid (mg g-1 fresh leaf)

300

Soluble sugars (mg g-1 dry leaf) Reducing sugars (mg g-1 dry leaf)

300 275 250 225 200 175

0.8

35 DAS AAI CL Mh Jh Bh Ar RRP

Experimental sites

Experimental sites

Fig. 4. Effect of air pollution on protein and ascorbic acid contents in leaves of palak plants with varying fertility levels at different experimental sites.

250

200

Catalase activity (mM H2O2 decomposed min-1 g-1 fresh leaf)

90 80 70 60 50 40 30 20

150

F0 F2

F1 F3

AAI

CL

Mh

Jh

Bh

Ar

RRP

Experimental sites

Fig. 6. Effect of air pollution on starch, soluble and reducing sugar contents in leaves of palak plant at 50 DAS sites with varying fertility levels at different experimental.

Peroxidase activity (mM purpurogallin formed min-1 g-1 fresh leaf)

5

4

3

2

1

0 AAI CL Mh Jh Bh Ar RRP

Experimental sites
Fig. 5. Effect of air pollution on catalase and peroxidase activity in leaves of palak plants at 50 DAS with varying fertility levels at different experimental sites.

peak concentrations of SO2 varied from 25 to 95 mg mÿ3, NO2 from 27 to 61 mg mÿ3 and O3 from 21 to 102 mg mÿ3 in periurban and urban areas (Pandey et al., 1992). Peak concentration affects vegetation more adversely than prolonged exposure to low concentrations (Lefohn and Jones, 1986). The lower O3 concentration recorded during the present study may also be ascribed to the presence of monitoring sites near roads, where O3 is quickly scavenged. The adverse effects of urban air pollutants are clearly evident on physiological and biochemical processes of palak plants during the present investigation. Photosynthetic pigments are fairly sensitive to air pollutants and their sensitivity may determine the responses of plants to pollutants. Significant negative correlations were obtained between air pollutants and total chlorophyll and carotenoid contents. Khan and Khan (1994) reported that combined treatments of O3 and SO2 at all concentrations had a significant negative effect on the leaf pigments. Exposure to SO2 causes more reduction in chlorophyll than carotenoids in wheat plants (Verma and Agrawal, 2001). El-Khatib (2003) also reported reductions in chlorophyll and carotenoid contents

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A. Singh et al. / Environmental Pollution 134 (2005) 385–395

caused by elevated levels of O3 in five common Egyptian plant species. Nutrient amendment in various combinations significantly lowered the magnitude of reduction in chlorophyll as compared to unamended plants (F0 treatment). Verma et al. (2000) also reported that nutrient amendment lowered the magnitude of reduction in chlorophyll content as compared to unamended ones, confirming the results of the present investigation. Nitrogen supply is reported to increase the leaf photosynthesis via the amount of N-containing components such as ribulose-1,5 bisphosphate carboxylase/ oxygenase activity (Sivasankar et al., 1993) and also by chlorophyll formation (Agrawal and Verma, 1997). However, N and P deficiency reduces the chlorophyll concentration (Rousseau and Reid, 1990). Air pollutants are known to induce the degradation of biologically important molecules such as proteins with the consequent release of malondialdehyde (Mudd, 1982). Protein content showed significant negative correlations with air pollutants and significant increase in protein content was observed due to application of mineral nutrients at all sites. The protein content depends upon N uptake and plants receiving a higher dose assimilate more N as compared to the unamended plants. Plants grown with higher N-supply invest a greater proportion of carbon in protein (Makino et al., 1984). Verma and Agrawal (1996) also noticed that N, P and K amendment in soybean plants significantly reduced the levels of decrease in protein content of SO2 exposed plants. Ascorbate is a ubiquitous soluble antioxidant in photosynthetic organisms and the most important reducing substrate for H2O2 detoxification. It has been suggested that pollutants produce oxyradicals in plants (Shimazaki et al., 1980; Sakaki et al., 1983). These radicals cause widespread damage to membranes and associated molecules including the chlorophyll pigments (Sakaki et al., 1983). Several authors have reported that ascorbic acid can serve as a free radical scavenger against O3 (van Hove et al., 2001; El-Khatib, 2003). The reduction in ascorbic acid concentration may be ascribed to its consumption during removal of cytotoxic free radicals generated in chain reactions after the penetration of oxidative pollutants into leaf tissues. Agrawal and Verma (1997) also observed higher ascorbic acid content in SO2 exposed plants amended with fertilizers. Peroxidase activity showed an increasing trend with increasing pollution levels at various sites, while catalase activity decreased. Singh (1998) also found that wheat plants exposed to O3 showed increase in peroxidase activity without any specific symptoms of O3 on foliage. Tingey et al. (1975) stated that stimulation of peroxidase activity in pollutant-exposed plants might be due to increased oxidative processes under pollutant stress. Present observations are in conformity with the earlier

findings of Ranieri et al. (1997), who showed that catalase activity decreased with increase in SO2 levels while peroxidase activity increased. Starch content showed significant negative correlation with individual pollutants. Rennenberg et al. (1996) suggested that O3 probably interacts with carbon allocation by inhibiting sucrose export. This causes an accumulation of starch in leaves, which results in reduction of photosynthesis and consequently reduces the level of starch in plants. Nutrient amendment in different combinations has significantly elevated the levels of starch in plants. Agrawal and Verma (1997) also reported reduction in foliar starch content of two cultivars of wheat treated with SO2 compared with untreated plants and also found significant increase after nutrients application. An increase in total soluble sugars was recorded with increasing pollution levels while reducing sugars reduced. The adverse effects of air pollutants are evident in the form of changes in pool volume of free carbohydrates in palak grown at different sites with various fertility levels. Katase et al. (1983) reported that SO2induced inhibition of photosynthesis in rice reduced the level of starch in the plants. An increase in concentration of sugar was associated with reduced starch content, suggesting increased hydrolysis of polysaccharides into monosaccharides due to gaseous pollution (Koziol and Jorden, 1978). Nutrient amendment has significantly reduced the level of decrease in starch and reducing sugar contents at various sites, which might be due to increased photosynthetic rate led by nutrient amendment. Meyer et al. (2000) reported that O3 caused inhibition of photosynthesis, and consequently decline in assimilate production. Agrawal et al. (2003) have also reported significant reductions in photosynthetic rate of a number plant species growing in a periurban area of Varanasi experiencing higher level of pollutants. Total biomass accumulation reduced in palak with increasing pollution load at various sites. This suggests that air pollutants directly interfere with various fundamental processes of plants, resulting in lower biomass accumulation. Ashmore et al. (1987) have also reported a decline in biomass accumulation in different plant parts along a gradient of air pollution around London. Agrawal et al. (2003) have also reported a negative correlation between ambient air pollutant levels and biomass accumulation in plants grown in the outskirts of Varanasi city experiencing similar climatic conditions. Verma et al. (2000) reported that 390 mg mÿ3 SO2 treatment for 4 h daily for 5 days weekÿ1 for 8 weeks resulted in a significant reduction in biomass accumulation and productivity in wheat plants were due to the integrated result of effects on a range of biochemical, physiological and metabolic activities in plants. The joint action of O3 and SO2 caused significant suppression in dry matter of tomato shoot and root at

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393

all concentrations (Khan and Khan, 1994). McKee et al. (1997) also reported that elevated O3 caused a 15% decline in total biomass accumulation in wheat plants. Fertilizer amendment has a significant effect on plant response to air pollutants. Agrawal and Verma (1997) reported that plant height and total biomass reduced significantly in SO2 treated plants, except those grown using recommended and twice recommended N, P and K applications. In the present investigation, F2 treatment showed the most positive impact on biomass accumulation and photosynthetic pigments by decreasing the negative impact of air pollutants. The percent reduction in total biomass at 50 DAS was 15.5 and 9.5%, respectively at RRP site in nutrient unamended and amended plants as compared to the same growing at Ar site experiencing lowest levels of air pollutants. The double recommended dose of nutrients showed maximum percent reduction in total biomass (29% at RRP site compared to Ar site) suggesting that this dose is supra optimal and caused negative influence on the plants. Application of nutrients higher than the demand has been shown to reduce the positive effects of fertilizers (Agrawal and Verma, 1997; Verma et al., 2000). Recently, Singh et al. (2003) also revealed that air pollutants suppressed the growth and yield of wheat plants grown at various urban and periurban sites of Allahabad but fertilizer amendment higher than the recommended dose resulted in a positive response by increasing the total biomass, weight of 1000 seeds and yield. The supply of macro nutrients N, P and K increased the total biomass in palak by increasing the levels of photosynthetic pigments, antioxidative property and metabolites in foliar tissue, which have further reduced the magnitude of reduction in biomass due to air pollutants compared to unfertilized plants. Coleman et al. (1989) have suggested that plants growing in nutrient poor conditions may be more sensitive to air pollution with respect to changes in carbon gain. N limitation has been shown to decrease chlorophyll and protein contents, RuBP carboxylase activity and increase the mesophyll resistance, which all limit CO2 fixation (Osman and Milthorpe, 1971). High P availability is found to increase the rate of photosynthesis (Rousseau and Reid, 1990). K fertilization is also beneficial due to its role in stomatal opening, photosynthesis, protein synthesis and osmotic and pH regulation (Wyn Jones and Pollard, 1983).

of palak plants grown at sites experiencing elevated pollutant concentrations. Increasing pollution load also deteriorated the nutritive quality of palak plants, as protein and carbohydrate contents were decreased. One and half times of recommended dose of NPK was most efficient in reducing the adverse effects of air pollutants on palak plants. The present investigation also suggests that urban air quality of Allahabad city is unfavourable for vegetable production in urban and periurban areas. Though the concentrations of individual pollutants were not very high except SO2, the levels of reductions were fairly significant. This clearly shows that pollutants in combination may have acted synergistically in causing greater adverse impact. Low concentrations of pollutants have been shown to increase the stomatal conductance thus facilitating the pollutant uptake and consequently greater negative response. Palak plants seem to be fairly sensitive to air pollutants under ambient conditions. More large-scale studies are, however, required to ascertain the potential for use of this plant as a biomonitor of air pollution.

Acknowledgements Authors wish to express sincere thanks to Prof. R.B. Lal, Vice Chancellor, Allahabad Agricultural Institute DU and Prof. P.W. Ramteke, Director (Research) for providing laboratory facilities and encouragements and to C.S.I.R. (New Delhi) for providing financial support. Authors are also grateful to Professor Madhoolika Agrawal (B.H.U.) and to anonymous reviewers for comments and fruitful suggestions.

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