MS Research Proposal

I. Title: Effect of sewage irrigation on growth and Bioavailability of

metals in spinach (Spinacia oleracea L.)

Submitted by

Nuaman Sharif

Reg. No.790-FBAS/MSES/F19

Supervised by
Dr. Muhammad Asad Ghufran
Associate Professor
2020

Department of Enviromental Science

FACULTY OF BASIC AND APPLIED SCIENCES
 INTERNATIONAL ISLAMIC UNIVERSITY, ISLAMABAD

II. Title: Effect of sewage irrigation on growth and

Bioavailability of metals in spinach (Spinacia
oleraceaL.)

Problem statement:
For all living things water is very important for their survival on the earth. From entire accessible
water, almost six percent used for household purposes, three percent for commercial &for
irrigation purposes 90% of water is used. There is scarcity of fresh water supply in Pakistan. This
scarcity is fulfilled by using underground water to raise domestic crops. Also by using sewage
water different vegetables can be grown in different areas(Lone, 1995).

Sewage water is increasing on routine basis by commercial and domestic sources due to their
increase usage (Qadir et al., 2008). Commercial, domestic and household activities generate
Sewage water which is a combination of heavy metals, pure water and chemicals comprising of
organic and inorganic constituents(Dixit et al., 2011).

Sewage water includes different heavy metals/oids like manganese (Mn), arsenic (As), lead (Pb),
boron (B), zinc (Zn), chromium (Cr), nickel (Ni) and copper (Cu) etc. most of citied heavy
metals are unessential which ultimately, affectliving organisms (Kanwar & Sandha, 2000).
Heavy metals accumulate in soil because of application of sewage water leading to penetration of
heavy metal into plant biomass which pose serious threats to human population. The plants that
are grown in sewage water have more concentration of heavy metals compare to those raised in
similar normal soil (Singh et al., 2010). The excess use of sewagewater for irrigation system has
enhanced contamination of As, Pb,Cd, and Ni causing long term health issues to consumers by
eatable part of vegetables (Sharma et al., 2006)

For irrigation, use of sewage water having organic and inorganic materials causes more plant
biomass giving more yield (Mitra and Gupta, 1999). This operation provides farmers nutrient
supplemented water which is an inexpensive and consistent disposal of waste water for society
(Ullah et al., 2011; Feigin et al., 1991). The first priority of the Pakistani farmers is to apply the
wastewater even if freshwater is accessible because by the usage of sewage water they earn
higher profits. Wastewater is more consistent source, in terms of availability and volume.
Additionally, the wastewater provides great resources to earn income and jobs for the farmers in
many ways. (Ensink et al., 2004).The use of fertilizers and increased application of sewage water
can have bad effects on the crops and accumulations of different metals in soil andvegetables
(Uwah et al. 2011)

Vegetables are regularly used for cooking purposes, Leafy vegetable like spinach is a good
source of high value nutrition and most important selling vegetable in different areas. (Sobukola
and Dairo, 2007). Spinach (leafy vegetable) is most important food that helps in prevention and
curing a lot of diseases (D’Mello, 2003). Vegetables like spinach and cabbage can grow easily
by waste water as compared to vegetables like radish (other than green) are very sensitive to
waste water (Bakhsh et al., 2005). By growing vegetables in the sewage water causes increased
accumulation of various metals having health effects and problems to living organisms (Qadir et
al., 1999).

Leafy green vegetables like spinach mostly used for cooking purposes. The quality of soup can
be enhanced by using green leafy vegetables like spinach and also for nutritional purposes
(Sobukola et al., 2007). Green vegetables are very significant safe food, good for the improving
health, and also for the treatment and prevention of many diseases (D’Mello, 2003). But these
can be dangerous for human beings and their health if increased accumulation of heavy metals
added by irrigation of sewage water. Vegetables like (Lettuce and radish) are more responsible
for the addition of heavy metals in the humans through eatable part than other green vegetables
(Intawongse and Dean 2006).
Some of the heavy metals are necessary for the growth of plants but some are harmful and are
unnecessary so if they accumulate in excess amount in the soil they transmitted to food chain and
cause serious effects to consumers (Malla et al., 2007; Ghafoor et al., 1995).Leafy green
vegetable such as Spinach response toward pollutants is very sensitive due to the accumulation
higher quantities of heavy metals, when these vegetable are taken up by the humans they cause
lethal health issues to the health of human beings and animals (Madhvi et al. 2014).

The amount of heavy metals that are directly absorbed by plantsbe determined by plantsspecies,
varieties, soil temperature and composition of soil.Vegetables that are eaten by human which
consist of large amount of heavy metals can cause toxicity in human and these metalspose many
health hazards to human (Adefemi et al., 2012; Duruibe et al. 2007)

Objective:

To minimize the effect of heavy metals on spinach by using dilution technique.

III. REVIEW OF LITERATURE:

Lone et al., (2003) conducted a study to evaluate the concentration of heavy metals in the leaves
of spinach and okra fruit by using sewage and tube well water. Tube well water and sewage
water are given to the fields which were chosen for this experiment and also mixture of
sewage/tube well water given to the fields, recommended dose of N100 kg ha-1 and P 50 kg ha-1
was given to both vegetables. Okra and spinach was planted in spring and winter season
respectively. Micronutrient and heavy metals were present in both vegetables significantly in
higher amount except copper and iron. In both crops Cu and Fe shows variations, amount of
sewage water is given more than tube well and mixture of sewage/tube well water. In this study
it was revealed that leaves of spinach showed higher concentration of heavy metals as compared
to okra.

Ahmad et al., (2006) investigated the effect of sewage water on the yield of spinach in a survey
based study in the district of R.Y. Khan. 70 spinach growers were interviewed and to know the
positive and negative effects on the yield of spinach, Cobb Douglas production function was
used. This survey shows the positive effects of sewage water on the yield but it reduces soil
productivity and fertility if used continuously. In orderto clean sewage water treatment plants are
requiredbecause of beneficial and economic aspects this is the only way to get benefit from this
type of water.

Perveen et al.,(2012) conducted a research in the city of Peshawar, Pakistan to analyze the effect
of sewage water on vegetable quality. Forty samples of vegetables leaves, soil and water from
forty different localities were selected for various physiochemical analyses. Water from Bara
River and Warsak Canal polluted with wastewater usedto irrigatethe crop. Samples of water were
slightly alkaline and the concentration of Pb (13%), Cd (98%)and Ni (18%) were more toxic as
compared to Cr which is in safe limit. Heavy metals in leaves were in the order of Ni,Cd,Cr,Pb
(90, 10, 78 and 15% respectively),and the plant Cd had a positive non-significant correlation
with soil Cd while other heavy metals in leaves had a significant positive correlation with the
heavy metals in soil. The vegetables tested were not safe for human use, particularly those
directly consumed by human beings.

Uwah et al., (2012) determined the concentrations of four heavy metals Zinc, Manganese, Iron
and Copper by using Atomic Absorption Spectrophotometer (AAS) techniques in the lettuce and
spinach’s leaves and rootswhile also in the soil where vegetables were grown at Potiskum, Yobe
State, Nigeria. It was concluded that heavy metal contamination was much more than the safe
limits of the vegetables. However in vegetables root area was contained more heavy metals along
with leaves of vegetables. So on Consumption these vegetables pose health hazard to living
being.

Khan et al., (2013) directed a survey to check the heavy metals into the eatable part of
vegetables, agricultural land by using sewage water. In the present study, thirty vegetable,
seventy six soil samples, forty leaf and twenty five sewage water samples in the year of 2009 and
2010 of different vegetables such as spinach, cauliflower, tomato and carrot were taken from
Lahore city areas.Samples were used to check for different heavy metal contents. The analysis
shows the presence of copper, Manganese, Nickel and Cadmium (100 %, 72%, 32% and 44%
respectively) but the concentration of iron, zincand lead werein the safe limits.

Madhvi et al., (2014)observed the effect of different types of water such as groundwater, treated
wastewater and untreated wastewater on the morphological characters of spinach in a field
experiment. This study shows that by using wastewater many of parameters like dry weight and
fresh weight, root, leaf and stem length were increased but vigour index, germination %
decreased and it also effects shape,color, and size of leaf. It concluded that by the irrigation of
sewage water many negative change occur in the important parameters in this experiment.

Saini et al., (2014) carried a research in Bhiwadi industrial area where they used to regulate the
applications of different metals in the Spinach which irrigated by wastewater. AAS is used to
check the amount of metals in the spinach. The heavy metals that were investigated in this
experiment were above the safe limits that were recommended. There is a lot of health problems
to the consumers of these vegetables and steps must be taken by the organization to minimize
this issue. The study shows that sewage water have many heavy metals and the spinach which
were grown by using wastewater as a bio indicator of pollution of many heavy metals and may
be helpfulfor phytoremediation purposes.

MATERIALS AND METHODS:

Collection of water and soil sample:

Waste water samples will be taken from the ponds of Chokera waste water treatment plant,
Faisalabad. Sewage water with distill water will be used to prepare different dilutions i.e. 80, 60,
40, and 20 times. Raw sewage water will be applied and a control in which distill water alone
will be used. Soil samples will be collected from the agriculture land of (UAF) University of
Agriculture Faisalabad, Institute of (ISES) Soil & Environmental Sciences. Then the collected
soil material will be sieved using 2 mm sieve and mixed thoroughly for further analysis and to
fill the pots.

Selection of plant cultivar for experiment:

Spinach will be chosen for the experiment and seeds will be purchased from Ayub Agriculture
Research Center (AARI), Faisalabad.

Experimental setup:

This experiment will be directed at wire house of (SARC) ISES, UAF. Before Applications
Sewage water will be analyzed for physical ,chemical and biological water characteristics like
color , turbidity , pH , EC , TSS , TDS , Ca+Mg , Na , SAR, RSC , CO 3, HCO3, Cr, Cd, Fe, Pb ,
Mn , Cu, BOD and COD. This experiment will be conduct in pots. Each pot will be filled with
12kg soil and five spinach seeds will be sown in each pot. N, P and K will be added according to
the suggested dose, at sowing time of spinach. Six treatments each with three replications, will
be applied and there will be 18 pots in total.

TREATMENTS DETAILS:
Treatment

T0= Control (Distilled water)

T1=80 Time Dilution (1:80)

T2=60 Time Dilution (1:60)

T3=40 Time Dilution (1:40)

T4=20 Time Dilution (1:20)

T5= Sewage water

After 30 days of sowing chlorophyll contents will be measured by using chlorophyll meter.
While osmotic potential will be measured by using Osmo-meter. There will be two harvest of
this experiment , 1st will be taken after 6 weeks of germination and 2 nd will be taken after 12
weeks of germination. After harvesting the growth and yield parameters will be recorded like:

Parameters to be studied:

No of plants
Shoot length
Fresh weight
Dry weight
 Root length
Root weight
Plant analysis (Cr, Cd, Fe, Pb, Mn and Cu)
Post harvesting soil analysis

Statistical analysis:
The data will be analyzed statistically using software 8.1.LSD test (P≤ 0.05) will be applied to
find out the significance among different treatments. (Steel et al., 1997).