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© 2021 by the authors; licensee Asian Online Journal Publishing Group 
 

Agriculture and Food Sciences Research 
Vol. 8, No. 2, 36-43, 2021 

ISSN(E) 2411-6653/ ISSN(P) 2518-0193 
DOI: 10.20448/journal.512.2021.82.36.43 

© 2021 by the authors; licensee Asian Online Journal Publishing Group 

  

 
 
 
Toxicological Indices of Wistar Rats Fed Formulated Chaw of Telfairia occidentalis 
Planted on Crude Oil Contaminated and Remediated Soil 

 
Reginald C. Ohiri1   
Eugene N. Onyeike2 
Augustine A. Uwakwe3 

  
( Corresponding Author) 

 
1,2,3Department of Biochemistry, Faculty of Science, University of Port Harcourt, Nigeria. 
1Email: raycohiri@yahoo.com  
2Email: eugene.onyeike@uniport.edu.ng  
3Email: augustine.uwakwe@uniport.edu.ng  

 
Abstract 

Toxicological indices of wistar rats fed formulated chaw of Telfairia occidentalis planted on crude 
oil contaminated and bioremediated soil was studied. Farmland, measuring 18 m2 was divided into 
three lots of 4 m2 and 2 m space was allowed between each lot (A, B and C). Two lots were 
polluted, subjected to 16 weeks of bioremediation and viable seeds of T. occidentalis were planted 
and grown for 28 days. The harvested vegetable leaves were analyzed. Leaves from natural 
attenuated soil showed high concentration of both aliphatic and polycyclic aromatic hydrocarbons 
with values of 9.34+0.03 mg/kg and 9.18+0.05 mg/kg for C36 and Fluoranthene respectively, 
while the bioaugmented soil had Lead as the highest heavy metal with a value of 0.10+0.02 
g/100g. Four groups, of 9 rats each (totalling 36 rats) of adult males and females, wealing males 
and females were subsequently sub-grouped into 3 and allowed to acclimatize for a period of 1 
week. These animals were fed formulated rat chaw of 50% dry weight of T. occidentalis, from the 
three lots (control, bioaugmented and natural attenuated soil) for a period of 28 days. Assay of 
animals’ toxicological index showed elevated concentrations of alkaline phosphatase, alanine and 
aspartate aminotransferases, urea and creatinine in their sera. A slight increase in serum amylase 
activity was recorded, while concentrations of their haemoglobin and Packed Cell Volume (PCV) 
reduced. The obtained results from these biomarkers were confirmed by the histopathological 
sections of the animal organs. 
 

Keywords: Accumulation, Biotransformation, Crude oil, Excretion, Ingestion, Tocixity. 

 
Citation | Reginald C. Ohiri; Eugene N. Onyeike; Augustine A. 
Uwakwe (2021). Toxicological Indices of Wistar Rats Fed 
Formulated Chaw of Telfairia occidentalis Planted on Crude Oil 
Contaminated and Remediated Soil. Agriculture and Food Sciences 
Research, 8(2): 36-43. 
History:  
Received: 10 September 2021 
Revised: 2 December 2021 
Accepted: 20 December 2021 
Published: 14 January 2022 
Licensed: This work is licensed under a Creative Commons 

Attribution 3.0 License  
Publisher:  Asian Online Journal Publishing Group 
 

Acknowledgement: All authors contributed to the conception and design of 
the study. 
Funding: This study received no specific financial support. 
Competing Interests: The authors declare that they have no conflict of 
interests. 
Transparency: The authors confirm that the manuscript is an honest, 
accurate, and transparent account of the study was reported; that no vital 
features of the study have been omitted; and that any discrepancies from the 
study as planned have been explained. 
Ethical: This study follows all ethical practices during writing.   

 

 

Contents 
1. Introduction ...................................................................................................................................................................................... 37 
2. Experimental Materials and Methodology ................................................................................................................................. 37 
3. Results ................................................................................................................................................................................................ 39 
4. Discussion .......................................................................................................................................................................................... 42 
References .............................................................................................................................................................................................. 43 
 

 
 

 

 

mailto:raycohiri@yahoo.com
mailto:eugene.onyeike@uniport.edu.ng
mailto:augustine.uwakwe@uniport.edu.ng
http://creativecommons.org/licenses/by/3.0/
http://creativecommons.org/licenses/by/3.0/
https://www.doi.org/10.20448/journal.512.2021.82.36.43
https://orcid.org/orcid-search/quick-search?searchQuery=AJAYI%20Boboye%20L.


Agriculture and Food Sciences Research, 2021, 8(2): 36-43 

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Contribution of this paper to the literature 
This study contributes to existing literature by confirming that accumulated hydrocarbons and heavy 
metals in plants (vegetables) will cause destruction of cellular architecture in animals upon ingestion, 
thereby causing impairments in cellular activities. 

 
1. Introduction 

One established mechanism by which plants decontaminate oil contaminated soil is by direct absorption of the 
contaminant hydrocarbons and subsequent accumulation of same in specific plant tissues and organelles. The 
Casparian strip has remain a structural foundation for uptake of substances through the plant roots. This system 
blocks the possible absorption of soil solution via the plant root to the xylem [1]. Substances inside root cells are 
moved to the aerial parts of the plant. Foliar uptake has been the most important pathway for petroleum 
hydrocarbon uptake [2]. Other factors that influence hydrocarbon uptake aside from foliar uptake include aqueous 
concentration of petroleum hydrocarbon, uptake/exposure period, plants bio-physical and chemical compositions. 
Kipopoulou, et al. [3], reported solubility, octanol-water and octanol-air partition coefficients and vapour pressure 
as good predictors of petroleum hydrocarbon accumulation in leaf tissues of vegetables. T. occidentalis is a 
commonly produced crop in the Niger delta area of Nigeria. This region has recorded exorbitant crude oil pollution 
since the advent of petroleum exploration in the area and there is a continuous annual elevation of its contaminant 
load in concord with industrial surge and energy uptake thus necessitating the need for environmental remediation. 
After remediation of petroleum hydrocarbon pollution within and agricultural area, cultivation and growing of 
plants are carried out without consideration to hydrocarbon recalcitrance in the post remediated soil. These post 
remediated hydrocarbons may be absorbed, accumulated and a possible high or low potent toxicant can be 
generated as an intermediary metabolite and stored in specific plant cells. The photosynthetic activities in leafy 
vegetable (which T. occidentalis is amongst them) enriches them with protein, minerals and carbohydrates [4]. Also 
referred as fluted pumpkin, T. occidentalis Hoof (Family: Cucurbitaceae) has been a vital commercial crop of 
cultivated within the humid lowland tropical region situated in the Western part of Africa [5]. Because of its 
nutritional value, T. occidentalis is prominent in the diet of many animals, including humans [4]. This research was 
geared towards the evaluation of the toxicological indices of wistar rats fed formulated chaw from T. occidentalis 
planted on crude oil contaminated and remediated soil.  
  

2. Experimental Materials and Methodology 
2.1. Source of Materials 

A volume of 30 dm3 of bonny light crude oil was gotten from flow station of Shell Petroleum Development 
Company located at Egbema Local Government area of Imo State, Nigeria. Forty kilograms (40 kg) of poultry 
manure was obtained from Godvine Poultry Farm located at Obio Akpor Local Government area of Rivers State, 
Nigeria. Viable T. occidentalis seeds were obtained from Rumuokoro market, also located in Obio Akpor Local 
Goverment Rivers State, Nigeria. While a total of 36 healthy wistar rats of 9 rats each of adult males and females, 
wealing males and females were selected from the research animal house of Biochemistry Department, University 
of Port Harcourt, River State, Nigeria. 
 

2.2. Source of Reagent Kits  
Alanine and aspartate aminotransferase, alkaline phosphatase, creatinine and urea kits were sourced from 

Randox laboratories Ltd, Antrim, United Kingdom, while the kit for alpha amylase was from Giesse Diagnostics, 
Rome, Italy.  
 

2.3. Description of Research Site. 
The research site was geographically located on longitude 70 10″ E and latitude 40 40″ N on the Eneka axis of 

Eneka-Oyigbo inter-city road in Obio Akpor, Rivers State, Nigeria. Soils of this research site has an approximate 
pH of 4.86 + 0.12, it belongs to the ultisols and are made up of humus topsoil with a deep mixture of sandy and clay 
soil. Pipeline vandalization and oil spillage has not been recorded on the research site and its environs.   
 

2.4. Pollution and Remediation of Research Site 
A farmland, measuring 18 m2 was divided into three lots (A, B and C) of 4 m2 and 2 m land space was allowed 

between each lot.  Two lots were polluted and remediated as follows: Lot B = 4 m2 farmland polluted with 40 dm3 
of bonny light crude oil and remediated with 40kg of poultry manure for 16 weeks (bioaugmented soil). Lot C = 4 
m2 un-remediated farmland polluted with 40 dm3 of bonny light crude oil and allowed for 16 weeks (natural 
attenuated soil), while Lot A = Unpolluted 4 m2 farmland (control soil). 
  

2.5. Planting and Growing and Collection of Vegetable Samples 
After 16 weeks of remediation, viable T. occidentalis seeds were planted on the 3 lots and 4 weeks of growth 

were allowed from their germination date. Vegetable leaves were harvested at the fourth week with a sterilized 
razor blade, placed into sterilized cellophane bags and closed with plastic bands. Each cellophane bag of harvested 
samples was marked using a water-resistant marker and subsequently taken to the laboratory within one hour of 
sample collection for hydrocarbon and heavy metal analysis. 
 

2.6. Aliphatic and Polycyclic Aromatic Hydrocarbons (PAH) and Heavy Metals 
Concentration of both aliphatic and polycyclic aromatic hydrocarbons were extracted and determined in T. 

occidentalis by the method described by the AOAC [6], while the method described by Jones Jr [7], was used to 
determine heavy metal concentration in the vegetables.  
 
 



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2.7. Formulation of Rat Chaw 
Harvested leaves of T. occidentalis were dried and weighed using model AB204 Mettler Toledo weighing 

balance and a Model 4 Thomas Scientific Wiley’s mill was use to grind the vegetable. The ground sample was 
formed into rat chaw and combined with normal rat feed.  
 

2.8. Acclimatization, Treatment and Sacrificing of Research Animals 
Four groups of wistar rats made up of 9 rats each (totalling 36 rats) of adult males and females, wealing males 

and females were subsequently sub-grouped into 3. Both the control and test animals were placed at normal room 
temperature in steel cages and allowed to acclimatize for a period of 1 week while having free access to normal rat 
feed and portable drinking water. The sub-groups were placed on rat chaw comprising of equal ratio of normal rat 
feed and dried T. occidentalis from the control, bioaugmented and natural attenuated lots. Using chloroform 
anaesthesia, the animals were sacrificed at the 28th day and blood sample was obtained via cardiac puncture into 
ethylene diamine tetracetic acid (EDTA) bottles and taken to the laboratory within one hour for assay of the 
following toxicological indices: alkaline phosphatase, alanine and aspartate aminotransferases, urea, creatinine, 
hemoglobin concentration and packed cell volume (PCV). The internal organs (liver, kidney and pancreas) were 
immediately harvested into a preservative container of 10% physiological saline and subsequently subjected to 
histological analyses. 
 

Table-1. Concentration of aliphatic hydrocarbon (mgkg-1) in T. occidentalis  from crude oil remediated soil. 

No. of Carbon atoms Control soil Bioaugmented soil Natural   attenuated soil 

C8 2.29 + 0.05a 1.30 + 0.03b 5.37 + 0.20c 
C9 1.60 + 0.03ab 1.50 + 0.03ab 2.78 + 0.04c 
C10 1.04 + 0.02ac 4.43 + 0.05b 3.02 + 0.03ac 
C11 1.07 + 0.07a 7.56 + 0.10b 2.11 + 0.04c 
C12 2.70 + 0.02a 1.23 + 0.01b 1.56 + 0.08c 
C13 2.05 + 0.02a 2.31 + 0.06b 1.02 + 0.01c 
C14 1.47 + 0.09ac 8.17 + 0.02b 1.43 + 0.01ac 
C15 1.66 + 0.09a 1.19 + 0.02b 1.02 + 0.02c 
C16 1.03 + 0.01a 3.12 + 0.01b 7.90 + 0.18c 
C17 1.02 + 0.05a 3.02 + 0.02b 5.83 + 0.24c 

Pristane 1.17 + 0.06a 6.65 + 0.21b 5.08 + 0.05c 
C18 1.32 + 0.03a 2.08 + 0.14b 6.95 + 0.28c 

Phytane 1.61 + 0.01a 1.11 + 0.02b 4.14 + 0.02c 
C19 1.90 + 0.05a 4.18 + 0.02b 6.83 + 0.06c 
C20 8.56 + 0.06a 1.65 + 0.02b 6.17 + 0.06c 
C21 1.53 + 0.08a 9.03 + 0.09b 1.86 + 0.09c 
C22 2.76 + 0.07a 2.46 + 0.10b 1.19 + 0.04c 
C23 5.49 + 0.04a 3.07 + 0.14b 1.80 + 0.02c 
C24 6.83 + 0.08a 2.30 + 0.03b 1.61 + 0.02c 
C25 3.14 + 0.06a 6.36 + 0.07b 1.04 + 0.00c 
C26 6.49 + 0.03a 2.02 + 0.02b 7.46 + 0.09c 
C27 7.80 + 0.09a 9.58 + 0.09b 9.10 + 0.20c 
C28 1.22 + 0.01a 1.66 + 0.07bc 1.83 + 0.11bc 
C29 1.59 + 0.05a 6.17 + 0.06b 7.35 + 0.15c 
C30 1.17 + 0.03a 3.89 + 0.03b 5.11 + 0.01c 
C31 1.54 + 0.10a 3.87 + 0.05b 5.46 + 0.08c 
C32 6.09 + 0.03a 9.03 + 0.05b 7.03 + 0.01c 
C33 5.23 + 0.07a 1.47 + 0.05b 3.11 + 0.01c 
C34 2.51 + 0.02a 5.08 + 0.03b 6.44 + 0.07c 
C35 4.18 + 0.10a 3.55 + 0.08b 7.60 + 0.02c 
C36 2.82 + 0.44a 4.03 + 0.00b 9.34 + 0.03c 
C37 9.08 + 0.04a 5.97 + 0.21b 2.09 + 0.01c 
C38 2.26 + 0.08ab 2.19 + 0.04ab 7.18 + 0.02c 
C39 6.37 + 0.14a 8.79 + 0.04b 5.84 + 0.09c 
C40 2.55 + 0.08a 6.43 + 0.05b 1.58 + 0.07c 

Note: 
Data presented are mean values + standard deviations of three observations.  
Superscript “a” indicates statistical significance at P < 0.05 when control lot is compared to the other lots.  
Superscript “b” indicates statistical significance at P < 0.05 when bioaugmented lot is compared to the other lots. 
Superscript “c” indicates statistical significance at P < 0.05 when natural attenuated lot is compared to the other lots. 
Superscript “ab” indicates statistical insignificance at P < 0.05 when control and bioaugmented lots are compared to each other,  
But statistically significant when both are compared to the natural attenuated lot. 
Superscript “ac” indicates statistical insignificance at P < 0.05 when control and natural attenuated lots are compared to each 
other,  
But statistically significant when both are compared to the bioaugmented lot.Superscript “bc” indicates statistical insignificance 
at P < 0.05 when bioaugmented and natural attenuated lots are compared to each other, but statistically significant when both 
are compared to the control lot. 

 

2.9. Assay of Some Toxicological Parameters of Wistar Rats Fed with Vegetables from Bioremediated Soil. 
The diacetyl monoxine method (DAM), described by Marsh, et al. [8], was used to assay for serum Urea 

concentration while the Alkaline picric acid procedure described by Slot [9], was used to determine the 

concentration of Creatinine. The 2-chloro-p-nitrophenyl-α-D-maltotrioside (CNPG3) kinetic procedure described 
by Hohenwallner, et al. [10] and the assay procedure of Kind and King [11], were used to determine Amylase and 
Alkaline phosphatase activities respectively, while the colorimetric end-point procedure of described by Reitman 
and Frankel [12] was used to determine both Aspartate and Alanine aminotransferases. Corash [13], procedure 
was used the haematocrit (PCV) volume and haemoglobin concentration, while histopathological assay of animal 



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tissues was done using haematoxylin and eosin procedure described by Conn [14]  and modified by Kumar and 
Gill [15].    
 

2.10. Statistical Analysis/Test of Significance   
The mean + standard deviations and One Way Analysis of Variance were calculated electronically for each 

group of observations using International Business Machine (IBM) Statistical Program for Social Sciences (SPSS) 
19 software at 95% confidence level, while Duncan [16], post hoc multiple range test was used to test for 
significance. 
 

3. Results 
Hydrocarbon and heavy metal content of vegetable samples. The mean values of both aliphatic hydrocarbons, 

polycyclic aromatic hydrocarbons and heavy metals obtained in T. occidentalis grown on the 3 lots (control, 
bioaugmented and natural attenuated) are presented in Tables 1-3. A significant increase at P<0.05 in aliphatic 
hydrocarbon content was observed in T. occidentalis harvested from natural attenuated and bioaugmented soils 
aside C20, C24 and C37 where higher concentrations were recorded in the control vegetables than in the remediated 
soils   Polycyclic aromatic hydrocarbon content of T. occidentalis increased in vegetables harvested from both 
natural attenuated and bioaugmented aside acenaphthene, phenanthrene and indeno(1,2,3-cd)pyrene. Higher 
concentration of acenaphthene was recorded in T. occidentalis from the control lot than in those from both the 
natural attenuated and bioaugmented soil. Phenanthrene concentrations for those grown on the control and natural 
attenuated soil were equal with a value of 2.67mgkg-1, while Phenanthrene concentration in vegetable samples from 
the bioaugmented soil increased significantly. A significantly lower concentration of Indeno(1,2,3-cd)pyrene was 
observed at P<0.05 in vegetables harvested from the natural attenuated soil than in those from the control soil. A 
significantly high concentration at P<0.05 of heavy metals were observed in vegetables from the bioaugmented soil 
as compared to those from both the control and the natural attenuated lots. 
 

Table-2. Polycyclic aromatic hydrocarbon (PAH) content (mgkg-1) of T. occidentalis from crude oil bioremediated soil. 

Polycyclic aromatic hydrocarbon Control soil Bioaugmented soil Natural attenuated soil 

Naphthalene 0.55 + 0.07a 3.98 + 0.05b 2.95 + 0.04c 
Acenaphthylene 1.58 + 0.03a 2.87 + 0.01b 3.61 + 0.01c 
Acenaphthene 1.87 + 0.05ab 1.73 + 0.03ab 1.57 + 0.08c 
Fluorene 2.60 + 0.05a 4.44 + 0.04b 5.34 + 0.01c 

Phenanthrene 2.67 + 0.01ac 3.02 + 0.03b 2.67 +0.04ac 
Anthracene 2.33 + 0.01a 3.56 + 0.04b 3.80 + 0.01c 
Fluoranthene 1.57 + 0.01a 7.03 + 0.02b 9.18 + 0.05c 
Pyrene NP NP NP 

Benzo(α)anthracene 1.77 + 0.08a 7.24 + 0.07b 6.09 + 0.04c 

Chrysene 0.07 + 0.01a 2.52 + 0.11b 2.73 + 0.02c 

Benzo(β)fluoranthene 1.69 + 0.06a 1.85 + 0.08b 2.68 + 0.04c 

Benzo(k)fluoranthene NP NP NP 

Benzo(α)pyrene 0.53 + 0.02a 2.98 + 0.10bc 3.02 +0.00bc 

Indeno (1,2,3)pyrene 2.27 + 0.01a 3.28 + 0.03b 4.15 + 0.03c 

Indeno(1,2,3-cd)pyrene 3.47 + 0.07a 6.75 + 0.06b 0.43 + 0.03c 
Dibenz(a,h)anthracene 1.66 + 0.00a 2.58 + 0.04b 1.89 + 0.05c 
Benzo(g,h,i)perylene NP NP NP 

                  Note: 
Presented are mean values + standard deviations of three observations. NP= Not Present.  
Superscript “a” indicates statistical significance at P < 0.05 when control lot is compared to the other lots.  
Superscript “b” indicates statistical significance at P < 0.05 when bioaugmented lot is compared to the other lots. 
Superscript “c” indicates statistical significance at P < 0.05 when natural attenuated lot is compared to the other lots. 
Superscript “ab” indicates statistical insignificance at P < 0.05 when control and bioaugmented lots are compared to each other,  
But statistically significant when both are compared to the natural attenuated lot. 
Superscript “ac” indicates statistical insignificance at P < 0.05 when control and natural attenuated lots are compared to each other,  
But statistically significant when both are compared to the bioaugmented lot. 
Superscript “bc” indicates statistical insignificance at P < 0.05 when bioaugmented and natural attenuated lots are compared to each 
other,  
But statistically significant when both are compared to the control lot. 

 
Table-3. Heavy metal concentrations (g/100g) of T. occidentalis from crude oil bioremediated soil. 

Heavy  metal Control soil Bioaugmented soil Natural attenuated soil 

Copper 0.01 + 0.0ac 0.03 + 0.00b 0.01 + 0.00ac 

Lead NP 0.10 + 0.02b 0.08 + 0.00c 

Chromium NP 0.06 + 0.00b NP 

Cadmium NP 0.03 + 0.00b 0.01 + 0.00c 

Arsenic NP NP NP 

              Note: 
Data presented are mean values + standard deviations of three observations. NP= Not Present.  
Superscript “a” indicates statistical significance at P < 0.05 when control lot is compared to the other lots.  
Superscript “b” indicates statistical significance at P < 0.05 when bioaugmented lot is compared to the other 
lots. 
Superscript “c” indicates statistical significance at P < 0.05 when natural attenuated lot is compared to the 
other lots. 
Superscript “ac” indicates statistical insignificance at P < 0.05 when control and natural attenuated lots are 
compared to each other,  
But statistically significant when both are compared to the bioaugmented lot. 

 

 
 
 



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3.1. Indicators of Tissue Damage and Haematological Parameters 
 

Table-4. Serum urea and creatinine concentration (mgdl-1) of rats fed T. occidentalis from crude oil bioremediated soil. 

 Control soil Bioaugmented soil Natural attenuated soil 

Urea 

Weanling male               30.19+0.13a 54.35+0.51bc 54.32+0.11bc 
Weanling female            31.40+1.07a 56.80+0.20bc 55.50+0.36bc 
Adult male                     33.62+0.18a 52.00+0.63b 58.92+0.47c 
Adult female                  32.93+0.19a 58.61+0.30bc 58.66+0.43bc 

Creatinine 
Weanling male              1.19+0.02a 1.73+0.08bc 1.82+0.09bc 
Weanling female           1.11+0.02a 1.79+0.07bc 1.72+0.02bc 
Adult  male                    0.94+0.03a 1.66+0.03bc 1.71+0.05bc 
Adult  female                 0.80+0.01a 1.90+0.03bc 1.77+0.08bc 

                  Note: 
Data presented are mean values + standard deviations of three observations.  
Superscript “a” indicates statistical significance at P < 0.05 when control lot is compared to the other lots.  
Superscript “b” indicates statistical significance at P < 0.05 when bioaugmented lot is compared to the other lots. 
Superscript “c” indicates statistical significance at P < 0.05 when natural attenuated lot is compared to the other lots. 
Superscript “bc” indicates statistical insignificance at P < 0.05 when bioaugmented and natural attenuated lots are compared to each 
other,  
But statistically significant when both are compared to the control lot. 

 
Table-5. Serum enzymes concentration (I.Ul-1) of rats fed T. occidentalis  grown on crude oil bioremediated soil. 

 Control soil Bioaugmented soil Natural attenuated soil 

Alkaline phosphatase 
Weanling male               284.00+5.10a 486.90+10.40b 357.00+3.20c 
Weanling female            268.22+1.49a 479.81+ 0.40b 344.02+5.63c 
Adult male                     243.71+1.91a 346.20+ 1.50b 318.07+1.20c 

Adult female                  226.53+1.72a 359.70+ 0.84b 304.39+0.75c 
Alanine aminotransferase 

Weanling male              18.00+0.68a 61.28+0.24b 67.42+2.02c 
Weanling female           20.60+0.41a 57.63+0.53b 64.19+1.15c 
Adult  male                    25.20+0.84a 48.45+1.31b 61.59+0.75c 

Adult  female                 24.00+0.63a 41.05+0.43b 60.42+0.40c 
Aspartate aminotransferase 

Weanling male              19.00+0.28a 61.00+1.61bc 63.05+0.76bc 
Weanling female           19.22+0.80a 57.37+033bc 58.03+0.07bc 
Adult  male                    32.81+1.14a 57.60+0.02b 62.40+0.60c 
Adult  female                 29.70+0.67a 55.00+1.02b 63.81+0.91c 

Amylase 
Weanling male              27.66+0.46a 29.49+0.54bc 29.07+0.23bc 
Weanling female           26.85+0.34a 31.09+0.61bc 31.47+1.03bc 
Adult  male                    32.83+0.36a 38.18+0.86b 35.17+0.82c 
Adult  female                 33.51+0.47a 36.89+0.50bc 36.73+0.37bc 

                    Note: 
Data presented are mean values + standard deviations of three observations.  
Superscript “a” indicates statistical significance at P < 0.05 when control lot is compared to the other lots. 
Superscript “b” indicates statistical significance at P < 0.05 when bioaugmented lot is compared to the other lots. 
Superscript “c” indicates statistical significance at P < 0.05 when natural attenuated lot is compared to the other lots. 
Superscript “bc” indicates statistical insignificance at P < 0.05 when bioaugmented and natural attenuated lots are compared to each 
other,  
But statistically significant when both are compared to the control lot. 

 
Table-6. Packed Cell Volume (PCV) (%) and Haemoglobin concentration of rats fed T.  occidentalis from crude oil bioremediated soil. 

 Control soil Bioaugmented soil Natural attenuated soil 

Packed Cell Volume (%) 
Weanling male               31.35+l.33a 24.35+1.53bc 24.00+1.01bc 

Weanling female            31.67+0.58a 25.00+2.04bc 21.68+0.59bc 
Adult male                     34.34+2.52a 25.00±3.61bc 25.33+2.52bc 
Adult female                  36.33+0.57a 26.36+1.16b 23.67+0.56c 

Haemoglobin (gdl-1) 
Weanling male              10.44 + 0.51a 7.97 + 0.62bc 8.20 + 0.48bc 

Weanling female           10.56 + 0.20a 8.33 + 0.67bc 7.22 + 0.19bc 
Adult  male                    11.44 + 0.84a 8.34 + 1.21bc 8.44 + 0.84bc 
Adult  female                 12.11 + 0.19a 8.78 + 0.39b 7.89 + 0.20c 

       Note: 
Data presented are mean values + standard deviations of three observations.  
Superscript “a” indicates statistical significance at P < 0.05 when control lot is compared to the other lots.  
Superscript “b” indicates statistical significance at P < 0.05 when bioaugmented lot is compared to the other lots. 
Superscript “c” indicates statistical significance at P < 0.05 when natural attenuated lot is compared to the other lots. 
Superscript “bc” indicates statistical insignificance at P < 0.05 when bioaugmented and natural attenuated lots are compared to each other,  
But statistically significant when both are compared to the control lot. 

 
Tables 4 – 6 shows mean concentrations of Urea, Creatinine, Amylase, Alkaline phosphatase (ALP), Aspartate 

and Alanine aminotransferases, Packed cell volume and Hemoglobin concentrations of rats fed with 50% 
formulated chaw from the research lots. A significant increase at P<0.05 were observed in all the maker 
compounds (urea, creatinine, amylase, alkaline phosphatase, alanine and aspartate aminotransferases) of rats fed 
50% chaw from the bioaugmented and natural attenuated lots, while a more pronounced decrease in Packed cell 
volume and Hemoglobin were recorded in rats fed 50% chaw formulated from vegetables gotten from the 



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bioaugmented lot than in those fed T. occidentalis chaw  formulated from vegetables gotten from the natural 
attenuated lot. 

 

3.2. Sections of Rat Organs Fed with T. Occidentalis from Crude Oil Bioremediated Lots 
Figure 1 shows histological scores of sections of rat organs fed with T. occidentalis formulated chaw from the 

crude oil bioremediated lots. The observed lesions in the organ sections were highest in the kidney, liver and 
pancreas of animals fed T. occidentalis from natural attenuated and bioaugmented lots, while organ sections of those 
fed T. occidentalis from the control lot had healthier sections with non-significant degrees of lesions. 
 

 
Values are means + standard deviations of three determination. 

Figure-1. Histological scores of sections of rat organs fed T. occidentalis from crude oil bioremediated soil. 
 

Sections of rat organs fed T. occidentalis from on crude oil bioremediated lots are presented in Figures 2 - 4. 
Animals fed T. occidentalis from the control lot had healthy kidneys with perfect structures of glomeruli and 
unaltered tissue architecture Figure 2a; while those fed T. occidentalis from both bioaugmented and natural 
attenuated lots had unhealthy distorted organs with shrunken glomeruli and multicystic spaces Figure 2b - 2c. 
 

 
Figure-2. Histology Sections of Rat Kidney Fed T. Occidentalis from Crude Oil Polluted and Remediated Soil   

 
(a). Representative section of the kidney of rat fed T. occidentalis from control lot. [Glomeruli perfectly in order 
(Black pointers)]. (b). Representative section of the kidney of rat fed with T. occidentalis from bioaugmented lot. 
[Observations: Distorted tissue architecture (black pointers), multicystic spaces (blue pointers), glomeruli 
shrunken (green pointers)]. (c.). Representative section of the kidney of rat fed T. occidentalis from natural 
attenuated lot. [Mild oedema (black pointers), tissue stromal proliferation with silt-like channel (blue pointers), 
glomeruli shrunken (green pointers), distorted tissue architecture (red pointers)]. 
 

Rats fed T. occidentalis from the control lot had healthy liver sections with well identified hepatocytes and 
pronounced central vein Figure 3a. The presence of slit like channels and proliferation of tissue was noticed around 
the central veins of the sections of those fed T. occidentalis from the bioaugmented lot Figure 3b, while those fed T. 
occidentalis from the natural attenuated lot had proliferated tissue, enlarged central vein and an altered tissue 
architecture Figure 3c. 
 

 
Figure-3. Histology Sections of Rat Liver Fed T. Occidentalis from Crude Oil Polluted and Remediated Soil. 

 
(a). Representative section of the liver of rat fed T. occidentalis from control lot. [Hepatocytes clearly identified 
(Black pointers), central vein perfectly in order (Blue pointers)]. (b). Representative section of the liver of rat fed T. 



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occidentalis from bioaugmented lot. [Presence of slit like channels (Black pointers), tissue proliferation (Blue 
pointers), central vein blocked (Red pointers)]. (c) Representative section of the liver of rat fed T. occidentalis from 
natural attenuated lot. [Tissue stromal proliferation (black pointers), distorted tissue architecture (blue pointers), 
hepatocytes not clearly identified (red pointers), central vein grossly enlarged (green pointers)].  

Pancreas sections of rats fed T. occidentalis from control lot revealed a healthy, undistorted pancreas Figure 4a, 
while those fed T. occidentalis from both the bioaugmented and natural attenuated lot had scanty stroma, altered 
tissue architecture and mild oedema Figure 4b and 4c. 
 

 
Figure-4. Histology Sections of Rat Pancreas Fed T. Occidentalis from Crude Oil Polluted and Remediated Soil. 

 
(a). Representative section of Pancreas of rat fed  T. occidentalis from control lot. [Secretory cells perfectly in order 
(Black pointers)]. (b). Representative section of Pancreas of rat fed T. occidentalis from bioaugmented lot. [Scanty 
stroma (Black pointers), stromal abrasions (Blue pointers), mild oedema (Green pointers)]. (c). Representative 
section of Pancreas of rat fed T. occidentalis from natural attenuated lot. [Scanty stroma (Black pointers), pancreatic 
acini not clearly seen (Blue pointers)]. 
 

4. Discussion 
The accumulation of petroleum hydrocarbons and heavy metals in the leaves T. occidentalis of both the 

bioaugmented and natural attenuated lots can be linked to partial degradation and recalcitrant hydrocarbons 

present in the remediated lots (Tables 1, 2 and 3). This result corroborates the work of Ziółkowska and 
Wyszkowski [17], where contamination of soil with petroleum was reported not to adversely affect the vegetative 
development of edible plant by can lead to hydrocarbon accumulation in their foliar parts.   The increase in 
concentration of petroleum hydrocarbons observed in the control may be due to trans-boundary contamination 
caused by horizontal flow of hydrocarbon or due human and animal activities on the research site Tables 1 and 2. 
Moreover, Ohiri, et al. [18] reported an increase in heavy metal content of crude oil polluted and remediated 
agricultural soil and they attributed it to heavy metal presence in the petroleum. Their report also corroborates 
that of Chicarelli, et al. [19] where heavy metals like nickel and copper are present in petroleum through 
association with porphyrins. 

Ingestion is the most common route of exposure of bioaccumulated toxic chemicals from edible plants to 
humans and other animals alike. However, most toxicants diffuse through the cell membrane in the non-ionized 
form, thus the degree of ionization is important in determining chemical absorptivity [20]. The increased 
toxicological parameters recorded in this study can be attributed to the ingestion and biotransformation of plant 
accumulated hydrocarbons and heavy metals by the liver and subsequent excretion via the renal processes of the 
kidney. (Table 4 and 5). The kidney has been responsible for excretion of toxic compounds but in the process of 
excretion, compounds that are soluble in lipids (e.g. aliphatic and polycyclic aromatic hydrocarbons) are easily 
reabsorbed through the renal tubule [21], thereby causing accumulation of toxic compounds in the kidney. This 
accumulation ultimately leads to impairment in renal function, resulting in increased urea and creatinine and 
destruction of kidney cells as noticed in rats fed T. occidentalis from bioaugmented and natural attenuated lots 
Table 4 and Figures 2b and 2c. 

Toxic substances in the liver are biotransformed or secreted into the bile. However, biliary excretion of 
toxicants does not tantamount to elimination of such toxicants from the body. Reabsorption of biliary metabolites 
results into an enterohepatic cycle, thereby keeping the toxicants in the body and subjecting the livers to more 
stress conditions. This can lead to destruction of hepatic cells and subsequent elevation hepatic marker enzymes 
like alkaline phosphatase, aspartate and alanine aminotransferases in the serum. Transformation of toxicants by the 
liver may produce a potential carcinogen [22], which may subsequently lead to cell proliferation as noticed in the 
livers of rats fed T occidentalis from the bioaugmented and natural attenuated lots Figure 3b and 3c. However, the 
slight increase in serum amylase activity observed in this study indicates minimal destruction of pancreatic cells. 
Table 5 and Figures 4a - 4c.  

The pronounced reduction in packed cell volume and hemoglobin concentration recorded this study Table 6 
may be as a result of destructions of the cells of the kidney and liver. This may cause an impairment in the secretion 
of erythropoietin thereby resulting to anaemia. Erythropoietin in the presence of colony-stimulating factors (CSF) 
regulates the differentiation of stem cell in the bone marrow, which ensures that erythrocyte precursor cells are 
converted to erythrocytes [21]. Aside neurological and developmental anomalies, exposure to elevated lead 
concentrations may lead to biochemical defects in both humans and animals. The resultant effect of such 
impairment may be on kidneys, gastrointestinal tract, joints and reproductive tissues, thereby affecting 
haemoglobin synthesis [23]. This tantamount to the observed decrease in packed cell volume and haemoglobin 
concentration recorded in rats fed T occidentalis from natural attenuated and bioaugmented lots. Conclusively, the 



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hydrocarbon and heavy metal concentration in crops harvested from crude oil polluted and remediated soil has to 
be evaluated before crops from such environment can be allowed for both human and animal consumption. 
 

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