Scientia Agriculturae
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E-ISSN: 2310-953X / P-ISSN: 2311-0228
DOI: 10.15192/PSCP.SA.2016.13.1.3741
Sci. Agri.
13 (1), 2016: 37-41
© PSCI Publications
Impaired renal and cellular functions in asymptomatic
petroleum depot workers in Calabar metropolis, south-south
Nigeria
Anthony Cemaluk C. Egbuonu*, Daniel C. Nkwazema, Godwin S. Aloh
Department of Biochemistry, Michael Okpara University of Agriculture Umudike, Nigeria
*
Corresponding author email: [email protected]
Paper Information
ABSTRACT
Location-determined diet/nutrition and environmental factors affect animal
Received: 18 October, 2015
response to toxicants. This study aimed to ascertain the renal and cellular
functions status in asymptomatic occupationally exposed and control male
Accepted: 24 December, 2015
respondents in Calabar metropolis, a coastal area largely dependent on
vegetables and sea foods, using standard protocols. The serum
Published: 20 January, 2016
concentration (mmol/L) in the exposed group for hydrogen carbonate,
HCO3− (20.61±2.40) was relatively lower than in the control (21.95 ±1.64)
whereas that for sodium ion, Na+ (141.31±3.52), potassium ion, K+
Citation
(4.54±3.77) and chloride ion, Cl− (104.91±2.68), respectively were
Egbuonu ACC, Nkwazema DC , Aloh GS. 2016.
relatively higher than in the control (136.98±2.19, 3.64±0.44 and
Impaired renal and cellular functions in asymptomatic
96.27±5.09). The observations in the exposed group for serum Na +:K+
petroleum depot workers in Calabar metropolis, southratio (31.13 ± 1.14), urinary creatinine clearance (µmol/L) (85.24 ± 9.65)
south Nigeria, 13 (1), 37-41. Retrieved from
and urinary creatinine concentration (µmol/L) (115.67 ± 10.27),
www.pscipub.com
(DOI:
respectively were relatively lower than in the control (37.63 ± 1.03, 91.88
10.15192/PSCP.SA.2016.13.1.3741)
± 7.97 and 119.96 ± 10.97). The serum concentration for urea (3.19 ± 0.84
mmol/L), creatinine (107.65 ± 8.17 µmol/L), uric acid (0.31 ± 0.05
mmol/L) and urea:creatinine ratio (0.03 ± 1.32) respectively in the control
were relatively lower than that in the exposed group (4.27 ± 1.28 mmol/L,
117.35 ± 7.87 µmol/L, 0.44 ± 0.07 mmol/L and 0.04 ± 0.81) The
observations apart from that for K+ were significant (p<0.05). Thus, the
study suggests impaired renal and cellular functions in the petroleum depot
workers, compares with that of similar but earlier studies and negates
possible location-determined diet/nutrition and environmental factorsassociated modulatory influence on the depot workers response to
petroleum products fumes intoxication.
© 2016 PSCI Publisher All rights reserved.
Key words: Renal function, cellular function, petroleum depot workers, uric acid, creatinine clearance
Introduction
Petroleum products vapour could persist in the environment (Schecter et al., 2006) and as pollutants could cause
adverse effect on the environment and in humans (Marilena and Elias, 2008) with varying severity based on the chemical
nature, concentration and persistence of the pollutant (Tietenberg, 2006). Thus, occupational exposure to petroleum
products vapour will have significant adverse health implications. Depot workers in Calabar metropolis in Nigeria do not
wear the personal protective kits which could worsen the adverse effects of petroleum products vapours exposure and the
attendant health risks. Earlier studies abound (Awodele, et al., 2014; Al-Helaly and Ahmed, 2014; Festus, et al., 2013;
Mahmood, et al., 2013), but were conducted in different locations.
However, it is a common knowledge that many factors, including location-determined diet/nutrition and
environmental factors, may affect animals response to toxicants. Generally, Calabar, a metropolis in Cross River State,
South-South, Nigeria, is a coastal area with the inhabitants largely inclined to eating vegetables and sea foods. Recent and
similar studies (Egbuonu and Nkwazema, 2015; Egbuonu et al., 2015) suggested adverse influence in the liver,
cardiovascular risks and impaired lipid metabolism.
These warranted this study aimed at ascertaining the renal and cellular functions status in petroleum depot
workers in Calabar metropolis, Nigeria, via set objectives, including the determination of urea, creatinine, chloride,
sodium, hydrogen carbonate and potassium ions concentration in the serum and, for creatinine only, in the urine, and
calculation of diagnostic ratios (Na+:K+ ratio and urea:creatinine ratio) from the value of corresponding serum results as
obtained in this study. The kidney is an important organ for maintaining a constant extracellular environment by adjusting
the excretion of water and electrolytes (Uboh et al., 2009). Thus, these bioindicators have been used to asses the
physiological status, including the functional capacity of the kidneys, in animals (Egbuonu and Ezeanyika, 2013; Egbuonu
Sci. Agri. 13 (1), 2016: 37-41
and Osakwe, 2011; Egbuonu et al., 2010; Uboh et al., 2009; Egbuonu et al., 2009; Zannan et al., 2008; Nwankwo et al.,
2006; Gidado et al., 2001).
Materials and Methods
Chemicals and equipment
The chemicals and equipment used were provided courtesy of the Chief Technologist, Chemical Pathology
Laboratory, University of Calabar Teaching Hospital, Calabar Cross River State Nigeria. The chemicals were of analytical
grade and products of reputable companies including Sigma-Aldrich, Germany and British drug House, London.
Experimental design
The participants, aged between 18 and 35, were asymptomatic male workers in 10 petroleum depots that do not
provide the personal protective equipment and male University of Calabar students (for 2 to 4 years) in Calabar
metropolis, Nigeria who are largely inclined to vegetable and sea food diets. The study adopted the sampling method of
Lindell et al. (2001) and a standard questionnaire (Lippincot and Wilkins, 2000). The respondents were informed and their
oral consent obtained before the administration of questionnaire and collection of blood and urine samples. Ethical
clearance was sought, and approval obtained, from the Ethical Committee of Biochemistry Department, Michael Okpara
University of Agriculture Umudike, Nigeria. Biochemical tests were conducted at the Chemical Pathology Laboratory of
University of Calabar Teaching Hospital.
Sample size determination and samples collection
The determination of the sample size for this study, sixty four (64), and blood sample collection to obtain the
serum was as described earlier (Egbuonu et al., 2015; Egbuonu and Nkwazema, 2015). Urine sample of each subject was
collected into wide mouth, screw-capped, transparent containers.
Biochemical tests
Flame photometric determination of serum sodium and potassium ions concentration
The concentration of sodium and potassium ions in the serum was determined by flame photometric estimation of
sodium and potassium, using compressed air. Dilute serum was sprayed as a fine mist of droplets (nebulised) into a nonluminous gas flame which emits the characteristic golden or lilac colour. Light of a wavelength corresponding to metal
being measured (sodium or potassium) was selected by a light filter and allowed to fall on a photo sensitive detector. The
amount of light emitted is proportional to the concentration of metallic ions present.
In brief, 1:200 dilutions each of the sample and the standard for sodium and potassium were made in a universal
container by diluting them with deionised water on the flame photometer and allowed to warm for 15 minutes. The
photometer reading was set at zero by using deionised water as blank. The equipment was calibrated with the diluted
solutions of the standards to give 140 mmol/L for sodium and 5.0 mmol/L for potassium. The various test sample
concentrations were then read on the digital read-out.
Determination of serum chloride ion concentration
The chloride ion concentration in the serum was determined by the method of Schales and Schales (1941). The
principle was based on the colorimetric estimation of chloride which when titrated with mercuric nitrate solution, using
diphenylcarbazone as indicator, gives a violet-blue complex that is measured colourimetrically and calculated using the
relation:
Chloride ion concentration =
Titration of test
×
Titration of standard
conc. of standard
1
Determination of serum hydrogen carbonate (bicarbonate) ion concentration
The hydrogen carbonate ion in the serum was determined by the method of Van Slyke and Neill (1924). The
principle was based on the colorimetric estimation of hydrogen carbonate in serum which when reacted with excess dilute
H2SO4 releases CO2 whereas the remaining H2SO4 on titration with NaOH using neutral red as an indicator gives an
orange coloured end point. The corresponding bicarbonate ion concentration was calculated using the relation:
Serum hydrogen carbonate ion concentration = 1 – titre of test × 50 mmol/L
Determination of creatinine concentration in serum and urine
The creatinine concentration in serum and urine was determined by the method of Barclay and Kenny (1947).
This was based on the principle that picric acid (50 mmol/L) could remove protein from the serum sample while the
creatinine in the resulting supernatant could react with creatinine buffer to form a yellow-orange colour. The absorbance of
the yellow-orange could be read at 520 nm, and the creatinine concentration (in µmol/L) calculated from the relation:
Absorbance of test
× conc. of standard
Absorbance standard
1
38
Sci. Agri. 13 (1), 2016: 37-41
The urinary creatinine clearance was calculated with Cockroft formula as in Egbuonu and Ezeanyika (2013), but for males
and with assumption of uniform age.
Serum urea concentration determination
The serum urea concentration was determined by the method of Natelson (1951). This was based on the principle
that urea could react with substances such as diacetyl in the presence of thiosemicarbazide and cadmium ions in acid
conditions to form a rose-purple coloured solution. The absorbance of the resultant coloured solution could be measured at
540 nm, and the serum urea concentration (mmol/L) calculated from the relation:
Absorbance of test × conc. of standard
Absorbance of standard
1
Diagnostic ratios
Diagnostic ratios, including Na+:K+ ratio and urea:creatinine ratio were calculated from the value of
corresponding serum results as obtained in this study.
Data analysis
Data were analyzed by simple student t- test using statistical package and service solutions (SPSS) for windows
version 7. Mean differences at p<0.05 were considered statistically significant. Results were presented as mean ± standard
deviation (SD).
Results
The serum concentration (mmol/L) in the exposed group for hydrogen carbonate, HCO3− (20.61±2.40) was
relatively lower than that in the control (21.95 ±1.64) by 6.11 % whereas that for sodium ion, Na + (141.31±3.52),
potassium ion, K+ (4.54±3.77) and chloride ion, Cl− (104.91±2.68), respectively were relatively higher than that in the
control (136.98±2.19, 3.64±0.44 and 96.27±5.09) by 3.16 %, 24.73 % and 8.94 % (Table 1). The observations apart from
that for K+ were significant (p<0.05).
The observation in the exposed group for serum Na+:K+ ratio (31.13 ± 1.14) in Table 1, urinary creatinine
clearance (µmol/L) (85.24 ± 9.65) and urinary creatinine concentration (µmol/L) (115.67 ± 10.27) (Table 2), respectively
were relatively lower (p<0.05) than that in the control (37.63 ± 1.03, 91.88 ± 7.97 and 119.96 ± 10.97) by 17.27 %, 7.22 %
and 3.58 %.
The serum concentration for urea (3.19 ± 0.84 mmol/L), creatinine (107.65 ± 8.17 µmol/L), uric acid (0.31 ± 0.05
mmol/L) and urea:creatinine ratio (0.03 ± 1.32) respectively in the control were relatively lower (p<0.05) than that in the
exposed group (4.27 ± 1.28 mmol/L, 117.35 ± 7.87 µmol/L, 0.44 ± 0.07 mmol/L and 0.04 ± 0.81) by 33.85 %, 9.01 %,
41.94 % and 33.33 % (Table 2).
Table 1. Serum Na+, K+, Cl− and HCO3− concentration and calculated Na+:K+ ratio in the control and exposed groups
Parameters
Na+ concentration
K+ concentration
Cl- concentration
HCO3− concentration
Na+:K+ ratio
Unit
mmol/L
mmol/L
mmol/L
mmol/L
Control Group
136.98 ± 2.19
3.64 ± 0.44
96.27 ± 5.09
21.95 ± 1.64
37.63 ± 1.03
Exposed Group
141.31 ± 3.52*
4.54 ± 3.77ns
104.91 ± 2.68*
20.61 ± 2.40*
31.13 ± 1.14
Difference relative to control (%)
+3.16
+24.73
+8.97
−6.11
−17.27
Result= Mean value ± SD for sample size, n = 64 *The difference was significant (p < 0.05). ns The difference was not significant
(p>0.05). + or – sign prefix denoted ‘increased by’ or ‘decreased by’, respectively.
Table 2. Serum urea, creatinine and uric acid concentration, calculated serum urea:creatine ratio and urinary creatinine clearance and
urinary creatinine concentration in the control and exposed groups
Parameters
Urea concentration
Creatinine concentration
Uric acid concentration
Urea:Creatinine ratio
Urinary creatinine clearance
Urinary creatinine concentration
Unit
mmol/L
µmol/L
mmol/L
µmol/L
µmol/L
Control Group
3.19 ± 0.84
107.65 ± 8.17
0.31 ± 0.05
0.03 ± 1.32
91.88 ± 7.97
119.96 ± 10.97
Exposed Group
4.27 ± 1.28*
117.35 ± 7.87*
0.44 ± 0.07*
0.04 ± 0.81*
85.24 ± 9.65
115.67 ± 10.27
Difference relative to control (%)
+33.85
+9.01
+41.94
+33.33
−7.22
−3.58
Result= Mean value ± SD for sample size, n = 64 *The difference was significant (p < 0.05). ns The difference was not significant
(p>0.05). + or – sign prefix denoted ‘increased by’ or ‘decreased by’, respectively.
Discussion
Generally, location-determined diet and environmental factors affect animal response to toxicants, warranting this
study in asymptomatic occupationally exposed and control male respondents in Calabar metropolis, a coastal area largely
dependent on vegetables and sea foods. The serum concentration for urea, creatinine and uric acid in the exposed group
were relatively higher than that in the control group, suggesting impaired excretion of these waste products that could
either cause or worsen renal impairment (Kuehi et al., 2000; Edmunds et al., 2001; Sheth et al., 2006). These wastes along
39
Sci. Agri. 13 (1), 2016: 37-41
with physiological electrolytes are usually eliminated through the urine, and their impaired elimination probably resulted
in the higher serum electrolytes concentration in the exposed group observed in this study. In particular, high serum urea
concentration suggested impaired kidney functions (Egbuonu et al., 2013; Siegel et al., 2004; Carrieri et al., 2006) and the
present result agrees with that of earlier studies (Nwanjo and Ojiako, 2007; Naza et al., 2013; Mahmood, et al., 2013;
Awodele, et al., 2014) but, respectively in petrol/gasoline station workers and among petrol tanker drivers. Also, high
serum uric acid concentration suggested renal impairment, in agreement with the result of earlier study (Uboh, et al., 2009)
though in rats exposed to kerosene and gasoline vapours. It could, in addition, suggest enhanced tissue breakdown and
release of intracellular nucleotides (Dykman and Simon, 1987), perhaps in apparent response to petroleum products
vapours intoxication.
The significantly higher serum creatinine in the exposed depot workers as observed in this study agrees with that
in gasoline filling station workers (Naza et al., 2013), petrol station workers (Festus, et al., 2013; Nwanjo and Ojiako,
2007), motor mechanics (Bartimaeus and Jacobs, 2003) and in rats exposed to kerosene and gasoline vapours (Uboh et al.,
2009), suggesting impaired kidney functions due probably to enhanced production of creatinine. This may further suggest
depletion in the intracellular arginine concentration and an inhibition of protein synthesis. Creatinine is a breakdown
product of creatine whereas arginine is utilized in creatine biosynthesis (Markus and Rima, 2000) and concomitant
decrease in the mRNA (involved in protein synthesis) content following high creatinine concentration has been reported
(Guthmiller et al., 1994). The possible resultant depletion of arginine (a semi-essential amino acid that is very essential in
nitric oxide synthesis) and inhibition of protein synthesis (vis a vis higher protein turnover suggested in this study) may be
fundamental steps in the petroleum products vapour related toxicity, warranting follow up.
Electrolyte status provides an indication of the renal integrity and cellular functions in animals (Uboh et al.,
2009). The decrease in HCO3− observed in this study contrasted with the increase reported earlier, but in petrol station
attendants in Owerri, Nigeria (Festus, et al., 2013). Generally, hydrogen carbonate, HCO3− maintains the acid-base
balance, hence the observed lower concentration in the serum of the exposed group may be indicating compromised
physiological buffering integrity in the exposed group, perhaps in response to apparent adverse effect following higher
concentration and longer exposure of the depot workers to petroleum products. The other studied serum electrolytes were
higher in the exposed group than in the control. The observation was, aside that of K +, significant (p<0.05), seemingly
indicating decreased excretion of these electrolytes (due perhaps to kidney and cellular dysfunctions) that may result in
pathological conditions. For instance, high blood Na+ and Cl− concentration was associated with increased blood pressure
(Overlack et al., 1993; Boegehold and Kotchen, 1991). In contrast to the result of the present study, Festus et al. (2013)
and Uboh et al. (2009) reported a decrease in the Na+ and Cl- concentration but in petrol station attendants and in rats
exposed to kerosene and gasoline vapours, respectively. The higher Na+ and Cl- concentration in the exposed group of this
study could be due to the higher concentration of, and longer exposure to, the petroleum products (Okoro et al., 2006) that
could adversely alter the electrolyte balance of the unprotected petroleum depot workers.
The observation in the exposed group for the calculated serum Na+:K+ ratio was relatively lower (p<0.05) than
that in the control by 17.27 %, suggesting compromised Na+:K+ ATPase activity that may result to the general body
weakness complained by the exposed subjects. The higher Na+ and K+ observed in the exposed group could up-regulate the
activity of ATP-dependent Na+:K+ ATPase pump that could, over time, deplete the available ATP consequently resulting
in compromised Na+:K+ ATPase activity and general body debility. Urea:creatinine ratio was higher (p<0.05) in the
exposed group by 33.33 %, suggesting higher protein turnover (Egbuonu et al., 2010) and decreased muscle mass
(Feinfeld et al., 2002) due perhaps to abnormal protein catabolism following compromised kidney integrity. The relatively
lower (p<0.05) urinary creatinine concentration and creatinine clearance in the exposed than in the control suggested
decreased muscle mass (Egbuonu and Ezeanyika, 2013) which may increase mortality risk in the exposed subjects. Low
muscle mass independently predisposed, though older, men to high mortality (Wannamethee et al., 2007).
Conclusion
The study suggests impaired renal and cellular functions in the petroleum depot workers, compares with that of
similar but earlier studies and thus, negates possible location-determined diet/nutrition and environmental factorsassociated modulatory influence on the depot workers response to petroleum products fumes intoxication. The study
underscores the need for the workers to wear personal protective equipment and to regularly assess their health status.
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