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Research Article
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Influence of Freezing on Some Phytotoxins and Micronutrients in the Leaves of Telfairia occidentalis (Fluted Pumpkin)
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Amanabo Musa
and
Emmanuel O. Ogbadoyi
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ABSTRACT
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Cold storage is one of the methods that is used to safeguard the quality of fresh foods (especially vegetables and fruits) and prevent food spoilage as the method reduces the activities of some microorganisms and enzymes that are involved in the sequence of some biochemical invents leading to deterioration reactions. Therefore, it is common practice to store fresh foods in refrigerator and freezer for some period of time. This study investigates the effect of freezing duration as it affects the concentrations of some inherent phytotoxins (cyanide, nitrate, soluble and total oxalates) and micronutrients which include vitamins (vitamin C and β-carotene; provitamin A) and mineral elements (Fe, Cu, Mg, Na and K) in the leaves of Telfairia occidentalis. The washed leaves of Telfairia occidentalis were stored in a deep freezer at-4°C for the period of four weeks and chemical analyses were done at weekly intervals. The result obtained showed that the concentrations of cyanide and Fe decreased significantly (p<0.05) with freezing time in the first and second weeks and their values remained significantly unchanged in the third and fourth week of storage. The nitrate, soluble and total oxalates and vitamin C contents in the leaves of Telfairia occidentalis decreased significantly (p<0.05) in the first week of freezing and remained significantly the same in the subsequent second, third and fourth weeks of storage. Similarly, the concentrations of β-carotene and Na in the vegetable were significantly (p<0.05) reduced in the second weeks of storage and their values obtained in the third and fourth weeks also indicated reduction which was not significantly different from the second weeks of freezing. While freezing had no significant effect on the Cu content in Telfairia occidentalis, the concentrations of Mg and K decreased significantly (p<0.05) in the third weeks and changed insignificantly in the fourth weeks of freezing. The result concludes that short period of freezing (especially one week of freezing) significantly reduced the plant toxins in Telfairia occidentalis and safeguard most of the micronutrients.
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Received:
March 12, 2013; Accepted: March 25, 2013;
Published: July 10, 2013 |
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INTRODUCTION
Telfairia occidentalis (Fluted pumpkin) is one of the leafy vegetables
that are grown across the low land humid tropic in West Africa (Nigeria, Ghana
and Sierra Leone being the major producers). Fluted pumpkin is a creeping vegetative
shrub that spread low across the ground with large lobed leave and long twisting
tendrils (Syndenham, 1985; Horsfall
and Spiff, 2005; Agatemor, 2006; Ojiako
and Igwe, 2008). Telfairia occidentalis prefers a loose, friable
ample humus and shaded position. Nitrogen is vital for sufficient vegetation
and should ideally be given in the form of manure (Schippers,
2000). Harvesting of fluted pumpkin takes place 120-150 days after planting
(Horsfall and Spiff, 2005). The vegetable is of commercial
importance in West African because of the high nutrient contents abound in it
(Nkang et al., 2003).
The leaves and seeds are widely eaten as they are good sources of micronutrient
such as mineral elements (Fe, Mg, Cu, Ca, Zn K, Na and P), vitamins (such as
vitamin C, β-carotene), fibres, fats (Schippers, 2000;
Nkang et al., 2003; Agatemor,
2006; Ogbadoyi et al., 2011; Musa
et al., 2011; Musa and Ogbadoyi, 2012a).
Telfairia occidentalis also accumulates various antinutrients and toxic
substances such as oxalate, phytate, cyanide and nitrate. These plant secondary
metabolites have been known to have deleterious effect on human and animals
health at high concentrations (Ogbadoyi et al.,
2011; Musa et al., 2011; Musa
and Ogbadoyi, 2012a). In order to maintain the quality of fresh foods and
avert food spoilage, it is a common practice to store the fresh foods in refrigerator
and freezer as this practice is known to reduce the activities of microorganisms
and enzymes that are associated with biochemical reactions leading to deterioration
of food substances. Thus this research was conducted to evaluate the influence
of freezing duration on the concentrations of some plant toxins and micronutrients
in the leaves of Telfairia occidentalis with a view to determine the
suitability of the storage conditions for the vegetable.
MATERIALS AND METHODS
Source of Telfairia occidentalis: The fresh samples of Telfairia
occidentalis was bought in three sets at different time from Maikunkele,
Bosso and Chanchanga markets in Minna town, Nigeria.
Chemicals: Except otherwise stated, all the chemicals used were of analytical
grade and were purchased from Sigma and BDH chemical companies, both of England.
Freezing: The leaves of Telfairia occidentalis were washed with
distilled water and kept in a well labelled polythene bag and stored in a freezer
at the temperature of-4°C for a period of 4 weeks. The concentrations of
the nutrients, antinutrients and toxic substances were determined at weekly
intervals over the four-weeks period.
Sample analysis: The nitrate concentration in the samples was determined
by the colourimetric method as described by Sjoberg and
Alanko (1994). While the alkaline picrate method of Ikediobi
et al. (1980) was used to analyse the cyanide content in the leaves
of Telfairia occidentalis. Both soluble and total oxalates in the samples
were determined by titrimetric method of Oke (1966).
The mineral elements (Fe, Cu, Mg, Na and K) in samples were determined according
to the method of Ezeonu et al. (2002). The ascorbic
acid concentration in the samples was determined by 2, 6-dichlorophenol indophenols
method of Jones and Hughes (1983). While β-carotene
concentration was determined by ethanol and petroleum ether extraction method
as described by Musa et al. (2010).
Statistical analysis: Analysis of variance (ANOVA) was carried out using
statistical package Minitab to determine variation between different freezing
time on the concentration of nutrients, antinutrients and toxic substances in
the leaves of Telfairia occidentralis. The Duncans
Multiple Range Test (DMRT) was used for comparison of means.
RESULTS
Cyanide concentration: Cyanide concentration was found to decrease with
freezing in Telfairia occidentalis. Significant (p<0.05) reduction
of cyanide content from 170.83-99.45 mg kg-1 was observed in the
first week of freezing. Value obtained in the second weeks (84.51 mg kg-1)
also indicated significant reduction of cyanide content which was not significantly
different from the mean values obtained for the third (70.66 mg kg-1)
and fourth weeks (69.04 mg kg-1) (Fig. 1).
Nitrate concentration: Significant (p<0.05) decreased in nitrate
content in Telfairia occidentalis from 2799.04 mg kg-1 in
fresh sample to 1514.83 mg kg-1 was observed during two weeks of
freezing. The mean values obtained for the second (1339.83 mg kg-1),
third (1072.22 mg kg-1) and fourth weeks (1035.17 mg kg-1)
also indicated reduction in concentration of nitrate which was not significantly
(p>0.05) different from the value obtained from the first week (Fig.
2).
Soluble oxalate concentration: The soluble oxalate concentration in
Telfairia occidentalis also decreased with freezing storage. Significant
reduction (p<0.05) in the parameter from 2.85 g kg-1 in fresh
sample to 2.40 g kg-1 was recorded in one week of freezing. The mean
values obtained in second weeks (2.29 g kg-1), third weeks (2.07
g kg-1) and fourth weeks (1.84 g kg-1) frozen leaves of
Telfairia occidentalis were significantly the same with that of the first
week (Fig. 3).
Total oxalate concentration: Analysis of total oxalate content in fresh
samples of Telfairia occidentalis showed that the oxalate concentration
decreased significantly (p<0.05) in one week of cold storage from 4.39 to
3.12 g kg-1.
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Fig. 1: |
Effect of freezing on cyanide content in Telfairia occidentalis.
Bars carrying the same letter are not significantly different (p>0.05) |
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Fig. 2: |
Effect of freezing on nitrate content in Telfairia occidentalis.
Bars carrying the same letter are not significantly different (p>0.05) |
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Fig. 3: |
Effect of freezing on soluble oxalate content in Telfairia
occidentalis. Bars carrying the same letter are not significantly different
(p>0.05) |
In the subsequent second, third and fourth weeks, the oxalate content remained
significantly unchanged and the values obtained were 3.12, 3.0 8 and 2.86 g
kg-1, respectively (Fig. 4).
β-carotene concentration: The β-carotene content in the fresh
sample (18859.00 μg/100 g) of Telfairia occidentalis decreased significantly
(p<0.05) in the second weeks of freezing to 16072.00 μg/100 g. The decreased
in the first week (17427.00 μg/100 g) was however, not significant. The
values obtained for the third (14941.70 μg/100 g) and fourth weeks (14690
μg/100 g) also indicated reduction in the concentration of β-carotene
which was not significantly different from those of the first and second weeks
(Fig. 5).
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Fig. 4: |
Effect of freezing on total oxalate content in Telfairia
occidentalis. Bars carrying the same letter are not significantly different
(p>0.05) |
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Fig. 5: |
Effect of freezing β-carotene content in Telfairia
occidentalis. Bars carrying the same letter are not significantly different
(p>0.05) |
Vitamin C concentration: The concentration of vitamin C in fresh sample
of Telfairia occidentalis decreased significantly (p<0.05) during
one week of freezing from 192.28 to 19.15 mg/100 g. The concentrations obtained
in the second, third and fourth weeks of freezing also indicate decreased in
the vitamin content which were insignificantly different from the first week
and the values obtained were 14.24, 12.60 and 11.50 mg/100 g, respectively (Fig.
6).
Iron concentration: The results obtained from the analysis of Fe content
in Telfairia occidentalis showed that its decreased significantly
(p<0.05) from 23.43 mg kg-1 in fresh sample to 19.55 mg kg-1
in one week of freezing.
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Fig. 6: |
Effect of freezing on vitamin C content in Telfairia occidentalis.
Bars carrying the same letter are not significantly different (p>0.05) |
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Fig. 7: |
Effect of freezing on iron content in Telfairia occidentalis.
Bars carrying the same letter are not significantly different (p>0.05) |
The mean value obtained in the second weeks (16.41 mg kg-1) was
significantly the same with the third weeks (15.42 mg kg-1) and fourth
weeks (15.29 mg kg-1) of freezing, however, these values were significantly
(p<0.05) lower than the first weeks of freezing (Fig. 7).
Copper concentration: Freezing had no significant effect on Cu concentration
in Telfairia occidentalis during the storage period. The mean values
obtained in fresh and those frozen samples for one, two, three and four weeks
were 18.70, 18.48, 18.35, 16.59 and 15.7 6 mg kg-1, respectively
(Fig. 8).
Magnesium concentration: The Mg concentration in Telfairia occidentalis
also decreased with freezing duration. Significant reduction (p<0.05) in
the mineral concentration from 48.81 mg kg-1 in fresh sample to 39.26
mg kg-1 was observed in the third weeks of storage in deep freezer.
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Fig. 8: |
Effect of freezing on copper content in Telfairia occidentalis.
Bars carrying the same letter are not significantly different (p>0.05) |
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Fig. 9: |
Effect of freezing on magnesium content in Telfairia occidentalis.
Bars carrying the same letter are not significantly different (p>0.05) |
The mean value obtained in the fourth weeks (37.57 mg kg-1) indicated
reduction in Mg content which was not significantly different from second weeks
(41.05 mg kg-1). The mineral concentration in first week (47.19 mg
kg-1) was however significantly higher compared to those in the third
and fourth weeks (Fig. 9).
Sodium concentration: Significant reduction (p<0.05) of Na concentration
in Telfairia occidentalis during freezing from 11.48 mg kg-1
in fresh sample to 8.76 mg kg-1 was observed in second weeks of storage.
The decreased in the first week (9.86 mg kg-1) was however, not significant.
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Fig. 10: |
Effect of freezing on sodium content in Telfairia occidentalis.
Bars carrying the same letter are not significantly different (p>0.05) |
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Fig. 11: |
Effect of freezing on potassium content in Telfairia occidentalis.
Bars carrying the same letter are not significantly different (p>0.05) |
The mean values obtained for the third (8.48 mg kg-1) and fourth
(8.10 mg kg-1) weeks also indicated reduction in concentration which
was not significantly different from those of the first and second weeks (Fig.
10).
Potassium concentration: The concentration of K in Telfairia occidentalis
also decreases with freezing time. Significant decreased (p<0.05) in the
mineral concentration from 183.92 mg kg-1 in fresh sample to 150.00
mg kg-1 was recorded in three weeks of storage in the deep freezer.
The mean value obtained in the fourth weeks (135.63 mg kg-1) indicated
reduction in mineral content which was not significantly different from second
weeks (161.25 mg kg-1). The K content in the first week (168.75 mg
kg-1) of storage was however, significantly higher (p<0.05) than
those in the third and fourth weeks (Fig. 11).
DISCUSSION
The decreasing effect of freezing on cyanide content in Telfairia occidentalis
agreed with the reported of Richard (1991) and McDonald
et al. (2006) to the effect that freezing ruptures the plant cells
which resulted in the release of cell contents including cyanide. The solubility
of cyanide in water was also responsible for the decreased of its concentration
during freezing (Budavari et al., 1989). Thus
cyanide may be trapped in the ice and released during thawing. The cyanide concentration
of 170.83 mg kg-1 in the fresh sample of Telfairia occidentalis
is very close to the maximum permissible level of 200 mg kg-1 of
cyanide in fresh weight (Everist, 1981; Richard,
1991) were reduced more than half of this value after one and subsequent
weeks of freezing. The result obtained indicates that the cyanide content in
the vegetable could be reduced to safe level by freezing.
The results in this study which show that there was a lower concentration of
nitrate in frozen samples than in the fresh sample of Telfairia occidentalis
is in line with the findings of Bakr and Gawish (1997),
who showed that freezing reduced the nitrate content in the vegetables. The
authors ascribed the decrease to the leaching of the cell content caused by
freezing and defrosting. The general weekly decline of nitrate content in the
vegetable during freezing is also in agreement with the submission of Bakr
and Gawish (1997), who reported that losses of nitrate during freezing storage
increase with freezing time. The nitrate concentration in fresh sample of Telfairia
occidentalis, was higher than the Acceptable Daily Intake of 220 mg (Macrae
et al., 1997) and 219 mg (Anjana et al.,
2007) for a 60 kg person (if 100 g day-1 is consumed). Thus consumption
of raw samples of the vegetable could lead to nitrate overload and could subject
the body to disease conditions associated with nitrate toxicities such as metheamoglobinaemia
and cancer (Galler, 1997; Macrae
et al., 1997; Mevissen, 1997; Mozolewski
and Smoczynski, 2004; Anjana et al., 2007).
However, one week of freezing is required to reduce the nitrate concentration
in Telfairia occidentalis to the recommended Acceptable Daily Intake.
The decreased in soluble and total oxalates concentration in the frozen leaves
compared to fresh leaves of Telfairia occidentalis recorded in this work
corroborates with the report of Ogbadoyi et al. (2006).
The authors observed that freezing of tissues at high moisture content results
in the formation of ice crystals within the cells. The sharp edges of the crystals
so formed are capable of shredding the cell membranes resulting in cell leakage.
This finding is in conformity with the report of Richard
(1991), Fellow (2000) and McDonald
et al. (2006) that freezing imparts physical injury to the cell,
alters the pigments which are sources of bioactive The soluble and total oxalates
concentration in fresh leaves of Telfairia occidentalis are more than
the permissible level of 250 mg kg-1 fresh weight as reported by
Oguchi et al. (1996). Thus regular consumption
of fresh leaves of the vegetable can cause oxalate toxicosis with attendant
health problems of kidney stone, reduction of bioavailability of elements, electrolyte
imbalance and irritation of digestive system (Antia
et al., 2006; Prohp et al., 2006; Musa
et al., 2011; Ogbadoyi et al., 2011;
Musa and Ogbadoyi, 2012a, b).Fortunately
one and two weeks of freezing reduced the soluble and total oxalates content
in Telfairia occidentalis to tolerable level, respectively.
The reduction of β-carotene content in Telfairia occidentalis during
freezing has been attributed to the physical injury and alteration of the plant
pigments (Fellow, 2000; McDonald et
al., 2006). Booth et al. (1992) stated
that the observed decrease in the β-carotene concentration in the vegetable
during storage in the freezer may be as a result of enzymatic activity coupled
with oxidation associated with conjugated double bond in the compound. Dutta
et al. (2005) observed that the loss of β-carotene in frozen
vegetables could be due to non-oxidative changes (cis-trans isomerisation, epoxide
formation) or oxidative changes on exposure light and oxygen. The insignificant
difference recorded in the β-carotene concentration in second to fourth
weeks of cold storage could be seen as a result of decrease in endogenous enzymatic
activity and reduction of oxidation of the provitamin A as freezing storage
progresses. The β-carotene concentration in the fresh and frozen leaves
of Telfairia occidentalis contained over and above the recommended adult
daily allowance of 900 μg of vitamin A (George, 1999;
Akanya, 2004). The implication of this is that because
β-carotene is abundant in Telfairia occidentalis and non-hydrosoluble,
its residual concentration in the frozen samples are high enough to meet the
normal adult recommended daily allowance. The results obtained therefore imply
that freezing the leaves of Telfairia occidentalis within the studied
periods may not require any pharmaceutical supplements of β-carotene.
The significant higher vitamin C concentration in the fresh sample of Telfairia
occidentalis than its frozen leaves is in agreement with findings of Olaofe
(1992), Yadav and Sehgal (1995), Yadav
and Sehgal (1997), Lisiewska and Kmiecik (2000),
Ejoh et al. (2005), Bergquist
et al. (2006), McDonald et al. (2006)
and Gebczynski (2006). The observation made in this study
and those of the above researchers contradicts the observations of Mathew
and Hall (1987), who found that total vitamin C concentration remained relatively
constant throughout the frozen storage period for all samples (pepper, strawberries,
green beans, etc). Olaofe (1992) attributed the reduction
of the vitamin concentration to be partly due to the enzymatic activities of
vitamin C oxidase, cytochrome oxidase and vitamin C peroxidase that were endogenously
present. The results agreed with the report of Bergquist
et al. (2006) that during freezing of vegetables the ascorbic acid
concentration decreased considerably and the dehydroascorbic acid/vitamin C
ratio increased. This finding could suggest that during cold storage, ascorbic
acid may undergo oxidation to dehydroascorbic acid by some of the endogenous
enzymes. Tosun and Yucecan (2007) added that the chemical,
biochemical and physical reactions that occurred in vegetables after harvesting
do not completely stop during frozen storage and this may lead to the oxidation
of susceptible molecules like ascorbic acid (vitamin C) while McDonald
et al. (2006) stressed that the observed decrease in the vitamin
C content during freezing is as a result of thawing. The insignificant difference
in the vitamin C content in first to fourth week of storage may be that during
the first week of freezing, the physical injury and alteration of pigments reported
by Fellow (2000) and McDonald et
al. (2006) and the enzymatic oxidation of ascorbic acid to dehydroascorbic
acid (Olaofe, 1992; Bergquist et
al., 2006) may be at the peak within one week of storage. However, these
processes might have declined considerably as freezing storage progresses. The
fresh leaves of Telfairia occidentalis could supply enough of vitamin
C to meet the recommended daily allowance of 60 mg (Olaofe,
1992; George, 1999) if 100 g of the samples are consumed.
However, Telfairia occidentalis contained some plant toxins that could
be reduced to tolerable level through freezing. Although, the residual vitamin
C concentration in the frozen leaves of the vegetable was lower than the recommended
daily allowance. Since vitamin C play important roles in human health and diseases
associated from its deficiency, pharmaceutical supplementation of the vitamin
will be necessary to augment its losses during freezing storage. This will enable
the body to meet the dietary requirement of this water soluble vitamin.
The reduction in mineral element (Fe, Cu, Mg, Na and K) concentrations in Telfairia
occidentalis with freezing time in the current study may be as results of
infliction of physical injuries to the cell wall of the leaves of the vegetable
by ice crystals that were formed during freezing which eventually leads to cell
leakage (Polo et al., 1992; Pruthi,
1999; Fellow (2000); Hui et
al., 2004; McDonald et al. (2006). Freezer
burn which also occurs during cold storage of vegetables and other soft
plant materials at the temperature of -4°C is known to decrease the bioactive
compounds including the mineral elements. The significant reduction in the mineral
elements content in some of the vegetables during freezing agrees with the report
of Hui et al. (2004), who reported that significant
amount of macro and micro mineral elements concentration in vegetables are lost
following freezing. Similarly the insignificant effect of freezing on Cu concentration
in the vegetable throughout the freezing duration also support the finding of
Polo et al. (1992) and Hui
et al. (2004). The authors stressed that, even though there was a
decreased in the mineral element in vegetables during freezing, it is not significant.
These two factions of the results obtained may suggest that the retention of
mineral elements in frozen samples or reductions of mineral in vegetables during
freezing storage depend to a great extend on the plant species/cultivars and
the form in which the mineral exist (Hui et al., 2004).
It is believed that mineral elements that exist as free ions can easily be leached
out during freezing when compared with those that are chemically bond or form
complexes with other compounds.
CONCLUSION
The results of this present study revealed that freezing the leaves of Telfairia
occidentalis at -4°C in a deep freezer for one week significantly reduced
the concentrations of the plant toxins to tolerable levels and safeguard the
micronutrients content in the vegetable. This finding thus imply that short
time freezing do not only maintaining the quality of fresh vegetable, but it
also reduces the phytotoxins content in the vegetable and thereby reduce the
incidence of disease conditions associated with ingestion of high concentration
of the phytotoxins.
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