Introduction:
Uziza seed is use to give a peppery flavor, this spice
is used in soups and stews. In particular banga soup and pepper soup. The
scientific name is piper guineense. According to[1] piper
guineense is a West African species of piper, the spice derived from its
dried fruit is known as West African pepper, Ashanti pepper, Benin pepper,
false cubeb, Guinea cubeb, Uziza pepper. Uziza seed is a close relative of
cubeb pepper and a relative of black pepper and long pepper. The seed is
prolate spheroids, small and smooth with reddish tinge. The uziza spice imparts
heat and a spicy, pungent aroma to classic West African soups, (stews).[2]
stated that uziza seeds have preservative and anti-oxidant properties.
Piper guineense (uziza) is a local spice that comprises of dillapiol,
5-8% of piperine, elemicine, 10% of myristine and safrole and these chemicals
exhibit bactericidal and antimicrobial effects on certain microorganisms[3].
Useful chemicals have being found embedded in uziza
seed and these chemicals exhibits bactericidial and antimicrobial effects on
certain microbes, these effects is associated with the presence of
phytochemicals like flavonoids, alkaloids, tannins, saponins, phenols,
phytates, steroids and essential oil.
According to[4], tannins are phenolic
compounds with proline-rich proteins that helps to inhibit the absorption of
iron when present in the gastro-intestinal lumen. Plants and herbs that
contains tannins as their primary components are astringent, thus highly
beneficiary for treating intestinal disorders. Alkaloids are natural products
present in p. guineense ‘uziza’. Alkaloids are made up of heterocyclic
nitrogen that possess antimalarial, pharmacological, antihypertensive,
antiarrhythymic and anticancer effects. Flavonoids on the other hand are
remarkable for their countless health benefits to human beings, especially due
to their antiplatelet, antiviral, anti-tumor, anti-oxidant and
anti-inflammatory abilities[5]. Researchers have equally identified
other phytochemicals in uziza seed, such phytochemicals include saponin,
steroid, phytate, phenol, and essential oil[6]. Phenols acts as
antifeedants, antioxidants and protective against uv light among others. They
are in general contribute to the bitterness associated to the astringency of
phenolics like tannins. Available research has reviewed that vitamins are
essential nutrients needed by the body to enhance good health.[7] classified
vitamins as either water-soluble or fat-soluble. According to him, fat soluble,
vitamins are (A,D,E and K) and water soluble vitamins in number (8B vitamins
and vitamin C). Research has reviewed that vitamins B1, B2, B3,A,
E and C are all present in uziza seed.However, minerals like Ca, Mg, Na, K, Fe
and P have all been identified in uziza seed (piper guineense). All
those vitamins and minerals are reason behind the health benefits of uziza
seed.
MATERIALS AND
METHODS:
The materials used areuziza seed, weighing balance,
oven, and beakers, moisture can, measuring cylinder, incinerator, test-tubes,
test-tube holder, water bath, dropper, flat bottomed flasks, stirrer, funnel,
crucible, funnel, spectrophometer, volumetric flask, desiccator, fume cupboard,
filter paper, kjeldahl distiller and flame photometer.
REGEANTS USED:
Acetic anhydride, ethanol, sulphuric acid (H2SO4),
distilled water, ferric chloride, pirovic acid, concentrated ammonia,
concentrated HCl, amyl alcohol, NaOH, EDTA, methyl red, vandatemolybate,
ammonia buffer, solochromedarkblue, Erichrome black T, hydroxylamine,
hydrochloride, phenolphthalein, potassium, hydroxide, isopropyl alcohol,
ethanolic sodium hydroxide, potassium dichromate, hydrogen peroxide, sodium
sulphate, potassium cyanide, starch indicator, and copper tetraoxosulphate
(VI).
Determination of Moisture:
This was done byAmerica Organization of Analytical
Chemist method. 10 g of each sample was measured into a pre-weighed moisture
can. The sample in the can was dried in the oven at 105
for 3 hours. It was cooled in a desiccator
and weighed. It was returned to the oven for further drying after which it was
left to cool and weighed repeatedly for an hour intervals until a constant
weight was obtained. The weight of the moisture lost was calculated as a
percentage of weight of sample analyzed. It was given by the expression.
% moisture content = 
Where
w1 = weight of empty moisture
W2 = weight of moisture can and sample
before drying
W3= weight of moisture can and sample dried
on constant weight.
Determination of Ash of Content:
This was done by furnace incineration according to the
America Organization of Analytical method. 3 g of the processed sample (uziza)
was poured into a previously weighed porcelain crucible. The sample was burnt
to ashes in a muffle furnace in a desiccator and weighed. The weight of the ash
was expressed in percentage of weight of sample analyzed as shown below:
% Ash= 
Where, 
= weight of empty crucible,
= weight of crucible + ash
Determination of Crude Fibre Content:
This is done by the America Organization of Analytical
Chemist AOAC method. 3 g of each processed samples was boiled in 150 ml of
1.25% H2SO4 solution for 30 min under reflux. The boiled
sample was washed in several portions of hot water using a two-fold muslin
cloth to trap the particles which were returned back to the flask and boiled
again in 150 ml of 1.25% NAOH for another 30 min under the same condition.
After washing in several portions of hot water, the samples was allowed to
drain dry before being transferred to a weighed crucible where it was dried in
an oven at 105 to a constant weight. It was burnt to
ashes in a muffle furnace. The weight of fibre was calculated as a percentage
of weight of sample analyzed.
Determination of Crude Protein:
This was done by kjeldahl method. The total N2was
determined and multiplied with factor 6.25 to obtain the protein content. 1.0 g
of processed sample was mixed with 10 ml of concentrated H2SO4
in a digestion flask. A tablet of selenium catalyst was added to it before it
was heated in a fume cupboard until a clear solution was obtained (i.e. the
digest) which was diluted to 100mls of the digest was mixed with equal volume
of 45% NaOH solution in a kjeldahl distillation apparatus. The mixture was
distilled into 10 ml of 4% buric acid containing 3drops of mixed indicator
(methyl red). A total of 5 ml of distilleries was collected and titrated
against 0.02M EDTA from green to deep red end point. The N2 content
and hence the protein content was calculated using the formular below:
% protein= %N2 x 6.25
% N2 = (
W= weight of sample,
N= normality of titrant (0.02) H2SO4,
Vt= total digest volume (100mls),
Va= volume of digest analysed (10ml),
T= titre value of sample,
B= titre value of blank.
Determination of Phosphorus Content in the Three
Spices:
This was determined by the molybdo vanadate method. A
measured volume of dry ash digest (2 mg) of samples was dispersed into a 50 ml
volumetric flask. The same volume of distilled water and standard phosphorus
solution were measured into different flask to serve as a regent blank and standard
respectively. 2 ml of phosphorus colour reagent (molybdo vanadate solution) was
added to each of the flasks and allowed to stand at room temperature for 15
min. The content of the flask was diluted to the 50 ml mark with distilled
water and its absorbance was measured in spectrophotometer at a wavelength of
540 nm with the regent blank at zero.
The phosphorus content was calculated, using the
formula;
P mg/100 g = 
Where, W= weight of ashed sample
Au= absorbance of test sample,
C = concentration of standard phosphorus solution
Vt = total volume of extract,
Va= volume of extract analyzed
Determination of Potassium and Sodium Content:
This was done using the flame photometry method. Jaway
digital flame photometer was set up according to the manufacturer’s
instruction. It was switched on and allowed about to 10 to 15 minutes to
equilibrate. Standard sodium and potassium solution were prepared separately
and diluted in series to contain 10, 8, 6, 4, and 2 g of sodium and potassium
respectively.
After equilibrating the instrument, 1 ml of each
standard was aspirated into it and sprayed over the non-luminous flame. The
optical density of the resulting emission from each standard solution was
recorded. Before filtrating, the appropriate element (Na and K) was put in
place with the standard course which was used to extrapolate the content of
each test element and calculated as shown below;
Na or K (mg/ 100g) = 
Where;
X = concentration of test elements from the curve.
Concentration of Calcium and Magnesium Content:
This was done by using the versanate EDTA titrimetric
method. 20 ml portion of the extract was dispersed into a conical flask and
treated with pinches of making agents (hydroxylamine hydrochloride, sodium
potassium ferrocyanide). The flask was shaken and the mixture dissolve. 20 ml
of ammonia buffer was added to it to raise the pH to 10.00 (a point at which
calcium and magnesium forms complexes with EDTA solution using crochorme
black-t as indicator. A regent blank was also done from deep red to permanent
blue end point. The titration value represents both Ca2+ and Mg2+
in the test sample.
A repeated sample was done to determine Ca2+
alone in the test sample. This was done in similarity with the above titration.
However, 10% NaOH was used in place of crochrome black-T. From the above values
obtained, the Ca2+ and Mg2+ content were calculated as
follows.
Ca/Mg (mg/100 g) = 
Where;
W= weight of sample, T= titre value of sample, B =
titre value of blank, Ca= calcium equivalent, Mg = magnesium equivalent, N=
normality of titrant (0.02N EDTA).
Test for the Presence of Alkaloid in the three Spices:
5 g of each sample was weighed into a 250 ml beaker
and 200 ml of 20% acetic acid in ethanol was added and covered to stand for 4
hours. This was the filtered and extract was concentrated using a water-bath to
one-quarter of the original volume. Concentrated ammonium hydroxide was added
drop wise to the extract until the precipitation was complete. The whole
solution was allowed to settle and then filtered. The precipitate was dried and
weighed [8-9]. The alkaloid content was calculated in percentage;
= 
Test for the Presence of Saponin:
20 g of each of the samples was dispersed in 200ml of
20% ethanol. The suspension was heated over a hot water bath for 4 h with
continuous stirring at about 55. The mixture was filtered and the residue
re-extracted with another 200ml of 20% ethanol. The combined extracts were
reduced to 40 ml over water-bath at about 90. The concentrate was transferred into 250
ml separator funnel and 20 ml of diethyl ether was added and shaken vigorously.
The aqueous layer was discarded. The purified process was repeated. 60 ml of
n-butanol was added. The combined n-butanol extract were washed twice with 10
ml of 5% aqueous sodium chloride. The remaining solution was heated in a water-bath.
After evaporation, the samples were dried in the oven to a constant weight.
=
Test for the Presence of Tannin in the Three Spice:
0.5g of the sample was extracted with distilled water.
It was shaken for 30minutes at room temperature and filtered. A standard tannic
acid solution was prepared. 2mls of the standard solution and equal volume of
distilled water were dispersed into a separate 50 ml volumetric flask to serve
as standard and regent blank respectively. Then 2mls of each of the samples
extract was mixed with 35 ml distilled water in 50 ml volumetric flask. 1.0ml
of the Folin-Denis regent was added followed by addition of 2.5 ml of saturated
Na2CO3solution and then distilled water. Shake the
mixture, incubate at room temperature for 90 min. Take the absorbance at 760 nm
with spectrophotometer.
= 
Where,
= absorbance of the test sample, 
= absorbance of blank, 
= concentration of the standard tannin
solution, 
= total volume of extract, 
= volume of extract analyzed, 
= weight of the sample used.
Determination of Phenol:
0.2g of the sample was dissolved in methanol and
filtered to extract phenol. 1ml of the filtrate was mixed with 1ml of
folin-ciecateau regent and 2 ml of 20% Na2CO3 solution
was added. The intensity of the developed colour was measured using
spectrophotometer at 560 nm. The standard phenol was treated the same way.
= 
Where;
= absorbance of the test sample,
Ab = absorbance of blank,
As = absorbance of standard phenolic solution,
C = concentration of standard phenolic solution,
D = dilution factor if any.
Test for Flavonoid:
10 g of sample was extracted, repeated with 100ml of
80% aqueous methanol at room temperature. The whole solution was filtered
through No 42 whatmann filter paper. The filtrate was later transferred into a
crucible and evaporated to dryness over a water bath and weighed to a constant
weight.
% Flavonoid = 
Test for steroid:
5g of each spice was hydrolyzed by boiling in 50ml
hydrochloric acid solution for about 30minutes. It was filtered paper, the
filtrate was transferred to the separating funnel. Equal volume of ethyl
acetate was added to it, mixed well and allowed to separate in two layers was
discharged. The extract was dried at 100oC for 5minutes in a steam
bath. It was then heated with concentrated amyl alcohol to extract the steroid.
The mixture becomes turbid and a reweighed Whatmann filter paper was used to
filter the mixture properly. The dry extract was then cooled in a desiccator
and weighed. The process was repeated two times and an average was obtained.
The concentration of steroid was determined and
expressed as a percentage thus
% steroid = 
Where,
W1 = weight of filter paper
W2= weight of paper + steroid
Test for Phytate:
The plant material was extracted with 0.2 M HCl such
that we have (3-30
m-1phytate solution). 0.5 ml of
extract was pipette into a test tube fitted with a ground glass stopper. 1ml of
ferric solution was added, the tube covered with the stopper and fix if with a
clip. The tube was heated in a boiling water bath for 30 min. Care was taken to
ensure that the first 5 min that the tube remains well stopped. After cooling
in ice water for 15 min, it was allow to adjust to room temperature. Once the
tubes have reached room temperature, the content of the tube was mixed and
centrifuge for 30 min at 3000 g. 1 ml of the supernatant was transferred to
another test tube and added to 5 ml of 2, 2-Bipyridine solution. The absorbance
was measured at 519 nm against distilled water. Then calibration with the
reference solutions as a substitute for the sample solution with each set of
analysis was made, then the absorbance of the test sample is used to obtain the
concentration from the calibration curve.
RESULTS AND
DISCUSSIONS:
The important phytochemicals of Piper guineense
seeds are shown in Table 4.1, which include alkaloids, saponins, flavonoids,
phenols, steroids, phytates and tannins. Some of the phytochemicals have
established medicinal values[10-12]. However, the steroids content
of Piperguineense seeds is 0.06 
0.00% for alkaloids. Alkaloids can
elucidate a wide range of physiological activities in the body when consumed
and are therefore widely applied in medicine. The percentage flavonoid in Piper
guineense seeds is 0.26 
0.00 %, this particular chemical
serves to protect the body against oxidative cells destruction through their
antioxidant activity in scavenging free radicals. Flavonoids are remarkable for
their countless health benefits to human beings, especially due to the
antiplatelet, antiviral, anti-tumor, antioxidant and anti-inflammatory
abilities.[13] also agreed that flavonoids are protective agent
inflammatory disorders. The saponins content of Piper guineense seeds is
found to be 0.25 ± 0.00% .This property gives them the ability to foam[14].
Table 4.1:Phytochemical composition ofPiper
guineense seed
|
Phytochemical
Composition
|
Piper
Guineense(Uziza)
|
|
Phytate
|
0.39 ± 0.01
|
|
Tannin
|
1.30 ± 0.01
|
|
Saponin
|
0.25 ± 0.00
|
|
Flavonoid
|
0.26 ± 0.00
|
|
Alkaloid
|
1.58 ± 0.01
|
|
Phenol
|
0.01 ± 0.01
|
|
Steroid
|
0.06 ± 0.00
|
|
Total
|
0.45 ± 0.56
|
Values are means
and standard deviations of three replicates
Table 4.2: Proximate
composition of Piper guineenseseed
|
Proximate
Composition
|
Piper Guineense(Uziza)
|
|
Moisture Content (%)
|
12.41 ± 0.06
|
|
Dry Matter (%)
|
87.60 ± 0.06
|
|
Ash (%)
|
8.75 ± 0.01
|
|
Crude Fibre (%)
|
2.67 ± 0.02
|
|
Ether Extract (%)
|
10.53 ± 0.11
|
|
Crude Protein (%)
|
6.67 ± 0.05
|
|
Total
|
8.76 ± 3.21
|
Values are means
and standard deviations of three replicates
Table 4.3: Vitamin composition of Piper
guineenseseeds
|
Vitamins
|
Piper
Guineense(Uziza)
|
|
Thiamine (B1)
|
0.02 ± 0.00
|
|
Riboflavin (B2)
|
0.09 ± 0.00
|
|
Niacin (B3)
|
0.08 ± 0.00
|
|
Retinol (A)
|
8.36 ± 0.00
|
|
Tocopherol (E)
|
6.46 ± 0.08
|
|
Ascorbic Acid (C)
|
2.52 ± 0.12
|
|
Total
|
2.92 ± 3.49
|
Values are means
and standard deviations of three replicates
Table 4.4: Mineral composition ofPiper
guineenseseed
|
Mineral
Composition
|
Piper guineense(Uziza)
|
|
Calcium
|
191.60 ± 0.57
|
|
Magnesium
|
25.16 ± 0.02
|
|
Sodium
|
19.60 ± 0.007
|
|
Potassium
|
19.60 ± 0.07
|
|
Iron
|
2.48 ± 0.06
|
|
Phosphorus
|
212.08 ± 0.16
|
|
Total
|
127.53 ±
129.53
|
Values are means
and standard deviations of three replicates
Table 4.5: Trace
metal composition of Piper guineenseseed
|
Trace metal
|
Piper
guineenseseeds
(ppm)
|
|
Pb(II)
|
0.240
|
|
Cr(III)
|
0.066
|
|
Zn(II)
|
0.750
|
|
Fe(II)
|
1.930
|
|
Cd(II)
|
0.026
|
Table 4.2 shows the proximate compositions of Piper
guineense(uziza seed), Table 4.5 shows trace metal composition of Piper
guineense(uziza seed), while Table 4.3 shows the amount of vitamins present
in Piper guineense and the vitamins include vitamin A, B1, B2,
B3, C, and E. The relatively high concentration of vitamin A
makes it good for the body, this vitamin is responsible for improved vision
etc.Vitamin C has anti-infective properties, promotes wound healing, may boost
the immune system and may help ward off infections [15]. The role of
these B-vitamins cannot be overemphasized. B1 is often called an
anti-stress vitamin because of its ability to protect the immune system. The
rich micro-nutrient content of the spices makes them beneficial and useful to
the physiological needs of man .Table 4.4 shows the result of the minerals
present in Piper guineense seeds as a reflection of its relative high
ash content.
CONCLUSION:
This research work has revealed the importance of Piper
guineeseseed for nutritional and medicinal purposes since they are regarded
as spices. The results shows the presence of protein, crude fibre, ash,
minerals and vitamins in appreciable quantities. It revealed that the seeds are
rich in phytochemicals, as regards the presence of anti-nutrients observed, it
should be noted that most anti nutrients are destroyed or reduced to
non-significant level by normal food processing methods of washing and boiling.
Therefore, the consumers are not at any reasonable risks.
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Tannins 1.30
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