<?xml version="1.0" encoding="UTF-8"?><!DOCTYPE article  PUBLIC "-//NLM//DTD Journal Publishing DTD v3.0 20080202//EN" "http://dtd.nlm.nih.gov/publishing/3.0/journalpublishing3.dtd"><article xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink" dtd-version="3.0" xml:lang="en" article-type="research article"><front><journal-meta><journal-id journal-id-type="publisher-id">AJAC</journal-id><journal-title-group><journal-title>American Journal of Analytical Chemistry</journal-title></journal-title-group><issn pub-type="epub">2156-8251</issn><publisher><publisher-name>Scientific Research Publishing</publisher-name></publisher></journal-meta><article-meta><article-id pub-id-type="doi">10.4236/ajac.2015.67058</article-id><article-id pub-id-type="publisher-id">AJAC-57608</article-id><article-categories><subj-group subj-group-type="heading"><subject>Articles</subject></subj-group><subj-group subj-group-type="Discipline-v2"><subject>Chemistry&amp;Materials Science</subject></subj-group></article-categories><title-group><article-title>
 
 
  Comparative Assessment of Phytochemicals, Proximate and Elemental Composition of &lt;i&gt;Gnetum africanum&lt;/i&gt; (Okazi) Leaves
 
</article-title></title-group><contrib-group><contrib contrib-type="author" xlink:type="simple"><name name-style="western"><surname>.</surname><given-names>O. Okerulu</given-names></name><xref ref-type="aff" rid="aff1"><sup>1</sup></xref></contrib><contrib contrib-type="author" xlink:type="simple"><name name-style="western"><surname>C.</surname><given-names>T. Onyema</given-names></name><xref ref-type="aff" rid="aff1"><sup>1</sup></xref></contrib></contrib-group><aff id="aff1"><addr-line>Department Pure and Industrial Chemistry, Nnamdi Azikiwe University, Awka, Nigeria</addr-line></aff><pub-date pub-type="epub"><day>17</day><month>06</month><year>2015</year></pub-date><volume>06</volume><issue>07</issue><fpage>604</fpage><lpage>609</lpage><history><date date-type="received"><day>21</day>	<month>May</month>	<year>2015</year></date><date date-type="rev-recd"><day>accepted</day>	<month>26</month>	<year>June</year>	</date><date date-type="accepted"><day>30</day>	<month>June</month>	<year>2015</year></date></history><permissions><copyright-statement>&#169; Copyright  2014 by authors and Scientific Research Publishing Inc. </copyright-statement><copyright-year>2014</copyright-year><license><license-p>This work is licensed under the Creative Commons Attribution International License (CC BY). http://creativecommons.org/licenses/by/4.0/</license-p></license></permissions><abstract><p>
 
 
  The leaves of 
  Gnetum africanum were studied for their proximate (nutrient), mineral (elemental) and phytochemical constituents. All analyses were done using standard analytical procedures and the results obtained showed a moisture content, crude protein, ash content, crude fibre, crude lipids and carbohydrates of 10.9%, 20.12%, 6.70%, 7.10%, 2.79% and 52.39% respectively. The mineral (elemental) analysis showed the presence of sodium, magnessium, calcium, iron, zinc, manganesse, potassium and copper in that order of decreasing concentration while notably absent was chromium and lead in the examined leaves. The phytochemicals detected in the leaves using various selected solvents were alkaloids, saponins, glycosides and tannins with various concentrations while flavonoids, phenols and steroids were totally absent. From the results of the aforementioned analyses, it could be concluded that the leaves of 
  Gnetum africanum contain some beneficial nutrients, mineral elements and secondary metabolites justifying the medicinal status and possible potency of the plant part.
 
</p></abstract><kwd-group><kwd>&lt;i&gt;Gnetum africanum&lt;/i&gt; Leaves</kwd><kwd> Phytochemicals</kwd><kwd> Proximate and Elemental Compositions</kwd></kwd-group></article-meta></front><body><sec id="s1"><title>1. Introduction</title><p>Gnetum africanum commonly called “Okazi” by Igbos and “Afang” by Efik of Nigeria is one of the most popular green leafy vegetables in Nigeria and is gaining popularity as a delicious food leaf in other African countries such as Cameroon, Gabon, Congo, Angola etc. [<xref ref-type="bibr" rid="scirp.57608-ref1">1</xref>] . Gnetum africanum leaves are widely consumed in the South Eastern Nigeria due to its palatability and taste and often cooked with water leaves (Talinium triangulare) to give the soup a special flavour. The plant grows as a wild evergreen climbing plant in the rain forest of Nigeria where it is searched for and highly priced in the regional markets. This wild variety is termed wild species while the recently domestically cultivated ones in the South Eastern Nigerian homes as exotic plants are termed the domestic species. Gnetum africanum belongs to the family Gnetacea and order Gnetales [<xref ref-type="bibr" rid="scirp.57608-ref2">2</xref>] . The seed of Gnetum africanum is oval in shape, small in size, greenish in colour when unripe and reddish when ripe. The seeds but most especially the leaves which are our interest in this work have shown medical efficiency in the treatment of enlarged spleen, sore throats, reduction of pain during child birth, antidotes to some forms of poison and snake bites. The seeds are specially used as fungicide for dressing fresh and septic wounds and can also be chewed raw for the management of excessive urination [<xref ref-type="bibr" rid="scirp.57608-ref3">3</xref>] .</p><p>Various chemical composition analyses have been done on these leaves to determine their proximate, phytochemicals, antinutrirional, mineral composition and other essential contents. Dike [<xref ref-type="bibr" rid="scirp.57608-ref4">4</xref>] carried out some analysis on the leaves of Gnetum africanum sourced from Umudike rainforest, Abia State while Ekpo and Eddy [<xref ref-type="bibr" rid="scirp.57608-ref5">5</xref>] studied the chemical composition of both wild and domestic species of Gnetum africanum collected from Urua Akpan Ndem in Uyo Akwa Ibom State and Akwa Ibom State University Obio Akpa Campus Teaching and Research Farms respectively. The results of the above works will be compared with those of our results obtained from species sourced from Ohianri Forest Umoli Village Mgbirichi on Ohaji Egbema L.G.A of Imo State Nigeria to discover any specific variations with respect to its chemical and elemental composition.</p></sec><sec id="s2"><title>2. Materials and Methods</title><sec id="s2_1"><title>2.1. Plant Collection, Identification and Preparation</title><p>Random samples of the leaves of Gnetum africanum (Okazi) were obtained from Ohianri forest Umoli village Mgbirichi in Ohaji Egbema L.G.A Imo state, Nigeria and identified by Mr. P.O. Ugwuozor a taxonomist of the Department of Botany, Nnamdi Azikiwe University, Awka, Anambra State, Nigeria. The leaves were removed from their stalks and air dried in the laboratory for twelve days. They were subsequently pulverized by pounding to ensure homogeneity and kept in an airtight container for further analysis. Leaves for moisture content determination were collected and analyzed in-situ.</p></sec><sec id="s2_2"><title>2.2. Proximate Analysis</title><p>The sample was analysed for moisture content, Crude protein, Crude fibre and ash content. Crude protein was determined using the kjeldahl method. The moisture and crude fat were determined using standard methods [<xref ref-type="bibr" rid="scirp.57608-ref6">6</xref>] and the percentage calculated on dry weight basis. Ash was determined by incineration in a muffle furnance and the weight of ash was calculated from difference with the sample taken and calculated on a dry weight basis. The carbohydrates were determined by difference of the sum of all the proximate composition from 100%.</p></sec><sec id="s2_3"><title>2.3. Elemental Analysis</title><p>2 g of the powerded sample was weighed into 50 ml beaker, 10 ml of conc. nitric acid (BDH analytical grade) and 10 ml of conc. perchloric acid (BDH analytical grade) were added to the sample. The mixture was put into a fume cupboard and heated until clear solution was obtained. The digested sample was filtered into a 250 ml volumetric flask using whatmann filter paper and made up to the mark with distilled water. The solution was subsequently analyzed for Ca, CU, Zn, Mg, K, Na, Pb and Cr using Atomic Absorption Spectrophotometer Varian AA 280 with appropriate hallow cathode lamps.</p></sec><sec id="s2_4"><title>2.4. Preliminary Phytochemical Screening</title><p>Phytochemcial constituents of the leaves both qualitatively using five different solvents (water, ethanol, methanol, n-hexane and diethyl ether) and quantitatively were determined according to the Harbone method for plant analysis [<xref ref-type="bibr" rid="scirp.57608-ref7">7</xref>] . The 50 ml of each solvent was used to extract the leaves separately and the presence of the phytochemicals: alkaloids, saponins, flavonoids, phenol, steroids, glycosides and tannins were tested. The phytochemicals present were estimated quantitatively.</p></sec></sec><sec id="s3"><title>3. Results and Discussion</title><p>The results as presented in <xref ref-type="table" rid="table1">Table 1</xref> showed a relatively low moisture content (10.9%) for Gnetum africanum against some values (31.6%) for Gnetum africanum seeds reported by Ekop [<xref ref-type="bibr" rid="scirp.57608-ref8">8</xref>] and some vegetables like Piper guineese and Gongromea latifolium [<xref ref-type="bibr" rid="scirp.57608-ref9">9</xref>] , 81.36% recorded in Bressica oleraecea [<xref ref-type="bibr" rid="scirp.57608-ref10">10</xref>] , 83.75% in Pterocarpus soyaubixii and Gnetum africanum [<xref ref-type="bibr" rid="scirp.57608-ref11">11</xref>] but however is within the range recorded for other vegetables like 6.82% for Cissus petiolate [<xref ref-type="bibr" rid="scirp.57608-ref12">12</xref>] and 7.60% - 8.55% for some vegetables from Nigeria [<xref ref-type="bibr" rid="scirp.57608-ref13">13</xref>] .</p><p>Moisture content which is an index of the water activity of many foods provides for greater activity of water soluble enzymes and coenzymes needed for the metabolic activities of these leaves. Higher moisture content indicates higher susceptibility to microbial attack during storage and shorter shelf life and also indicative of high total solids [<xref ref-type="bibr" rid="scirp.57608-ref14">14</xref>] [<xref ref-type="bibr" rid="scirp.57608-ref15">15</xref>] . The crude protein content of the leaves of Gnetum africanum (20.12%) was relatively high as in <xref ref-type="table" rid="table1">Table 1</xref> and compared favourably with 17.5% recorded by Epko [<xref ref-type="bibr" rid="scirp.57608-ref8">8</xref>] for the seeds of Gnetum africanum and 19.67% and 20.80% for the leaves of Gnetum africanum and C. pepo respectively [<xref ref-type="bibr" rid="scirp.57608-ref13">13</xref>] . It is however, lower than 32.95% recorded in undefatted leaves of A. hybridus [<xref ref-type="bibr" rid="scirp.57608-ref13">13</xref>] . Plant protein still remains a major source of food nutrient for the less priviledged population in developing countries including Nigeria such that protein content of the leaves makes it suitable for consumption and a rich source of vegetable protein [<xref ref-type="bibr" rid="scirp.57608-ref10">10</xref>] .</p><p>The relatively high protein content suggests the high amount of essential acids which serve as an alternative source of energy when the carbohydrate metabolism is impaired via glucogenesis [<xref ref-type="bibr" rid="scirp.57608-ref13">13</xref>] . The carbohydrate content of the sample (52.39%) as presented in <xref ref-type="table" rid="table1">Table 1</xref>, could be said to be high for a vegetable material but this value was justified by the 87.62% for Gnetum africanum seeds [<xref ref-type="bibr" rid="scirp.57608-ref8">8</xref>] , 52.32% reported for Pachira glabra and 45.92% for A. Africana seed flowers [<xref ref-type="bibr" rid="scirp.57608-ref14">14</xref>] , 52.18% for Amaranthus hybridus [<xref ref-type="bibr" rid="scirp.57608-ref16">16</xref>] . According to Emebu and Anyika [<xref ref-type="bibr" rid="scirp.57608-ref10">10</xref>] most vegetables are generally not good sources of carbohydrates. As far as vegetables are concerned, some of them are rich sources while others contain traces of the nutrients. They provide the body with a source of fuel and energy for daily activities [<xref ref-type="bibr" rid="scirp.57608-ref17">17</xref>] . The crude fibre content of 7.1% in the leaves of G. africanum in <xref ref-type="table" rid="table1">Table 1</xref> is higher than 0.80% recorded for the seeds of G. africanum [<xref ref-type="bibr" rid="scirp.57608-ref8">8</xref>] . The composition of leaves with high crude fibre may contribute to a reduction in the incidence of certain diseases like colon cancer, coronary heart disease, diabetis, high blood pressure, obesity and other digestive disorders [<xref ref-type="bibr" rid="scirp.57608-ref18">18</xref>] . From <xref ref-type="table" rid="table1">Table 1</xref>, the low fat content (2.79%) indicated that the leaves contain low quantities of lipid biomolecules [<xref ref-type="bibr" rid="scirp.57608-ref13">13</xref>] and cannot serve as main source of these biomolecules that are important for body metabolism. Epko (2007) recorded a value of 3.15% for G. africanum seeds which is still in unison with our obtained value (2.79%). The ash content of 6.70% in the leaves was also in line with the low ash content (1.2%) obtained by Epko [<xref ref-type="bibr" rid="scirp.57608-ref8">8</xref>] for the seeds of G. africanum, 4.34% for R. glabra [<xref ref-type="bibr" rid="scirp.57608-ref14">14</xref>] and 4.03% for A. Africana but lower than some vegetables such as P. mildbraedi (20.6%) [<xref ref-type="bibr" rid="scirp.57608-ref19">19</xref>] and Talinum triangulare (20.05%) [<xref ref-type="bibr" rid="scirp.57608-ref20">20</xref>] . The ash content is an indication of the mineral contents of the leaves as low ash content suggests low mineral composition or high organic components a claim verified by the results of the Elemental composition (<xref ref-type="table" rid="table2">Table 2</xref>).</p><p>From the results in <xref ref-type="table" rid="table2">Table 2</xref>, the Major elements present in the leaves were Fe, Ca, Mg and Na in that order with Cr and Pb totally absent. The absent of Cr and Pb which are toxic metals shows that these leaves do not pose any health risks. The high concentration of Ca (11.20 mg/l), a vital element that helps in bone formation and blood coagulation whose deficiency may contribute to rickets, curvature of the spine and pelvic and thoracic deformities [<xref ref-type="bibr" rid="scirp.57608-ref21">21</xref>] was also noticed. Magnessium was equally in high concentration (12.00 mg/l) which is in line with the fact that an adult human body contains about 25 grams of Magnessium and that most vegetables are rich in magnesium (more than 500 mg/kg fresh weight) [<xref ref-type="bibr" rid="scirp.57608-ref22">22</xref>] . The presence of Mg in these leaves is also known to prevent cardiomyopathy, impaired spermatogenesis and bleeding disorders [<xref ref-type="bibr" rid="scirp.57608-ref23">23</xref>] .</p><p>Iron, a key element was also present in high amount (7.23 mg/l) as in <xref ref-type="table" rid="table2">Table 2</xref> and this helps in the metabolism of almost all living organisms. In humans, iron is an essential component of hundreds of proteins and enzymes [<xref ref-type="bibr" rid="scirp.57608-ref24">24</xref>] [<xref ref-type="bibr" rid="scirp.57608-ref25">25</xref>] . The iron content of the leaves was higher than the FAO/WHO (1988) recommend dietary allowance for males (1.37 mg/day) and females (2.94 mg/day) [<xref ref-type="bibr" rid="scirp.57608-ref26">26</xref>] . Iron as an essential trace metal plays numerous biochemical roles in the body, including oxygen blinding in haemoglobin and acting as an important catalytic centre in many enzymes [<xref ref-type="bibr" rid="scirp.57608-ref27">27</xref>] . The high level of Fe could be the reason behind the common use of these leaves in tackling iron deficiency associated disease (Anemia).</p></sec><sec id="s4"><title>4. Phytochemcial Analysis</title><p>Phytochemicals are natural bioactive compounds found in plants that work with nutrients and dietary fiber for disease protection [<xref ref-type="bibr" rid="scirp.57608-ref28">28</xref>] . The phytochemical results showed the absence of flavonoids, phenols and steroids in all the solvents used in the extraction as in <xref ref-type="table" rid="table3">Table 3</xref>. Alkaloids and saponins were present in moderate amount in ethanol extract with percentage composition of 4.56 and 1.47 respectively as in <xref ref-type="table" rid="table4">Table 4</xref>. Saponin was also equally</p><table-wrap id="table1" ><label><xref ref-type="table" rid="table1">Table 1</xref></label><caption><title> Proximate Composition of Gnetum africanum leaves</title></caption><table><tbody><thead><tr><th align="center" valign="middle" >Parameters</th><th align="center" valign="middle" >Composition (%)</th></tr></thead><tr><td align="center" valign="middle" >Moisture content</td><td align="center" valign="middle" >10.90</td></tr><tr><td align="center" valign="middle" >Crude protein</td><td align="center" valign="middle" >20.12</td></tr><tr><td align="center" valign="middle" >Ash content</td><td align="center" valign="middle" >6.70</td></tr><tr><td align="center" valign="middle" >Crude fibre</td><td align="center" valign="middle" >7.10</td></tr><tr><td align="center" valign="middle" >Lipids</td><td align="center" valign="middle" >2.79</td></tr><tr><td align="center" valign="middle" >Carbohydrate</td><td align="center" valign="middle" >52.39</td></tr><tr><td align="center" valign="middle" >Vit A.</td><td align="center" valign="middle" >0.129 mg/g</td></tr><tr><td align="center" valign="middle" >Vit C.</td><td align="center" valign="middle" >0.360 mg/l</td></tr></tbody></table></table-wrap><table-wrap id="table2" ><label><xref ref-type="table" rid="table2">Table 2</xref></label><caption><title> Elemental composition of G. africanum leaves</title></caption><table><tbody><thead><tr><th align="center" valign="middle" >Parameters</th><th align="center" valign="middle" >Compositon mg/l</th></tr></thead><tr><td align="center" valign="middle" >Ca</td><td align="center" valign="middle" >11.20</td></tr><tr><td align="center" valign="middle" >Cu</td><td align="center" valign="middle" >0.30</td></tr><tr><td align="center" valign="middle" >Zn</td><td align="center" valign="middle" >2.40</td></tr><tr><td align="center" valign="middle" >Cr</td><td align="center" valign="middle" >-</td></tr><tr><td align="center" valign="middle" >Fe</td><td align="center" valign="middle" >7.23</td></tr><tr><td align="center" valign="middle" >Pb</td><td align="center" valign="middle" >-</td></tr><tr><td align="center" valign="middle" >Mg</td><td align="center" valign="middle" >12.00</td></tr><tr><td align="center" valign="middle" >K</td><td align="center" valign="middle" >0.43</td></tr><tr><td align="center" valign="middle" >Na</td><td align="center" valign="middle" >23.00</td></tr><tr><td align="center" valign="middle" >Mn</td><td align="center" valign="middle" >1.56</td></tr></tbody></table></table-wrap><table-wrap id="table3" ><label><xref ref-type="table" rid="table3">Table 3</xref></label><caption><title> Qualitative composition of the phytochemicals of G. africanum leaves</title></caption><table><tbody><thead><tr><th align="center" valign="middle" >Parameters/Solvent</th><th align="center" valign="middle" >Water</th><th align="center" valign="middle" >Diethylether</th><th align="center" valign="middle" >Ethanol</th><th align="center" valign="middle" >n-hexane</th><th align="center" valign="middle" >Methanol</th></tr></thead><tr><td align="center" valign="middle" >Alkaloids</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >++</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td></tr><tr><td align="center" valign="middle" >Saponins</td><td align="center" valign="middle" >+</td><td align="center" valign="middle" >+</td><td align="center" valign="middle" >++</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >++</td></tr><tr><td align="center" valign="middle" >Flavonoids</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td></tr><tr><td align="center" valign="middle" >Phenols</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td></tr><tr><td align="center" valign="middle" >Steroids</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td></tr><tr><td align="center" valign="middle" >Glycosides</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >+</td><td align="center" valign="middle" >-</td></tr><tr><td align="center" valign="middle" >Tannins</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >-</td><td align="center" valign="middle" >++</td><td align="center" valign="middle" >++</td></tr></tbody></table></table-wrap><p>Key : +: Slightly present; +++: Highly present; ++: Moderately present; -: Absent</p><table-wrap id="table4" ><label><xref ref-type="table" rid="table4">Table 4</xref></label><caption><title> Quantitative composition of the phytochemicals of G. africanum leaves</title></caption><table><tbody><thead><tr><th align="center" valign="middle" >Parameter</th><th align="center" valign="middle" >Composition (%)</th></tr></thead><tr><td align="center" valign="middle" >Alkaloids</td><td align="center" valign="middle" >4.56</td></tr><tr><td align="center" valign="middle" >Glycosides</td><td align="center" valign="middle" >0.25</td></tr><tr><td align="center" valign="middle" >Saponins</td><td align="center" valign="middle" >1.47</td></tr><tr><td align="center" valign="middle" >Tannins</td><td align="center" valign="middle" >4.23</td></tr></tbody></table></table-wrap><p>present in water and diethyl ether in slight proportions. The valiue (1.47%) was higher than that (0.25%) previously obtained by Dike (2010) [<xref ref-type="bibr" rid="scirp.57608-ref4">4</xref>] . The presence supports its anti-inflammatory property. Glycosides was spotted in very low concentration only in n-hexane with quantitative compositon of 0.25% showing its medicinal property in prevention of heart diseases as in <xref ref-type="table" rid="table3">Table 3</xref> and <xref ref-type="table" rid="table4">Table 4</xref>. Glycosides and alkaloids have been reported to exert inhibiting activity against most bacteria [<xref ref-type="bibr" rid="scirp.57608-ref29">29</xref>] [<xref ref-type="bibr" rid="scirp.57608-ref30">30</xref>] .</p><p>Tannins an agent for toning of vital organs such as kidney and liver were only substantially present in n-hexane and methanol as in <xref ref-type="table" rid="table3">Table 3</xref> and <xref ref-type="table" rid="table4">Table 4</xref>.</p><p>Each of these phytochemicals is known for various protective and therapeutic effects [<xref ref-type="bibr" rid="scirp.57608-ref31">31</xref>] .</p></sec><sec id="s5"><title>5. Conclusion</title><p>The present study has shown the safety of this plant part based on the absence of the heavy metals with adverse health effects and its nutritive composition given the presence of requisite elements like Ca, Fe, Mg, Na, food classes like proteins, carborhydrates, crude fibre etc. Its phytochemical composition determine the medicinal values of these edible vegetable leaves and also could serve as a stating materials for the synthesis of new drugs in pharmaceutical industries.</p></sec><sec id="s6"><title>6. Recommendations</title><p>Further studies have to be carried out to isolate, characterise and elucidate the structures of the bioactive compounds from the plant for industrial drug formulation.</p></sec><sec id="s7"><title>NOTES</title></sec></body><back><ref-list><title>References</title><ref id="scirp.57608-ref1"><label>1</label><mixed-citation publication-type="other" xlink:type="simple">Eyo, E. and Abel, U. 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