# Qualitative and Quantitative Determination of Phytochemical Contents of Indigenous Nigerian Softwoods

Authors: Chukwuma S. Ezeonu; Chigozie M. Ejikeme
Journal: New Journal of Science
 (2016)

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## Abstract

The phytochemical contents of some milled Nigerian softwood chips were carried out in a quest to evaluate their potentials as sources of alternative medicine as well as uses in other industrial applications. The qualitative and quantitative analysis were ascertained. Tannin was found in all the Nigerian softwoods examined with the highest quantities obtained inSterculia oblonga(1240 mg/100 g) andBarteria nigritiana(1230 mg/100 g). Highest quantities of alkaloid were obtained inCordia millenii(11.2%) andSterculia oblonga(10.4%).Barteria nigritiana(14.2%) andMoringa oleifera(12.2%) recorded more flavonoid content than other individual softwoods. Saponin was more inAnogeissus leiocarpus(12.5%) andDichrostachys cinerea(9.8%). Oxalate was found to be higher inCombretodendron macrocarpum(5.84 g/100 g) andGlyphaea brevis(3.55 g/100 g).Pentaclethra macrophylla(890 mg/100 g) andMoringa oleifera(880 mg/100 g) contained more cyanogenic glycosides.Sacoglottis gabonensis(4.68 mg/g) andPentaclethra macrophylla(4.04 mg/g) showed the highest contents of phenol, while more lipids (8% and 7.2%) were found inAnogeissus leiocarpusandKaempferia galanga, respectively. The results showed that these Nigerian softwoods grains could be a source for the exploitation of these phytochemicals beneficial in the pharmaceutical and alternative medicine industries.


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## Body

## 1. Introduction

Nigeria abounds in diverse forest and natural resources. Forest trees are found in various parts of the country where they are lumbered and sold locally for house building and various construction purposes. Phytochemical studies of softwood stemvis-à-visNigerian timber components have been inadequate as the focuses of most research are on leaves and fruits of these plants. Some of these softwood stems contain phytochemicals locally extracted using alcohol or in aqueous form by traditional drug peddlers, showing that they have undocumented medicinal values. The medicinal value of plant lies in the phytochemical (bioactive) constituents of the plant which shows various physiological effects on human body. Therefore, through phytochemical screening one could detect the various important compounds which may be used as the bases of modern drugs for curing various diseases [1]. Chemical compounds produced as a result of metabolic reaction during plant growth are known as phytochemicals. Harborne [2] and Okwu [3] refer to such metabolic chemicals as “secondary metabolites” which include alkaloids, flavonoids, coumarins, tannins, terpenes, terpenoids, phenols, gums, polysaccharides, and glycosides. Human bodies under stress condition produce less enzymatic antioxidants (e.g., superoxide dismutase (SOD), glutathione peroxidase (GPx), and catalase) and nonenzymatic antioxidants (e.g., ascorbic acid (vitamin C), tocopherol (vitamin E)) but more reactive oxygen species (ROS) (e.g., superoxide anion radicals, hydroxyl radicals, and hydrogen peroxide) [4]. An imbalance could result from the above causing damage to the body cell [5–8] and other health challenges [9]. Preventive medicine has been immensely enhanced by the use of these natural plant antioxidants. Plants contain lots of free radical scavenging molecules some of which include alkaloids, amines, betalains, vitamins, terpenoids, phenolic acids, lignins, stilbenes, and tannins as well as other secondary metabolites with high level of antioxidant activity [4]. Most phytochemicals are antioxidant agents which essentially reduce the damages caused in tissue during physiological processes. This research is aimed at determining the qualitative and quantitative phytochemical components in Nigerian softwood as indices of producing plant secondary metabolites which have medicinal values as well as application in industries.


## 2. Materials and Methods

### 2.1. Materials

Soft wood samples were locally collected at saw mill in different states in Nigeria and identified as reported elsewhere [10, 11].


### 2.2. Wood Sample Preparation

Twenty-four identified softwoods were prepared before analysis in a finely grounded chip form as reported by Ejikeme et al. [11].


### 2.3. Phytochemical Constituents of Nigerian Softwood Samples

Methodologies for the determination of phytochemical used in this research were adapted from those reported by Keay et al. [10] and Ejikeme et al. [11], respectively.


### 2.4. Qualitative Analysis of the Phytochemicals of the Wood Sample

#### 2.4.1. Test for Tannins

Analysis used was the method reported by Ejikeme et al. [11]. Each wood powder sample (0.30 g) was weighed into a test tube and boiled for 10 minutes in a water bath containing 30 cm3 of water. Filtration was carried out after boiling using number 42 (125 mm) Whatman filter paper. To 5 cm3 of the filtrate was added 3 drops of 0.1% ferric chloride. A brownish green or a blue black colouration showed positive test.


#### 2.4.2. Test for Phlobatannins

Analytical method is according to Ejikeme et al. [11]. To each sample (0.30 g) weighed into a beaker was added 30 cm3 of distilled water. After 24 hours of extraction, aqueous extract (10 cm3) of each wood sample was boiled with 5 cm3 of 1% aqueous hydrochloric acid. Deposit of red precipitate showed positive test.


#### 2.4.3. Test for Saponin

Methodology is as reported by Ejikeme et al. [11]. Distilled water (30 cm3) was added to wood powder samples (0.30 g) and boiled for 10 minutes in water bath and filtered using Whatman filter paper number 42 (125 mm). A mixture of distilled water (5 cm3) and filtrate (10 cm3) was agitated vigorously for a stable persistent froth. The formation of emulsion on addition of three drops of olive oil showed positive result.


#### 2.4.4. Test for Steroid

Analytical method used is according to Ejikeme et al. [11]. Each sample (0.30 g) weighed into a beaker was mixed with 20 cm3 of ethanol; the component was extracted for 2 hours. To the ethanolic extract of each sample (5 cm3) was added 2 cm3 acetic anhydride followed with 2 cm3 of concentrated tetraoxosulphate (VI) acid. A violet to blue or green colour change in sample(s) indicates the presence of steroids.


#### 2.4.5. Test for Terpenoids

Methodology is as reported by Ejikeme et al. [11]. Each wood powder sample (0.30 g) was weighed into a beaker and extracted with 30 cm3 and component extracted for 2 hours. A mixture of chloroform (2 cm3) and concentrated tetraoxosulphate (VI) acid (3 cm3) was added to 5 cm3 of each extract to form a layer. The presence of a reddish brown colouration at the interface shows positive results for the presence of terpenoids.


#### 2.4.6. Test for Flavonoids

The test for flavonoid adopted is as reported by Sofowara [12] and Harborne [2]. Each sample (0.30 g) weighed into a beaker was extracted with 30 cm3 of distilled water for 2 hours and filtered with Whatman filter paper number 42 (125 mm). To 10 cm3 of the aqueous filtrate of each wood extract was added 5 cm3 of 1.0 M dilute ammonia solution followed by the addition of 5 cm3 of concentrated tetraoxosulphate (VI) acid. Appearance of yellow colouration which disappeared on standing shows the presence of flavonoids.


#### 2.4.7. Test for Alkaloids

Test for flavonoid used is as reported by Hikino et al. [13]. Extraction of component from 2 grams of each wood powder sample was carried out using 5% tetraoxosulphate (VI) acid (H2SO4) (20 cm3) in 50% ethanol by boiling for 2 minutes and filtered through Whatman filter paper number 42 (125 mm). The filtrate was made alkaline using 5 cm3 of 28% ammonia solution (NH3) in a separating funnel. Equal volume of chloroform (5.0 cm3) was used in further solution extraction in which chloroform solution was extracted with two 5 cm3 portions of 1.0 M dilute tetraoxosulphate (VI) acid. This final acid extract was then used to carry out the following test: 0.5 cm3 of Dragendorff’s reagent (Bismuth potassium iodide solution) was mixed with 2 cm3 of acid extract and precipitated orange colour infers the presence of alkaloid.


#### 2.4.8. Test for Glycoside

Glycoside test was conducted according to the method reported by Hikino et al. [13]. To 2.00 g of each sample was added 20 cm3 of water, heated for 5 minutes on a water bath and filtered through Gem filter paper (12.5 cm). The following tests were carried out with the filtrate:(a)
0.2 cm3 of Fehling’s solutions A and B was mixed with 5 cm3 of the filtrate until it became alkaline (tested with litmus paper). A brick-red colouration on heating showed a positive result.(b)
Instead of water, 15 cm3 of 1.0 M sulphuric acid was used to repeat the above test and the quantity of precipitate obtained compared with that of (a) above. High precipitate content indicates the presence of glycoside while low content shows the absence of glycoside.


### 2.5. Quantitative Determination of Phytochemical Constituents of Woods

#### 2.5.1. Tannin

Analytical method for quantitative determination of tannin was according to Amadi et al. [14] and Ejikeme et al. [11]. By dissolving 50 g of sodium tungstate (Na2WO4) in 37 cm3 of distilled water, Folin-Denis reagent was made. To the reagent prepared above, 10 g of phosphomolybdic acid (H3PMo12O40) and 25 cm3 of orthophosphoric acid (H3PO4) were added. Two-hour reflux of the mixture was carried out, cooled, and diluted to 500 cm3 with distilled water. One gram of each wood powder (sample) in a conical flask was added to 100 cm3 of distilled water. This was boiled gently for 1 hour on an electric hot plate and filtered using number 42 (125 mm) Whatman filter paper in a 100 cm3 volumetric flask. Addition of 5.0 cm3 Folin-Denis reagent and 10 cm3 of saturated Na2CO3 solution into 50 cm3 of distilled water and 10 cm3 of diluted extract (aliquot volume) was carried out after being pipetted into a 100 cm3 conical flask for colour development. The solution was allowed to stand for 30 minutes in a water bath at a temperature of 25°C after thorough agitation. With the aid of a Spectrum Lab 23A spectrophotometer optical density was measured at 700 nm and compared on a standard tannic acid curve. Dissolution of 0.20 g of tannic acid in distilled water and dilution to 200 cm3 mark (1 mg/cm3) were used to obtain tannic standard curve. Varying concentrations (0.2–1.0 mg/cm3) of the standard tannic acid solution were pipetted into five different test tubes to which Folin-Denis reagent (5 cm3) and saturated Na2CO3 (10 cm3) solution were added and made up to the 100 cm3 mark with distilled water. The solution was left to stand for 30 minutes in a water bath at 25°C. Optical density was ascertained at 700 nm with the aid of a Spectrum Lab 23A spectrophotometer. Optical density (absorbance) versus tannic acid concentration was plotted.The following formula was used in the calculation:(1)Tannic acidmg100g=C×extract volume×100Aliquot volume×weight of sample,where C is concentration of tannic acid read off the graph.


#### 2.5.2. Determination of Alkaloids

Quantitative determination of alkaloid was according to the methodology by Harborne [2]. Exactly 200 cm3 of 10% acetic acid in ethanol was added to each wood powder sample (2.50 g) in a 250 cm3 beaker and allowed to stand for 4 hours. The extract was concentrated on a water bath to one-quarter of the original volume followed by addition of 15 drops of concentrated ammonium hydroxide dropwise to the extract until the precipitation was complete immediately after filtration. After 3 hours of mixture sedimentation, the supernatant was discarded and the precipitates were washed with 20 cm3 of 0.1 M of ammonium hydroxide and then filtered using Gem filter paper (12.5 cm). Using electronic weighing balance Model B-218, the residue was dried in an oven and the percentage of alkaloid is expressed mathematically as(2)%  Alkaloid=Weight of alkaloidWeight of sample×100.


#### 2.5.3. Determination of Flavonoid

Flavonoid determination was by the method reported by Ejikeme et al. [11] and Boham and Kocipai [15]. Exactly 50 cm3 of 80% aqueous methanol added was added to 2.50 g of sample in a 250 cm3 beaker, covered, and allowed to stand for 24 hours at room temperature. After discarding the supernatant, the residue was reextracted (three times) with the same volume of ethanol. Whatman filter paper number 42 (125 mm) was used to filter whole solution of each wood sample. Each wood sample filtrate was later transferred into a crucible and evaporated to dryness over a water bath. The content in the crucible was cooled in a desiccator and weighed until constant weight was obtained. The percentage of flavonoid was calculated as (3)%  Flavonoid=Weight of flavonoidWeight of sample×100.


#### 2.5.4. Determination of Saponin

Saponin quantitative determination was carried out using the method reported by Ejikeme et al. [11] and Obadoni and Ochuko [16]. Exactly 100 cm3 of 20% aqueous ethanol was added to 5 grams of each wood powder sample in a 250 cm3 conical flask. The mixture was heated over a hot water bath for 4 hours with continuous stirring at a temperature of 55°C. The residue of the mixture was reextracted with another 100 cm3 of 20% aqueous ethanol after filtration and heated for 4 hours at a constant temperature of 55°C with constant stirring. The combined extract was evaporated to 40 cm3 over water bath at 90°C. 20 cm3 of diethyl ether was added to the concentrate in a 250 cm3 separator funnel and vigorously agitated from which the aqueous layer was recovered while the ether layer was discarded. This purification process was repeated twice. 60 cm3 of n-butanol was added and extracted twice with 10 cm3 of 5% sodium chloride. After discarding the sodium chloride layer the remaining solution was heated in a water bath for 30 minutes, after which the solution was transferred into a crucible and was dried in an oven to a constant weight. The saponin content was calculated as a percentage:(4)%  Saponin=Weight of saponinWeight of sample×100.


#### 2.5.5. Determination of Oxalate

Oxalate quantitative determination was carried out using the method reported by Ejikeme et al. [11] and Munro and Bassir [17]. Exactly 20 cm3 of 0.3 M HCl in each wood powder sample (2.50 g) was extracted three (3) times by warming at a temperature of 50°C for 1 hour with constant stirring using a magnetic stirrer. For oxalate estimation, 1.0 cm3 of 5 M ammonium hydroxide was added to 5.0 cm3 of extract to ensure alkalinity. Addition of 2 drops of phenolphthalein indicator, 3 drops of glacial acetic acid, and 1.0 cm3 of 5% calcium chloride to make the mixture acidic before standing for 3 hours was followed by centrifugation at 3000 rpm for 15 minutes. After discarding the supernatant, the precipitate was washed three times using hot water by mixing thoroughly each time centrifugation. Then, to each tube, 2.0 cm3 of 3 M tetraoxosulphate (VI) acid was added and the precipitate dissolved by warming in a water bath at 70°C. Freshly prepared 0.01 M potassium permanganate (KMnO4) was titrated against the content of each tube at room temperature until the first pink colour appears throughout the solution. The solution was allowed to stand until it returned colourless, after which it was warmed on an electric hot plate at 70°C for 3 minutes, and retitrated again until a pink colour appears and persists for at least 30 seconds.Titration reaction of oxalate in sample was calculated as(5)C2O2-4+8H++MnO2-4=2CO2+4H2O+Mn2+Ratio of reacting ions=1 : 1FromM1V1=M2V2,where M1 is molarity of KMnO4, M2 is molarity of extract (oxalate), V1 is volume of extract (oxalate), and V2 is volume of KMnO4 (Titre Value).(6)Molecular Weight ofCaCO3=100Weight of oxalate in titre=M2×molecular weight=XgWeight of oxalate in titrand2Cm3=Xg1000×2=Y100Cm3of oxalate extract will contain=Y2.5×100g=W%  oxalate compositiong/100g=W2.5×1001.


#### 2.5.6. Determination of Cyanogenic Glycoside

Cyanogenic glycoside quantitative determination methodology used in this research is that by Amadi et al. [14] as reported by Ejikeme et al. [11]. It was weighed into a 250 cm3 round bottom flask and about 200 cm3 of distilled water was added to one gram of each dry wood powder sample and allowed to stand for 2 hours for autolysis to occur. Full distillation was carried out in a 250 cm3 conical flask containing 20 cm3 of 2.5% NaOH (sodium hydroxide) in the sample after adding an antifoaming agent (tannic acid). Cyanogenic glycoside (100 cm3), 8 cm3 of 6 M NH4OH (ammonium hydroxide), and 2 cm3 of 5% KI (potassium iodide) were added to the distillate(s), mixed, and titrated with 0.02 M AgNO3 (silver nitrate) using a microburette against a black background. Turbidity which was continuous indicates the end point.Content of cyanogenic glycoside in the sample was calculated as(7)Cyanogenic glycosidemg100g=Titre ValueCm3×1.08×exact volumeAliquot volumeCm3×sample weightg×100.


#### 2.5.7. Determination of Percentage Lipid

Into a thimble connected to a soxhlet extractor chamber with a preweighed flat bottom was added each dry wood powder (2.50 g) and connected to a condenser. Petroleum ether (100 mL) enough to cause a reflux was added to the flask and the lipid from the wood sample was extracted for 3 hours by heating on an electric hot plate at 50°C. The extractant (petroleum ether) was distilled off and the lipid recovered by cooling the flask in a dessicator and its value calculated by reweighed flask and content. This methodology for lipid quantitative analysis is reported elsewhere [18]. The calculated percentage of lipid content was thus (8)%  Lipid=Weight of LipidWeight of Sample×100.


#### 2.5.8. Determination of Phenols

Defatting of 2 g wood powder sample was carried out for 2 hours in 100 cm3 of ether using a soxhlet apparatus. The defatted sample (0.50 g) was boiled for 15 minutes with 50 cm3 of ether for the extraction of the phenolic components. Exactly 10 cm3 of distilled water, 2 cm3 of 0.1 N ammonium hydroxide solution, and 5 cm3 of concentrated amyl alcohol were also added to 5 cm3 of the extract and left to react for 30 minutes for colour development. The optical density was measured at 505 nm. 0.20 g of tannic acid was dissolving in distilled water and diluted to 200 mL mark (1 mg/cm3) in preparation for phenol standard curve. Varying concentrations (0.2–1.0 mg/cm3) of the standard tannic acid solution were pipetted into five different test tubes to which 2 cm3 of NH3OH, 5 cm3 of amyl alcohol, and 10 cm3 of water were added. The solution was made up to 100 cm3 volume and left to react for 30 minutes for colour development. The optical density was determined at 505 nm [10].


## 2.1. Materials

Soft wood samples were locally collected at saw mill in different states in Nigeria and identified as reported elsewhere [10, 11].


## 2.2. Wood Sample Preparation

Twenty-four identified softwoods were prepared before analysis in a finely grounded chip form as reported by Ejikeme et al. [11].


## 2.3. Phytochemical Constituents of Nigerian Softwood Samples

Methodologies for the determination of phytochemical used in this research were adapted from those reported by Keay et al. [10] and Ejikeme et al. [11], respectively.


## 2.4. Qualitative Analysis of the Phytochemicals of the Wood Sample

### 2.4.1. Test for Tannins

Analysis used was the method reported by Ejikeme et al. [11]. Each wood powder sample (0.30 g) was weighed into a test tube and boiled for 10 minutes in a water bath containing 30 cm3 of water. Filtration was carried out after boiling using number 42 (125 mm) Whatman filter paper. To 5 cm3 of the filtrate was added 3 drops of 0.1% ferric chloride. A brownish green or a blue black colouration showed positive test.


### 2.4.2. Test for Phlobatannins

Analytical method is according to Ejikeme et al. [11]. To each sample (0.30 g) weighed into a beaker was added 30 cm3 of distilled water. After 24 hours of extraction, aqueous extract (10 cm3) of each wood sample was boiled with 5 cm3 of 1% aqueous hydrochloric acid. Deposit of red precipitate showed positive test.


### 2.4.3. Test for Saponin

Methodology is as reported by Ejikeme et al. [11]. Distilled water (30 cm3) was added to wood powder samples (0.30 g) and boiled for 10 minutes in water bath and filtered using Whatman filter paper number 42 (125 mm). A mixture of distilled water (5 cm3) and filtrate (10 cm3) was agitated vigorously for a stable persistent froth. The formation of emulsion on addition of three drops of olive oil showed positive result.


### 2.4.4. Test for Steroid

Analytical method used is according to Ejikeme et al. [11]. Each sample (0.30 g) weighed into a beaker was mixed with 20 cm3 of ethanol; the component was extracted for 2 hours. To the ethanolic extract of each sample (5 cm3) was added 2 cm3 acetic anhydride followed with 2 cm3 of concentrated tetraoxosulphate (VI) acid. A violet to blue or green colour change in sample(s) indicates the presence of steroids.


### 2.4.5. Test for Terpenoids

Methodology is as reported by Ejikeme et al. [11]. Each wood powder sample (0.30 g) was weighed into a beaker and extracted with 30 cm3 and component extracted for 2 hours. A mixture of chloroform (2 cm3) and concentrated tetraoxosulphate (VI) acid (3 cm3) was added to 5 cm3 of each extract to form a layer. The presence of a reddish brown colouration at the interface shows positive results for the presence of terpenoids.


### 2.4.6. Test for Flavonoids

The test for flavonoid adopted is as reported by Sofowara [12] and Harborne [2]. Each sample (0.30 g) weighed into a beaker was extracted with 30 cm3 of distilled water for 2 hours and filtered with Whatman filter paper number 42 (125 mm). To 10 cm3 of the aqueous filtrate of each wood extract was added 5 cm3 of 1.0 M dilute ammonia solution followed by the addition of 5 cm3 of concentrated tetraoxosulphate (VI) acid. Appearance of yellow colouration which disappeared on standing shows the presence of flavonoids.


### 2.4.7. Test for Alkaloids

Test for flavonoid used is as reported by Hikino et al. [13]. Extraction of component from 2 grams of each wood powder sample was carried out using 5% tetraoxosulphate (VI) acid (H2SO4) (20 cm3) in 50% ethanol by boiling for 2 minutes and filtered through Whatman filter paper number 42 (125 mm). The filtrate was made alkaline using 5 cm3 of 28% ammonia solution (NH3) in a separating funnel. Equal volume of chloroform (5.0 cm3) was used in further solution extraction in which chloroform solution was extracted with two 5 cm3 portions of 1.0 M dilute tetraoxosulphate (VI) acid. This final acid extract was then used to carry out the following test: 0.5 cm3 of Dragendorff’s reagent (Bismuth potassium iodide solution) was mixed with 2 cm3 of acid extract and precipitated orange colour infers the presence of alkaloid.


### 2.4.8. Test for Glycoside

Glycoside test was conducted according to the method reported by Hikino et al. [13]. To 2.00 g of each sample was added 20 cm3 of water, heated for 5 minutes on a water bath and filtered through Gem filter paper (12.5 cm). The following tests were carried out with the filtrate:(a)
0.2 cm3 of Fehling’s solutions A and B was mixed with 5 cm3 of the filtrate until it became alkaline (tested with litmus paper). A brick-red colouration on heating showed a positive result.(b)
Instead of water, 15 cm3 of 1.0 M sulphuric acid was used to repeat the above test and the quantity of precipitate obtained compared with that of (a) above. High precipitate content indicates the presence of glycoside while low content shows the absence of glycoside.


## 2.4.1. Test for Tannins

Analysis used was the method reported by Ejikeme et al. [11]. Each wood powder sample (0.30 g) was weighed into a test tube and boiled for 10 minutes in a water bath containing 30 cm3 of water. Filtration was carried out after boiling using number 42 (125 mm) Whatman filter paper. To 5 cm3 of the filtrate was added 3 drops of 0.1% ferric chloride. A brownish green or a blue black colouration showed positive test.


## 2.4.2. Test for Phlobatannins

Analytical method is according to Ejikeme et al. [11]. To each sample (0.30 g) weighed into a beaker was added 30 cm3 of distilled water. After 24 hours of extraction, aqueous extract (10 cm3) of each wood sample was boiled with 5 cm3 of 1% aqueous hydrochloric acid. Deposit of red precipitate showed positive test.


## 2.4.3. Test for Saponin

Methodology is as reported by Ejikeme et al. [11]. Distilled water (30 cm3) was added to wood powder samples (0.30 g) and boiled for 10 minutes in water bath and filtered using Whatman filter paper number 42 (125 mm). A mixture of distilled water (5 cm3) and filtrate (10 cm3) was agitated vigorously for a stable persistent froth. The formation of emulsion on addition of three drops of olive oil showed positive result.


## 2.4.4. Test for Steroid

Analytical method used is according to Ejikeme et al. [11]. Each sample (0.30 g) weighed into a beaker was mixed with 20 cm3 of ethanol; the component was extracted for 2 hours. To the ethanolic extract of each sample (5 cm3) was added 2 cm3 acetic anhydride followed with 2 cm3 of concentrated tetraoxosulphate (VI) acid. A violet to blue or green colour change in sample(s) indicates the presence of steroids.


## 2.4.5. Test for Terpenoids

Methodology is as reported by Ejikeme et al. [11]. Each wood powder sample (0.30 g) was weighed into a beaker and extracted with 30 cm3 and component extracted for 2 hours. A mixture of chloroform (2 cm3) and concentrated tetraoxosulphate (VI) acid (3 cm3) was added to 5 cm3 of each extract to form a layer. The presence of a reddish brown colouration at the interface shows positive results for the presence of terpenoids.


## 2.4.6. Test for Flavonoids

The test for flavonoid adopted is as reported by Sofowara [12] and Harborne [2]. Each sample (0.30 g) weighed into a beaker was extracted with 30 cm3 of distilled water for 2 hours and filtered with Whatman filter paper number 42 (125 mm). To 10 cm3 of the aqueous filtrate of each wood extract was added 5 cm3 of 1.0 M dilute ammonia solution followed by the addition of 5 cm3 of concentrated tetraoxosulphate (VI) acid. Appearance of yellow colouration which disappeared on standing shows the presence of flavonoids.


## 2.4.7. Test for Alkaloids

Test for flavonoid used is as reported by Hikino et al. [13]. Extraction of component from 2 grams of each wood powder sample was carried out using 5% tetraoxosulphate (VI) acid (H2SO4) (20 cm3) in 50% ethanol by boiling for 2 minutes and filtered through Whatman filter paper number 42 (125 mm). The filtrate was made alkaline using 5 cm3 of 28% ammonia solution (NH3) in a separating funnel. Equal volume of chloroform (5.0 cm3) was used in further solution extraction in which chloroform solution was extracted with two 5 cm3 portions of 1.0 M dilute tetraoxosulphate (VI) acid. This final acid extract was then used to carry out the following test: 0.5 cm3 of Dragendorff’s reagent (Bismuth potassium iodide solution) was mixed with 2 cm3 of acid extract and precipitated orange colour infers the presence of alkaloid.


## 2.4.8. Test for Glycoside

Glycoside test was conducted according to the method reported by Hikino et al. [13]. To 2.00 g of each sample was added 20 cm3 of water, heated for 5 minutes on a water bath and filtered through Gem filter paper (12.5 cm). The following tests were carried out with the filtrate:(a)
0.2 cm3 of Fehling’s solutions A and B was mixed with 5 cm3 of the filtrate until it became alkaline (tested with litmus paper). A brick-red colouration on heating showed a positive result.(b)
Instead of water, 15 cm3 of 1.0 M sulphuric acid was used to repeat the above test and the quantity of precipitate obtained compared with that of (a) above. High precipitate content indicates the presence of glycoside while low content shows the absence of glycoside.


## 2.5. Quantitative Determination of Phytochemical Constituents of Woods

### 2.5.1. Tannin

Analytical method for quantitative determination of tannin was according to Amadi et al. [14] and Ejikeme et al. [11]. By dissolving 50 g of sodium tungstate (Na2WO4) in 37 cm3 of distilled water, Folin-Denis reagent was made. To the reagent prepared above, 10 g of phosphomolybdic acid (H3PMo12O40) and 25 cm3 of orthophosphoric acid (H3PO4) were added. Two-hour reflux of the mixture was carried out, cooled, and diluted to 500 cm3 with distilled water. One gram of each wood powder (sample) in a conical flask was added to 100 cm3 of distilled water. This was boiled gently for 1 hour on an electric hot plate and filtered using number 42 (125 mm) Whatman filter paper in a 100 cm3 volumetric flask. Addition of 5.0 cm3 Folin-Denis reagent and 10 cm3 of saturated Na2CO3 solution into 50 cm3 of distilled water and 10 cm3 of diluted extract (aliquot volume) was carried out after being pipetted into a 100 cm3 conical flask for colour development. The solution was allowed to stand for 30 minutes in a water bath at a temperature of 25°C after thorough agitation. With the aid of a Spectrum Lab 23A spectrophotometer optical density was measured at 700 nm and compared on a standard tannic acid curve. Dissolution of 0.20 g of tannic acid in distilled water and dilution to 200 cm3 mark (1 mg/cm3) were used to obtain tannic standard curve. Varying concentrations (0.2–1.0 mg/cm3) of the standard tannic acid solution were pipetted into five different test tubes to which Folin-Denis reagent (5 cm3) and saturated Na2CO3 (10 cm3) solution were added and made up to the 100 cm3 mark with distilled water. The solution was left to stand for 30 minutes in a water bath at 25°C. Optical density was ascertained at 700 nm with the aid of a Spectrum Lab 23A spectrophotometer. Optical density (absorbance) versus tannic acid concentration was plotted.The following formula was used in the calculation:(1)Tannic acidmg100g=C×extract volume×100Aliquot volume×weight of sample,where C is concentration of tannic acid read off the graph.


### 2.5.2. Determination of Alkaloids

Quantitative determination of alkaloid was according to the methodology by Harborne [2]. Exactly 200 cm3 of 10% acetic acid in ethanol was added to each wood powder sample (2.50 g) in a 250 cm3 beaker and allowed to stand for 4 hours. The extract was concentrated on a water bath to one-quarter of the original volume followed by addition of 15 drops of concentrated ammonium hydroxide dropwise to the extract until the precipitation was complete immediately after filtration. After 3 hours of mixture sedimentation, the supernatant was discarded and the precipitates were washed with 20 cm3 of 0.1 M of ammonium hydroxide and then filtered using Gem filter paper (12.5 cm). Using electronic weighing balance Model B-218, the residue was dried in an oven and the percentage of alkaloid is expressed mathematically as(2)%  Alkaloid=Weight of alkaloidWeight of sample×100.


### 2.5.3. Determination of Flavonoid

Flavonoid determination was by the method reported by Ejikeme et al. [11] and Boham and Kocipai [15]. Exactly 50 cm3 of 80% aqueous methanol added was added to 2.50 g of sample in a 250 cm3 beaker, covered, and allowed to stand for 24 hours at room temperature. After discarding the supernatant, the residue was reextracted (three times) with the same volume of ethanol. Whatman filter paper number 42 (125 mm) was used to filter whole solution of each wood sample. Each wood sample filtrate was later transferred into a crucible and evaporated to dryness over a water bath. The content in the crucible was cooled in a desiccator and weighed until constant weight was obtained. The percentage of flavonoid was calculated as (3)%  Flavonoid=Weight of flavonoidWeight of sample×100.


### 2.5.4. Determination of Saponin

Saponin quantitative determination was carried out using the method reported by Ejikeme et al. [11] and Obadoni and Ochuko [16]. Exactly 100 cm3 of 20% aqueous ethanol was added to 5 grams of each wood powder sample in a 250 cm3 conical flask. The mixture was heated over a hot water bath for 4 hours with continuous stirring at a temperature of 55°C. The residue of the mixture was reextracted with another 100 cm3 of 20% aqueous ethanol after filtration and heated for 4 hours at a constant temperature of 55°C with constant stirring. The combined extract was evaporated to 40 cm3 over water bath at 90°C. 20 cm3 of diethyl ether was added to the concentrate in a 250 cm3 separator funnel and vigorously agitated from which the aqueous layer was recovered while the ether layer was discarded. This purification process was repeated twice. 60 cm3 of n-butanol was added and extracted twice with 10 cm3 of 5% sodium chloride. After discarding the sodium chloride layer the remaining solution was heated in a water bath for 30 minutes, after which the solution was transferred into a crucible and was dried in an oven to a constant weight. The saponin content was calculated as a percentage:(4)%  Saponin=Weight of saponinWeight of sample×100.


### 2.5.5. Determination of Oxalate

Oxalate quantitative determination was carried out using the method reported by Ejikeme et al. [11] and Munro and Bassir [17]. Exactly 20 cm3 of 0.3 M HCl in each wood powder sample (2.50 g) was extracted three (3) times by warming at a temperature of 50°C for 1 hour with constant stirring using a magnetic stirrer. For oxalate estimation, 1.0 cm3 of 5 M ammonium hydroxide was added to 5.0 cm3 of extract to ensure alkalinity. Addition of 2 drops of phenolphthalein indicator, 3 drops of glacial acetic acid, and 1.0 cm3 of 5% calcium chloride to make the mixture acidic before standing for 3 hours was followed by centrifugation at 3000 rpm for 15 minutes. After discarding the supernatant, the precipitate was washed three times using hot water by mixing thoroughly each time centrifugation. Then, to each tube, 2.0 cm3 of 3 M tetraoxosulphate (VI) acid was added and the precipitate dissolved by warming in a water bath at 70°C. Freshly prepared 0.01 M potassium permanganate (KMnO4) was titrated against the content of each tube at room temperature until the first pink colour appears throughout the solution. The solution was allowed to stand until it returned colourless, after which it was warmed on an electric hot plate at 70°C for 3 minutes, and retitrated again until a pink colour appears and persists for at least 30 seconds.Titration reaction of oxalate in sample was calculated as(5)C2O2-4+8H++MnO2-4=2CO2+4H2O+Mn2+Ratio of reacting ions=1 : 1FromM1V1=M2V2,where M1 is molarity of KMnO4, M2 is molarity of extract (oxalate), V1 is volume of extract (oxalate), and V2 is volume of KMnO4 (Titre Value).(6)Molecular Weight ofCaCO3=100Weight of oxalate in titre=M2×molecular weight=XgWeight of oxalate in titrand2Cm3=Xg1000×2=Y100Cm3of oxalate extract will contain=Y2.5×100g=W%  oxalate compositiong/100g=W2.5×1001.


### 2.5.6. Determination of Cyanogenic Glycoside

Cyanogenic glycoside quantitative determination methodology used in this research is that by Amadi et al. [14] as reported by Ejikeme et al. [11]. It was weighed into a 250 cm3 round bottom flask and about 200 cm3 of distilled water was added to one gram of each dry wood powder sample and allowed to stand for 2 hours for autolysis to occur. Full distillation was carried out in a 250 cm3 conical flask containing 20 cm3 of 2.5% NaOH (sodium hydroxide) in the sample after adding an antifoaming agent (tannic acid). Cyanogenic glycoside (100 cm3), 8 cm3 of 6 M NH4OH (ammonium hydroxide), and 2 cm3 of 5% KI (potassium iodide) were added to the distillate(s), mixed, and titrated with 0.02 M AgNO3 (silver nitrate) using a microburette against a black background. Turbidity which was continuous indicates the end point.Content of cyanogenic glycoside in the sample was calculated as(7)Cyanogenic glycosidemg100g=Titre ValueCm3×1.08×exact volumeAliquot volumeCm3×sample weightg×100.


### 2.5.7. Determination of Percentage Lipid

Into a thimble connected to a soxhlet extractor chamber with a preweighed flat bottom was added each dry wood powder (2.50 g) and connected to a condenser. Petroleum ether (100 mL) enough to cause a reflux was added to the flask and the lipid from the wood sample was extracted for 3 hours by heating on an electric hot plate at 50°C. The extractant (petroleum ether) was distilled off and the lipid recovered by cooling the flask in a dessicator and its value calculated by reweighed flask and content. This methodology for lipid quantitative analysis is reported elsewhere [18]. The calculated percentage of lipid content was thus (8)%  Lipid=Weight of LipidWeight of Sample×100.


### 2.5.8. Determination of Phenols

Defatting of 2 g wood powder sample was carried out for 2 hours in 100 cm3 of ether using a soxhlet apparatus. The defatted sample (0.50 g) was boiled for 15 minutes with 50 cm3 of ether for the extraction of the phenolic components. Exactly 10 cm3 of distilled water, 2 cm3 of 0.1 N ammonium hydroxide solution, and 5 cm3 of concentrated amyl alcohol were also added to 5 cm3 of the extract and left to react for 30 minutes for colour development. The optical density was measured at 505 nm. 0.20 g of tannic acid was dissolving in distilled water and diluted to 200 mL mark (1 mg/cm3) in preparation for phenol standard curve. Varying concentrations (0.2–1.0 mg/cm3) of the standard tannic acid solution were pipetted into five different test tubes to which 2 cm3 of NH3OH, 5 cm3 of amyl alcohol, and 10 cm3 of water were added. The solution was made up to 100 cm3 volume and left to react for 30 minutes for colour development. The optical density was determined at 505 nm [10].


## 2.5.1. Tannin

Analytical method for quantitative determination of tannin was according to Amadi et al. [14] and Ejikeme et al. [11]. By dissolving 50 g of sodium tungstate (Na2WO4) in 37 cm3 of distilled water, Folin-Denis reagent was made. To the reagent prepared above, 10 g of phosphomolybdic acid (H3PMo12O40) and 25 cm3 of orthophosphoric acid (H3PO4) were added. Two-hour reflux of the mixture was carried out, cooled, and diluted to 500 cm3 with distilled water. One gram of each wood powder (sample) in a conical flask was added to 100 cm3 of distilled water. This was boiled gently for 1 hour on an electric hot plate and filtered using number 42 (125 mm) Whatman filter paper in a 100 cm3 volumetric flask. Addition of 5.0 cm3 Folin-Denis reagent and 10 cm3 of saturated Na2CO3 solution into 50 cm3 of distilled water and 10 cm3 of diluted extract (aliquot volume) was carried out after being pipetted into a 100 cm3 conical flask for colour development. The solution was allowed to stand for 30 minutes in a water bath at a temperature of 25°C after thorough agitation. With the aid of a Spectrum Lab 23A spectrophotometer optical density was measured at 700 nm and compared on a standard tannic acid curve. Dissolution of 0.20 g of tannic acid in distilled water and dilution to 200 cm3 mark (1 mg/cm3) were used to obtain tannic standard curve. Varying concentrations (0.2–1.0 mg/cm3) of the standard tannic acid solution were pipetted into five different test tubes to which Folin-Denis reagent (5 cm3) and saturated Na2CO3 (10 cm3) solution were added and made up to the 100 cm3 mark with distilled water. The solution was left to stand for 30 minutes in a water bath at 25°C. Optical density was ascertained at 700 nm with the aid of a Spectrum Lab 23A spectrophotometer. Optical density (absorbance) versus tannic acid concentration was plotted.The following formula was used in the calculation:(1)Tannic acidmg100g=C×extract volume×100Aliquot volume×weight of sample,where C is concentration of tannic acid read off the graph.


## 2.5.2. Determination of Alkaloids

Quantitative determination of alkaloid was according to the methodology by Harborne [2]. Exactly 200 cm3 of 10% acetic acid in ethanol was added to each wood powder sample (2.50 g) in a 250 cm3 beaker and allowed to stand for 4 hours. The extract was concentrated on a water bath to one-quarter of the original volume followed by addition of 15 drops of concentrated ammonium hydroxide dropwise to the extract until the precipitation was complete immediately after filtration. After 3 hours of mixture sedimentation, the supernatant was discarded and the precipitates were washed with 20 cm3 of 0.1 M of ammonium hydroxide and then filtered using Gem filter paper (12.5 cm). Using electronic weighing balance Model B-218, the residue was dried in an oven and the percentage of alkaloid is expressed mathematically as(2)%  Alkaloid=Weight of alkaloidWeight of sample×100.


## 2.5.3. Determination of Flavonoid

Flavonoid determination was by the method reported by Ejikeme et al. [11] and Boham and Kocipai [15]. Exactly 50 cm3 of 80% aqueous methanol added was added to 2.50 g of sample in a 250 cm3 beaker, covered, and allowed to stand for 24 hours at room temperature. After discarding the supernatant, the residue was reextracted (three times) with the same volume of ethanol. Whatman filter paper number 42 (125 mm) was used to filter whole solution of each wood sample. Each wood sample filtrate was later transferred into a crucible and evaporated to dryness over a water bath. The content in the crucible was cooled in a desiccator and weighed until constant weight was obtained. The percentage of flavonoid was calculated as (3)%  Flavonoid=Weight of flavonoidWeight of sample×100.


## 2.5.4. Determination of Saponin

Saponin quantitative determination was carried out using the method reported by Ejikeme et al. [11] and Obadoni and Ochuko [16]. Exactly 100 cm3 of 20% aqueous ethanol was added to 5 grams of each wood powder sample in a 250 cm3 conical flask. The mixture was heated over a hot water bath for 4 hours with continuous stirring at a temperature of 55°C. The residue of the mixture was reextracted with another 100 cm3 of 20% aqueous ethanol after filtration and heated for 4 hours at a constant temperature of 55°C with constant stirring. The combined extract was evaporated to 40 cm3 over water bath at 90°C. 20 cm3 of diethyl ether was added to the concentrate in a 250 cm3 separator funnel and vigorously agitated from which the aqueous layer was recovered while the ether layer was discarded. This purification process was repeated twice. 60 cm3 of n-butanol was added and extracted twice with 10 cm3 of 5% sodium chloride. After discarding the sodium chloride layer the remaining solution was heated in a water bath for 30 minutes, after which the solution was transferred into a crucible and was dried in an oven to a constant weight. The saponin content was calculated as a percentage:(4)%  Saponin=Weight of saponinWeight of sample×100.


## 2.5.5. Determination of Oxalate

Oxalate quantitative determination was carried out using the method reported by Ejikeme et al. [11] and Munro and Bassir [17]. Exactly 20 cm3 of 0.3 M HCl in each wood powder sample (2.50 g) was extracted three (3) times by warming at a temperature of 50°C for 1 hour with constant stirring using a magnetic stirrer. For oxalate estimation, 1.0 cm3 of 5 M ammonium hydroxide was added to 5.0 cm3 of extract to ensure alkalinity. Addition of 2 drops of phenolphthalein indicator, 3 drops of glacial acetic acid, and 1.0 cm3 of 5% calcium chloride to make the mixture acidic before standing for 3 hours was followed by centrifugation at 3000 rpm for 15 minutes. After discarding the supernatant, the precipitate was washed three times using hot water by mixing thoroughly each time centrifugation. Then, to each tube, 2.0 cm3 of 3 M tetraoxosulphate (VI) acid was added and the precipitate dissolved by warming in a water bath at 70°C. Freshly prepared 0.01 M potassium permanganate (KMnO4) was titrated against the content of each tube at room temperature until the first pink colour appears throughout the solution. The solution was allowed to stand until it returned colourless, after which it was warmed on an electric hot plate at 70°C for 3 minutes, and retitrated again until a pink colour appears and persists for at least 30 seconds.Titration reaction of oxalate in sample was calculated as(5)C2O2-4+8H++MnO2-4=2CO2+4H2O+Mn2+Ratio of reacting ions=1 : 1FromM1V1=M2V2,where M1 is molarity of KMnO4, M2 is molarity of extract (oxalate), V1 is volume of extract (oxalate), and V2 is volume of KMnO4 (Titre Value).(6)Molecular Weight ofCaCO3=100Weight of oxalate in titre=M2×molecular weight=XgWeight of oxalate in titrand2Cm3=Xg1000×2=Y100Cm3of oxalate extract will contain=Y2.5×100g=W%  oxalate compositiong/100g=W2.5×1001.


## 2.5.6. Determination of Cyanogenic Glycoside

Cyanogenic glycoside quantitative determination methodology used in this research is that by Amadi et al. [14] as reported by Ejikeme et al. [11]. It was weighed into a 250 cm3 round bottom flask and about 200 cm3 of distilled water was added to one gram of each dry wood powder sample and allowed to stand for 2 hours for autolysis to occur. Full distillation was carried out in a 250 cm3 conical flask containing 20 cm3 of 2.5% NaOH (sodium hydroxide) in the sample after adding an antifoaming agent (tannic acid). Cyanogenic glycoside (100 cm3), 8 cm3 of 6 M NH4OH (ammonium hydroxide), and 2 cm3 of 5% KI (potassium iodide) were added to the distillate(s), mixed, and titrated with 0.02 M AgNO3 (silver nitrate) using a microburette against a black background. Turbidity which was continuous indicates the end point.Content of cyanogenic glycoside in the sample was calculated as(7)Cyanogenic glycosidemg100g=Titre ValueCm3×1.08×exact volumeAliquot volumeCm3×sample weightg×100.


## 2.5.7. Determination of Percentage Lipid

Into a thimble connected to a soxhlet extractor chamber with a preweighed flat bottom was added each dry wood powder (2.50 g) and connected to a condenser. Petroleum ether (100 mL) enough to cause a reflux was added to the flask and the lipid from the wood sample was extracted for 3 hours by heating on an electric hot plate at 50°C. The extractant (petroleum ether) was distilled off and the lipid recovered by cooling the flask in a dessicator and its value calculated by reweighed flask and content. This methodology for lipid quantitative analysis is reported elsewhere [18]. The calculated percentage of lipid content was thus (8)%  Lipid=Weight of LipidWeight of Sample×100.


## 2.5.8. Determination of Phenols

Defatting of 2 g wood powder sample was carried out for 2 hours in 100 cm3 of ether using a soxhlet apparatus. The defatted sample (0.50 g) was boiled for 15 minutes with 50 cm3 of ether for the extraction of the phenolic components. Exactly 10 cm3 of distilled water, 2 cm3 of 0.1 N ammonium hydroxide solution, and 5 cm3 of concentrated amyl alcohol were also added to 5 cm3 of the extract and left to react for 30 minutes for colour development. The optical density was measured at 505 nm. 0.20 g of tannic acid was dissolving in distilled water and diluted to 200 mL mark (1 mg/cm3) in preparation for phenol standard curve. Varying concentrations (0.2–1.0 mg/cm3) of the standard tannic acid solution were pipetted into five different test tubes to which 2 cm3 of NH3OH, 5 cm3 of amyl alcohol, and 10 cm3 of water were added. The solution was made up to 100 cm3 volume and left to react for 30 minutes for colour development. The optical density was determined at 505 nm [10].


## 3. Results

Table1 is a list of the various softwoods whose powdered mills were examined. Their local names in Igbo, Hausa, and Yoruba as well as where they are found in Nigeria are clearly shown. Qualitative determination of phytochemicals as seen in Table 2 showed various plant secondary metabolites in which there was no heavy presence of alkaloid in all the softwood timber powder examined. However, there was slight presence of alkaloid in all of them. Heavy presence of flavonoid was noted only inBarteria nigritiana; other softwoods showed mere presence of it. Phlobatannin was absent in all the softwoods examined. Some presence of terpenoid was observed in some with others not having any presence of the metabolite in them (Table 2).Anogeissus leiocarpus,Dichrostachys cinerea,Afrormosia laxiflora,Sacoglottis gabonensis,Khaya ivorensis, andKaempferia galanga all showed heavy presence of saponin. With the exception ofProtea elliottiiandBarteria nigritiana, other softwoods had slight presence of saponin. The presence of tannin was observed heavily in the following:Protea elliottii,Glyphaea brevis,Uapaca guineensis,Albizia adianthifolia,Dichapetalum barteri,Afzelia bella,Tetrapleura tetraptera,Cordia millenii,and Combretodendron macrocarpum(this was corroborated by Itoandon et al. [19]). Moringa oleifera,Barteria nigritiana,Anogeissus leiocarpus,Allanblackia floribunda,Albizia adianthifolia,Afzelia bipindensis,Pentaclethra macrophylla, andCassipourea barteriwere seen to contain heavy content of steroids. Glycoside was heavily present inMoringa oleifera,Afzelia bipindensis,Tetrapleura tetraptera,Combretodendron macrocarpum, andCordia millenii.Table 1
Table of the botanical and local names as well as location of various Nigerian softwoods.
S/N Wood sample (botanical name) Classification Botanical families Igbo Yoruba Hausa Location 1 Monodora tenuifolia Softwood Annonaceae Ehuru ofia Lakesin Guyiyadanmiya Port Harcourt 2 Moringa oleifera Softwood Moringaceae Okwe oyibo Ewe igbale Zogalla gandi Lagos, Ibadan 3 Protea elliottii Softwood Proteaceae Okwo Dehinbolorun Halshena Nsukka 4 Barteria nigritiana Softwood — Ukwoifia Oko Idon zakara Nsukka, Enugu 5 Anogeissus leiocarpus Softwood Combretaceae Atara Egba Marike Onitsha, Awka 6 Allanblackia floribunda Softwood Guttiferae Egba Orogbo Guthiferae eku Calabar, Ikom 7 Glyphaea brevis Softwood Tiliaceae Anyasu alo Eso, shishi Bolukonu kanana Calabar 8 Sterculia oblonga Softwood Sterculiaceae Ebenebe Aworlwo Kukuki Ibadan 9 Uapaca guineensis Softwood Euphorbiaceae Obia — Wawan kurmi Onitsha 10 Amphimas pterocarpoides Softwood Leguminosae Awo Ogiya Wawan kurmii Umuahia, Ikom 11 Albizia adianthifolia Softwood Leguminosae-Mimosoideae Avu Anyimebona Gamba Enugu, Nsukka 12 Dichapetalum barteri Softwood Dichapetalaceae Ngbu ewu Ira Kirni Onitsha, Agulu 13 Afzelia bipindensis Softwood Fabaceae Aja Olutoko Rogon daji Benin 14 Afzelia bella Softwood Fabaceae Uzoaka Peanut Epa Owerri, Orlu 15 Dichrostachys cinerea Softwood Fabaceae Amiogwu Kara Dundu Onitsha 16 Pentaclethra macrophylla Softwood Leguminosae Ugba Apara Kiriya Onitsha 17 Tetrapleura tetraptera Softwood Leguminosae-Mimosoideae Oshosho Aridan Dawo Onitsha, Akpaka 18 Afrormosia laxiflora Softwood Leguminosae-Papilionoideae Abua ocha Shedun Idon zakara Sokoto 19 Sacoglottis gabonensis Softwood Rhizophoraceae Nche Atala Chediya Rivers 20 Cassipourea barteri Softwood Lecythidaceae Itobo Itobo Odu Eket 21 Combretodendron macrocarpum Softwood Ochnaceae Anwushi Anwushi Akasun Udi, Owerri 22 Cordia millenii Softwood Meliaceae Okwe Okwe — Owerri, Onitsha 23 Khaya ivorensis Softwood Bignoniaceae Ono Oganwo Madachi Calabar 24 Kaempferia galanga Softwood Zingiberaceae Shanty — — EnuguTable 2
Table of the qualitative determination of the phytochemical components of Nigerian softwoods.
S/N Wood sample (botanical name) Alkaloids Flavonoids Glycoside Phlobatannins Tannins Terpenoids Saponins Steroids 1 Monodora tenuifolia ++ + ++ − ++ + + ++ 2 Moringa oleifera ++ ++ +++ − ++ + + +++ 3 Protea elliottii + + ++ − +++ + − ++ 4 Barteria nigritiana + +++ ++ − ++ − − +++ 5 Anogeissus leiocarpus + + − − ++ + +++ +++ 6 Allanblackia floribunda + + ++ − − ++ ++ +++ 7 Glyphaea brevis + + ++ − +++ + + + 8 Sterculia oblonga ++ − ++ − ++ + + − 9 Uapaca guineensis + ++ ++ − +++ ++ + − 10 Amphimas pterocarpoides ++ + ++ − ++ + + − 11 Albizia adianthifolia + + + − +++ − + +++ 12 Dichapetalum barteri + + ++ − +++ + + − 13 Afzelia bipindensis + + +++ − ++ ++ ++ +++ 14 Afzelia bella + + + − +++ − ++ + 15 Dichrostachys cinerea + + ++ − + + +++ − 16 Pentaclethra macrophylla + + ++ − − − + +++ 17 Tetrapleura tetraptera + + +++ − +++ + + ++ 18 Afrormosia laxiflora + + ++ − + + +++ − 19 Sacoglottis gabonensis + + ++ − + + +++ − 20 Cassipourea barteri + + ++ − − − + +++ 21 Combretodendron macrocarpum + + +++ − +++ + + ++ 22 Cordia millenii + + +++ − +++ + + ++ 23 Khaya ivorensis + + ++ − + + +++ − Heavily present: +++; slightly present: ++; present: +; absent: −.Quantitative analysis (Table3) showed that generally small quantities of tannin were found in all the Nigerian softwoods examined with the highest quantities obtained inSterculia oblonga (1240 mg/100 g),Barteria nigritiana (1230 mg/100 g),Albizia adianthifolia (1190 mg/100 g),Afzelia bella (1160 mg/100 g), andCombretodendron macrocarpum (1150 mg/100 g). The highest quantities of alkaloid were obtained inCordia millenii (11.2%),Sterculia oblonga (10.4%),Amphimas pterocarpoides (9.8%), andKhaya ivorensis (9.4%).Barteria nigritiana (14.2%),Moringa oleifera (12.2%) (flavonoid value higher than obtained elsewhere [20] from methanolic extract ofMoringa oleifera leaf (31.73±2.66 mg/g) and seed (20.73±4.16 mg/g)),Combretodendron macrocarpum (9.5%),Cordia millenii (8.2%),Afrormosia laxiflora, andSacoglottis gabonensis (8%) contain the highest quantities of flavonoids. Quantities of saponin inAnogeissus leiocarpus (12.5%) (comparably higher to value (89.5±0.57 mg/dL) obtained elsewhere [21] from stem barks of plant),Dichrostachys cinerea (9.8%),Afzelia bipindensis (6.8%), andSacoglottis gabonensis (6.6%) were highest among the softwoods analyzed. Highest quantities of oxalate were found inCombretodendron macrocarpum (5.84 g/100 g),Glyphaea brevis (3.55 g/100 g),Cordia millenii (3.05 g/100 g),Moringa oleifera (3.2 g/100 g), andBarteria nigritiana (2.91 g/100 g). Cyanogenic glycoside inPentaclethra macrophylla (890 mg/100 g),Moringa oleifera (880 mg/100 g),Tetrapleura tetraptera (890 mg/100 g),Protea elliottii (810 mg/100 g), andCassipourea barteri (860 mg/100 g) was higher than the other softwoods. The highest quantities of phenol found inSacoglottis gabonensis (4.68 mg/g),Pentaclethra macrophylla (4.04 mg/g),Moringa oleifera (3.58 mg/g), andAnogeissus leiocarpus (3.40 mg/g) were comparable but slightly lower than value (5.20±0.17 mg/dL) obtained elsewhere [21] in stem bark ofAnogeissus leiocarpus.Anogeissus leiocarpus (8%),Kaempferia galanga (7.2%),Dichrostachys cinerea (6.8%), andAllanblackia floribunda (6.4%) contained highest quantities of lipids among the softwoods examined.Table 3
Table of quantitative phytochemicals of Nigerian softwoods.
S/N Wood sample(botanical name) Tannin(mg/100 g) Alkaloid(%) Flavonoid(%) Saponin(%) Oxalate(g/100 g) Cyanogenic glycoside(mg/100 g) Phenol(mg/g) Lipid(%) 1 Monodora tenuifolia 900 8.6 7.4 4.4 0.45 680 1.82 3.6 2 Moringa oleifera 930 8.0 12.2 4.2 3.2 880 3.58 3.2 3 Protea elliottii 1030 7.6 5.4 1.6 1.31 810 3.14 0.4 4 Barteria nigritiana 1230 8.4 14.2 1.6 2.91 660 1.90 2.4 5 Anogeissus leiocarpus 920 4.8 3.0 12.5 0.58 340 3.40 8.0 6 Allanblackia floribunda 690 1.6 4.6 5.2 3.05 650 1.58 6.4 7 Glyphaea brevis 980 4.8 7.2 4.4 3.55 730 2.04 4.0 8 Sterculia oblonga 1240 10.4 3.4 4.0 1.31 620 1.66 4.4 9 Uapaca guineensis 1120 8.0 9.2 4.4 0.8 700 1.66 4.8 10 Amphimas pterocarpoides 870 9.8 7.5 3.6 0.69 410 0.86 1.2 11 Albizia adianthifolia 1190 8.0 7.2 2.2 2.56 440 0.49 1.6 12 Dichapetalum barteri 1110 6.0 4.0 3.0 1.02 500 0.04 2.8 13 Afzelia bipindensis 930 7.6 2.0 6.8 2.0 760 0.26 3.6 14 Afzelia bella 1160 5.6 7.0 5.8 1.83 730 0.18 6.0 15 Dichrostachys cinerea 840 4.4 1.8 9.8 2.21 680 1.76 6.8 16 Pentaclethra macrophylla 690 6.4 4.0 4.4 1.6 890 4.04 4.0 17 Tetrapleura tetraptera 1090 4.0 4.4 3.6 1.73 860 1.66 1.2 18 Afrormosia laxiflora 710 8.8 8.0 4.2 0.8 520 1.74 3.6 19 Sacoglottis gabonensis 1090 8.0 8.0 6.6 0.21 300 4.68 4.4 20 Cassipourea barteri 990 9.0 4.8 3.8 0.73 860 2.38 2.8 21 Combretodendron macrocarpum 1150 8.8 9.2 4.6 5.84 730 1.78 3.6 22 Cordia millenii 840 11.2 8.2 3.2 3.05 520 1.70 2.4 23 Khaya ivorensis 790 9.4 7.8 2.8 2.78 520 1.72 1.2 24 Kaempferia galanga 820 3.6 4.0 6.2 2.12 670 1.74 7.2


## 4. Discussion

Softwoods such asSterculia oblonga,Barteria nigritiana,Albizia adianthifolia,Afzelia bella, andCombretodendron macrocarpum, whose qualitative and quantitative determination in this study were shown to contain tannins, could be exploited for their industrial usage. Tannin is one of the major active ingredients found in plant based medicines [22]; it serves as caustics for cationic dyes (tannin dyes) used in the dyestuff industry as well as in the production of inks (iron gallate ink). Other uses of tannin are for wine, fruit juice, and beer clarification in food industries [23] Tannins are used in the dyestuff industry as caustics for cationic dyes (tannin dyes), and also in the production of inks (iron gallate ink), textile dyes, antioxidants in beverages, and coagulant in rubber production [24] as well as possessing antiviral, antibacterial, and antitumor activity [22, 25, 26]. Tannin has been reported to selectively inhibit HIV replication [27]. Therefore, the above listed Nigerian softwoods have potential in the provision of tannin.According to Braunwald et al. [28] cardiac glycoside has been used in treatment of congestive heart failure due to its direct action which increases the force of myocardial contraction. They also explained that in the vascular system cardiac glycoside acts directly on the smooth muscles. Their effects on neural tissues and indirect effect on electrical activities of the heart and vascular resistance as well as capacitance are equally reported [28].Moringa oleifera,Afzelia bipindensis,Tetrapleura tetraptera,Combretodendron macrocarpum, andCordia milleniiin this study were shown to contain glycosides which could be exploited for their medicinal properties.Flavonoids are known to have antioxidant effects and have been shown to inhibit the initiation, promotion, and progression of tumors [29]; reduction of coronary heart disease has been reported to be associated with intake of flavonoid [30]. This research has shown thatBarteria nigritiana,Moringa oleifera,Combretodendron macrocarpum,Cordia millenii,Afrormosia laxiflora, andSacoglottis gabonensis contain appreciable quantity of flavonoid. Apart from the antioxidant properties of flavonoid, other biological functions it possesses include protection against platelet aggregation, microorganisms, hepatotoxins, viruses, tumors, ulcers, free radicals, inflammation, and allergies [31].Phenols are antioxidants in human and plants [32]. Some softwoods in this research such asSacoglottis gabonensis,Pentaclethra macrophylla,Moringa oleifera, andAnogeissus leiocarpusmay be exploited for their saponin content. Haslam [33] points to current interest in the potential for amelioration of diseases simply by improving the dietary intake of nutrients with antioxidant properties, such as vitamin E, vitamin C, β-carotene, and carotenoids, and plant phenolics such as tannins and flavonoids.Plant lipids are important in paints production, pharmaceutical, and cosmetic raw materials as well as being sources for production of animal feeds. From the foregoing, extraction of lipids from nonfoliage parts of softwood (e.g., stems) is ideal for use as listed above. Thus stems ofAnogeissus leiocarpus,Kaempferia galanga,Dichrostachys cinerea, andAllanblackia floribunda from the result of this research are ideal sources for lipid extraction.Appreciable quantities of saponin are found inAnogeissus leiocarpus,Kaempferia galanga,Dichrostachys cinerea, andAllanblackia floribunda as shown in Tables 2 and 3 in this research. Saponins protects against microbial attack in plants; it is also useful in treating yeast and fungal infections [1].According to Sodipo et al. [34] most phytochemicals serve as natural antibiotics, which assist the body in fighting microbial invasion and infections. Alkaloids, for instance, consist of chemical compounds that contain mostly basic nitrogen atoms which occur naturally, mainly, in plants but may be produced by bacteria, fungi, and animals. In this research, appreciable quantities of alkaloid were obtained inCordia millenii,Sterculia oblonga,Amphimas pterocarpoides, andKhaya ivorensis(Table 3). Extraction of alkaloid from these softwoods could serve as a cheap and steady means of providing this secondary metabolite in industries where they are greatly needed. According to literature, alkaloids have a wide range of pharmacological activities including antimalarial (e.g., quinine), anticancer (e.g., homoharringtonine) [35], antibacterial (e.g., chelerythrine) [36], and antihyperglycemic activities (e.g., piperine) [37]. Alkaloids have equally been exploited for their importance in traditional pharmaceutical usage. Other alkaloids possess psychotropic (e.g., psilocin) and stimulant activities (e.g., cocaine, caffeine, and nicotine) and have been used as recreational drugs [37]. Although alkaloids carry out many metabolic activities in humans and other animals, they almost uniformly evoke a bitter taste [38].


## 5. Conclusion

Nigerian softwoods apart from their timber uses are also potential for traditional medicine. There have been well known usages of most of their foliage parts such as seeds and leaves in traditional medicine, but the stem has had less usage in the phytochemical application in industries and medicine. This research result has established through the investigation of their phytochemistry that they have potential in the industries particularly for pharmaceutical usages.



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*Source: 5601327-2016-10-20.xml*