Nigerian Food Journal Official Journal of Nigerian Institute of Food Science and Techonology www.nifst.org

NIFOJ Vol. 31 No. 2, pages 1 – 7, 2013

Evaluation of Chemical Properties of Mistletoe Leaves from Three Different Trees (Avocado, African Oil Bean and Kola)* Ishiwu, C.N.1, Obiegbuna, J.E.1 and Aniagolu, N.M.2

ABSTRACTA comparative evaluation of the chemical composition of mistletoe leaves (Viscum album) growing on three different trees: avocado pear (Persea Americana), African oil bean (Pentaclethra macrophylla) and kola (Kola nitida) was undertaken. Fresh mistletoe leaves were obtained from the three different trees and thoroughly washed with water after which they were dried at room temperature (28 ±10C) for 18 days during the dry season (December). After, the dried leaves were shredded and packaged. The samples were then subjected to phyto-chemical, vitamin and mineral analyses using standard methods of analyses. Anthocyanins ranged between 0.3.4 ± 0.003 and 0.37 ± 0.181 mg/g, Tannin, 2.09 ± 1.141 and 3.24±0.003 mg/g, Chlorophyll a, 0.38 ± 0.004 and 0.39 ± 0.018 mg/g, Chlorophyll b 0.52 ± 0.120 and 0.60 ± 0.001 mg/g. Vitamin B1 ranged between 0.85 ± 0.038 and 0.94 ± 0.001 mg/g, Vitamin B2, 0.55 ± 0.007 and 0.61 ± 0.003 mg/g, Vitamin C, 0.77 ± 0.003 and 1.98 ± 0.003 mg/g, Folate 0.53 ± 0.004 and 0.58 ± 0.004 mg/g, Magnesium ranged between 0.21 ± 0.002 and 0.92 ± 0.003 mg/g, Calcium 2.14 ± 0.004 and 2.26 ± 0.001 mg/g, Sodium 0.01 ± 0.000. Iron, 1.24 ± 0.005 and 1.42 ± 0.006 mg/g. Sample obtained from oil bean tree showed lower and differed significantly (p < 0.05) in Anthocyanin and Chlorophyll b with samples obtained from avocado and kola trees. The sample obtained from avocado tree showed higher Tannin content compared with samples obtained from oil bean and kola trees (P ≤ 0.05). There were significant differences (P < 0.05) between the samples in all the vitamins. The minerals showed significant differences in the samples (P < 0.05) except in sodium content which was very low in all the samples. Mistletoe leaves are shown to have a rich chemical composition and could thus serve as a source of these phyto-chemicals, vitamins and minerals.

Keywords: Trees, mistletoe, phyto-chemicals, vitamins, minerals

IntroductionAfrican mistletoes (Loranthus micranthus) have tough, oblong, green leaves, attractive flowers and waxy, translucent white berries with a viscous mesocarp (Kernzehola et al., 1997). Mistletoes are considered as parasitic plants growing wild on the branches or trunks of economic trees in the tropics. Mistletoe grows wild on various economic trees in Nigeria and are usually cut down and destroyed. This is due to the damages they cause to their host trees which lead to huge economic loss. Mistletoes anchor themselves to their host by means of

adventitious roots that penetrate through the host tissues and adversely affect the growth and fruiting performance of the host tree. Mistletoe takes water and minerals from its host, but not carbohydrates which it can produce by photosynthesis (Hostanska et al., 1995; Solar et al., 1998).

The North Americans and Germans use mistletoe as a ceremonial plant, while the Greeks believe that it has mystical powers (Brown, 1992). The Druids use mistletoe to welcome the New Year and also for religious rites and medicinal purposes, such as treating sterility, diabetes, epilepsy and some other chronic diseases (David, 2001).

It is scantly used in Nigeria by a few individuals as a remedy for several human and animal ailments which

1 Department of Food Science and Technology, Nnamdi Azikiwe University, Awka-Nigeria.

2 Department of Food Technology Institute of Management and Technology, Enugu-Nigeria

* corresponding author : nwaejaka@yahoo.com

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include diabetes, dysentery, wounds, diarrhoea, cancer and hypertension (Akinmoladum et al., 2007). Several studies have revealed that mistletoe has considerably antioxidant potential, justifying its therapeutic use in herbal medicine (Obatomi et al., 1994; Deeni and Sadiq, 2002). Aqueous extract of mistletoe has been reported to normalize blood sugar and cholesterol levels in rat (Iheanacho et al., 2008). Mistletoe contains lectins which are protein that could bind sugars and possibly reduce the blood sugar level (Hostanska et al., 1995).

A few rural subsistent farmers use the leaves to feed their goats that have newly given birth to kids. However, most individuals do not even know about its nutritional and health benefits or perhaps they are skeptical about its chemical composition which has not been well published. The amount of chemical compounds may vary in mistletoe leaves depending on the source of the leaves.

The objective of this study is to compare the chemical composition of mistletoe leaves plucked

from three different trees (Avocado pear tree, African oil bean tree and kola tree) in Nigeria.

Materials and MethodSource of raw materialFresh mistletoe leaves were harvested from three different trees such as Avocado pear, African oil bean and kola tree respectively from Affa Town in Udi local Government Area, Enugu, Nigeria.

Preparation of raw materialThe fresh mistletoe leaves were thoroughly washed with clean water separately according to how they were collected from different trees; they were then dried during the dry season at room temperature (28±1oC) for 18 days, after which they were separately shredded and packaged.

Key:O = Mistletoe leaf obtained from oil bean treeA = Mistletoe leaf obtained from avocado pear

treeK = Mistletoe leaf obtained from kola tree.

Fig. 1: Fresh mistletoe leaf from avocado (Persea americana) tree

Evaluation of Chemical Properties of Mistletoe Leaves from Three Different Trees ... Ishiwu et al. 3

Fig. 2: Fresh Mistletoe leaf from Kola (Kola nitida) tree

Fig. 3: Fresh <mistletoe leaf from Oil bean (Pentaclethra macrophylla) tree

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AnalysisDetermination of anthocyaninThis was done gravimetrically by the method of Harborne (1973). A 5 g of each test sample was hydrolyzed by boiling in 100 ml or 2 M HCl solution for 30 min. The hydrolysate was filtered using Whatman No. 41 filter paper. The filtrate was transferred into a separating funnel and an equal volume of ethyl acetate was added to it, mixed and allowed to separate into two layers. The ethyl acetate layer (supernatant) was recorded while the aqueous layer was discarded. The supernatant was concentrated to dryness in the crucible over a steam bath. The dried extract was then treated with conc. amyl alcohol which extracted the anthocyanin. After extraction, the alcohol extract and the filtrate were transferred to a weighed evaporating dish and evaporated to dryness. It was then dried in the oven at 30oC for 30 min and cooled in a desicator. The weight of anthocyanin was determined and expressed as mg/g.

Determination of tanninTannin content was determined by Folin-Denis colorimetric method described by Kirk and Sawyer (1998). 5 g sample powder was dispersed in 50 ml hot (80oC) distilled water and shaken. The mixture was allowed to cool to 28oC before it was filtered through Whatman No. 41 filter paper. 2 ml of the extract was dispersed into a 50 ml flask. Similarly, 2 ml standard tannic acid solution and 2 ml of distilled water were put in separate volumetric flasks to serve as blank. 2.5 ml standard Folin-Denis reagent was added to each of the flask and 2.5 ml of saturated NaCO3 solution also added. The content of each flask was made up to 50 ml with distilled water and incubated at 28oC for 90 min. Their respective absorbance was measured in a spectrophotometer (Spectronic 21) at 260 nm using the blank to calibrate the instrument at zero.

Determination of Vitamin C2, 6-dichlorophenol indophenol titration method was used for the determination of Vitamin C on 5 g sample as described by Pearson (1991).

Determination of Vitamins B1, B2 and folate These were determined using the method of AOAC (1990).

Determination of mineralsThe concentrations of selected minerals in the leaves were determined using Atomic Absorption Spectrophotometer (AA320N), while sodium was determined using flame photometer. The minerals were determined after wet oxidation of 2 g of the sample powder using concentrated nitric acid and perchloric acid as described by Osborn and Voogt (1978).

Statistical analysis of dataData generated from all analyses were subjected to statistical analysis using SPSS version 17.0 through one way analysis of variance (ANOVA). Significant difference between samples was tested at P = 0.05 using LSD.

Results and DiscussionFigures 1, 2 and 3 show the pictures of the mistletoe leaves plucked from the avocado, kola and oil bean trees respectively. These leaves are closely related in texture (tough) and appearance (green).

The phyto-chemical composition of the samples is shown in Table 1. The samples showed significant and high content of anthocyanin (0.34 ± 0.003 – 0.37 ± 0.0181 mg/g) compared with eight different Nigerian green leafy vegetables whose anthocyanin ranged between 0.0 2 ± 0.00 and 0.1 ± 0.02 g/100 g (Onyeka and Nwambekwe, 2007). Generally, green leafy vegetables are not known to contain very high amount of anthocyanins (Dewanta et al., 2002). The highest recorded amount of anthocyanin appears to be specific in the seed coat of black soybean containing 2000 mg/100 g (Choung, 2001). The tannin content was high in all the samples (2.09 ± 1.141 – 3.24 ± 0.003 mg/g) and chlorophyll A (0.38 ± 0.004 – 0.39 ± 0.018 mg/g) and chlorophyll B (0.52 ± 0.120 – 0.60 ± 0.001 mg/g). The result showed that the sample obtained from avocado tree has the highest anthocyanin and tannin, but this variation may be due to the composition of the host tree. With regard to chlorophyll A

Evaluation of Chemical Properties of Mistletoe Leaves from Three Different Trees ... Ishiwu et al. 5

and B, samples A and K have the highest values. The phyto-chemicals have been linked with the bioactive principles responsible for medicinal activities of most medicinal plants and herbs and this dictates why efforts have been expanded in studies aimed at elucidating their levels in medicinal plant (Edeoga et al., 2006). Other researchers have also reported that the medicinal value of plants and vegetables are dictated by their phyto-chemicals and Ether extractable chemical constituents (Fallah et al., 2005). This result also revealed that the leaves obtained from oil bean tree differed significantly with leaves obtained from avocado and kola nut tree (P < 0.05) in Anthocyanin and Chlorophyll A

and B. There was a significant difference between the three samples in tannin content. The sample obtained from avocado tree showed the highest tannin content followed by the sample obtained from oil bean tree. The sample obtained from kola tree ranked least in tannin content. Excessive consumption of a tannin-based diet could bind with proteins and make protein unavailable. It has been reported that phyto-chemicals are plant chemicals that give plants colours and smell and are plant compounds that contain natural disease-resistant compounds and also have been associated with reducing risk of degenerative diseases in humans (Anderson and Barrett, 1994).

Table 1: Phyto-chemical composition of the samples (mg/g)

Sample Anthocyanin Tannin Chlorophyll A Chlorophyll B

O 0.34 ± 0.003b 2.79 ± 0.002b 0.38 ± 0.004a 0.56 ± 0.004b

A 0.37 ± 0.005a 3.24 ± 0.003a 0.39 ± 0.003a 0.60 ± 0.001a

K 0.37 ± 0.181a 2.09 ± 1.141c 0.39 ± 0.018a 0.52 ± 0.120c

Data are means of triplicate determinations ± SD. Data in the same column bearing different superscripts differed significantly (P < 0.05)

Key:O = Mistletoe leaf obtained from oil bean treeA = Mistletoe leaf obtained from avocado pear treeK = Mistletoe leaf obtained from kola tree.

Table 2 shows the vitamins content of the samples. Vitamin C was the highest in the samples obtained from avocado (1.98 ± 0.003 mg/g), kola tree (1.95 ± 0.005 mg/g) and oil bean tree (0.77 ± 0.003 mg/g). Vitamin B1 ranked next to vitamin C even though none of the samples was up to 1 mg/g of vitamin B1. Vitamin B2 ranged between 0.55 ± 0.007 and 0.61 ± 0.003 mg/g and folate ranged between 0.53 ± 0.004 and 0.58 mg/g. Vitamins are micronutrients necessary for normal development, maintenance and repair of body tissues. The result showed that the sample obtained from avocado tree exhibited the highest values of vitamin B1, B2 and

C comparatively, and there is significant difference (P < 0.05) between the samples in all the vitamins. Mistletoe leaves, therefore, could serve as source of these vitamins.

The mineral contents of the samples are shown in Table 3. The samples showed highest value for calcium (2.14 ± 0.005 – 2.26 ± 0.001 mg/g) followed by iron (1.24 ± 0.005 – 1.42 ± 0.0060 mg/g). None of the samples had magnesium content of up to 1 mg/g. The magnesium content of the samples ranged between 0.21 ± 0.002 and 0.92 ± 0.003 mg/g. Sample obtained from kola tree had the lowest magnesium content (0.021 ± 0.002

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mg/g). All the samples showed very low amount of sodium which was 0.001 mg/g for all the samples.

Magnesium is an important mineral element whose deficiency is in connection with circulatory diseases such as heart disease. Without magnesium, many enzymes in the human body would function less efficiently and magnesium contributes to calcium and potassium metabolism and therefore essential for bone strengthening (McLean, 1994). The sample from avocado tree showed the highest value of magnesium. There is significant difference (P < 0.05) between the magnesium content of samples from avocado tree and the other trees. Calcium is important in teeth, bone and muscle metabolism (Turan et al., 2003). RDA for calcium for adults is

800 mg/day (Alaimo, 1994). According to Shills and Young (1988), a food rich in calcium content should be greater than 1 mg/g and poor if less than 0.5 mg/g. From the result, the dried mistletoe leaves samples are very rich in calcium and iron which is an essential trace element for haemoglobin formation, for normal central nervous system and energy metabolism (Ishida et al., 2000). The sodium content of all the samples was too low (≤0.01 mg/g). The implication of low concentration of sodium in the samples is that consumption of extract of mistletoe leaves sourced from any of the three trees cannot aggravate hypertension in consumers of the extract which high sodium intake has been linked with.

Table 2: Vitamin composition of the samples (mg/g)

Sample Vit. B1 Vit. B2 Vit. C Folate

O 0.87 ± 0.004b 0.58 ± 0.002b 0.77 ± 0.003c 0.58 ± 0.004a

A 0.94 ± 0.001a 0.61 ± 0.003a 1.98 ± 0.003a 0.57 ± 0.003b

K 0.85 ± 0.038c 0.55 ± 0.007c 1.95 ± 0.005b 0.53 ± 0.004c

Data are means of triplicate determinations ± SD. Data in the same column bearing different superscripts differed significantly (P < 0.05)

Table 3: Mineral composition of the sample (mg/g)

Sample Mg Ca Fe Na

O 0.81 ± 0.003b 2.14 ± 0.004a 1.27 ± 0.003b 0.01 ± 0.000a

A 0.92 ± 0.003a 2.26 ± 0.001b 1.24 ± 0.005c 0.01 ± 0.000a

K 0.21 ± 0.002c 2.14 ± 0.003a 1.42 ± 0.006a 0.01 ± 0.000a

Data are means of triplicate determinations ± SD. Data in the same column bearing different superscripts differed significantly (P < 0.05)

Evaluation of Chemical Properties of Mistletoe Leaves from Three Different Trees ... Ishiwu et al. 7

ConclusionThe comparative analyses of chemical composition of mistletoe leaves from different trees showed variations in the amount of these chemical compounds evaluated. Mistletoe leaves obtained from avocado tree showed the highest concentration of tannin, vitamin B1, vitamin C and calcium. The leaves obtained from oil bean tree showed highest concentration of anthocyanin, magnesium, chlorophyll b and folate, while sample obtained from kola tree exhibited highest amount of iron when the leaves from the three different trees were compared. However, there were no significant difference in the chlorophyll a and sodium content of the samples.

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