Evaluation Of Proximate Constituents In Leafs And Roots Of Simarouba Glauca D.C (Paradise Tree)

Project and Seminar Material for Biochemistry

Evaluation Of Proximate Constituents In Leafs And Roots Of Simarouba Glauca D.C (Paradise Tree)


The proximate content; moisture, crude protein, crude ash, crude fat, crude fibre and carbohydrate, of Simarouba glauca were determined, along with other properties to include the glucose and amino acid contents, and the qualitative macromolecular content of the plant. The proximate properties of the plant were determined based on the A.O.A.C (1990) methods for moisture, crude protein and crude ash; A.O.A.C (1984) for crude fibre and carbohydrates; and the acid base extraction technique described by Phillips et.al. (2001). The concentration of different amino acids were determined using the ninhydrin, while the glucose content was determined Nelson-Simogy’s method. The moisture contents of the

leaves and the roots were quite high, over 60% in both. The root of the plant was also found to be rich in fibre and low in fat, meanwhile the leaves recorded a higher fat concentration than fibre. With the moisture and fibre content of the roots taking a large percentage, it was found to be pretty low in crude ash, carbohydrates, crude protein and crude fat. The leafs however was much higher in ash, carbohydrates and protein, and a little higher in fat. Among the essestial amino acids, methionine was the highest while phenyl alanine was negative. The highest amino acid was cysteine, which is a conditionally essential amino acid. From these results, it is obvious that Simarouba glauca roots and leaf extract might not enough nutritional supplements while being used medicinally. They can serve as a good source of animal feed, however, proteins and fat would need to be supplemented.

Chapter One


1.1 Introduction

Plants undergo photosynthesis and they constitute a primary resource of carbon, vitamins, minerals, protein, essential fatty acids, and utilizable energy for food production (Young and Pelett, 1994). Plants have played a significant role in maintaining the health and improving the quality of human life for thousands of years. (Mishra, 2010). They provide a major source of food and nourishment for man and animal.

Nutrition is a science of food and its relationship to health. Nutrition refers to nourishment that sustains life. The study of nutrient requirements and the diet providing these requirements is also known as ‘nutrition’ (Chutani, 2008). Pike and Brown, 1984 defined it as “the science that interprets the relationship of food to the functioning of living organism. It includes the uptake of food, liberation of energy, elimination of wastes and all the processes of synthesis essential for maintenance, growth and reproduction (Chutani, 2008). Apart from maintaining normal body functioning, nutrition is important in fighting infections and in the recuperation of an ill person. Nutrition interacts with infections in a synergistic manner, such that recurrent infections lead to a loss of body nitrogen and worsen nutritional status; the resulting malnutrition, in its turn, produces a greater susceptibility to infection (Kurpad, 2005). Aristotle (384-322 B.C.) was the first to suggest that the composition of foods in the normal diet might contribute to health.

In an 1897 literature on metabolic investigations, Atwater divided food composition into five classes; protein, fat carbohydrate, energy and water. However, today, proximate composition is the term usually used to describe six components of food namely; moisture, crude protein, crude ash, crude fibre, crude fat and carbohydrate (nitrogen free extract) which are all expressed in percentage (%) or gram per 100 grams (g/100g). The study of proximate analysis on foods was devised over a hundred years ago by two German scientists, Henneberg and Stohmann, and even though new techniques have been introduced, their system of proximate still forms the basis for the statutory declaration of the composition of foods. (Dublecz, 2011).

1.2 Moisture Content

Water is essential for every living organism. In the human body, water content ranges from 50-70% in different tissues. It is present in different fluid compartments of the human body- Intracellular (fluid inside the cells) and extra cellular. Plasma, interstitial fluid, cerebrospinal fluid, ocular fluid, lymph, peritoneal, pericardial, pleural and synovial fluids are part of the extra cellular fluid (Chutani, 2008). However, the moisture content of a feed is seldom of interest nutritionally as water is usually taken on its own.

The active ingredients from the view of feed nutrition are present in the part of dry matter (solid matter); therefore the level of moisture content is an important factor in both economy and storage. At high temperature and humidity the risk of putrefaction is predicted due to the proliferation of molds, etc., or self-digestion by enzymes in the feed when moisture in the feed is not less than about 15%. As the assay for moisture in the feed measures loss on drying by heating at normal pressure as moisture, the result includes most of volatile substances other than H2O. Therefore, it may be more appropriate to be referred to as volatile matter rather than moisture for accuracy. Organic acids such as acetic acid and butyric acid in silage as well as ammonia and flavor components in feed materials are also vaporized and thus measured as moisture. Because the content of these in most feed is extremely low, there has hardly been a need to consider their influence on the measured value. (Chutani, 2008).

1.3 Carbohydrates

Photosynthesis is a process used by plants and other organisms to convert light energy from the sun, into chemical energy that can be later released to fuel the organisms’ activities. The light energy harnessed from the sun drives the reduction of carbon from CO2 to produce O2 and fixed carbon in form of carbohydrate.

Early in the twentieth century, it was mistakenly thought that light absorbed by photosynthetic pigments directly reduced CO2 which then combined with water to form carbohydrate. In fact, photosynthesis in plants is a two stage process in which light energy is harnessed to oxidise H2O:

2H2O → O2 + 4 [H+]

The electrons thereby obtained subsequently reduce CO2:

4H+ + CO2 → (CH2O)n + H2O

The two stages of photosynthesis are traditionally referred to as the light reactions and the dark reactions:

1.In the light reactions, specialised pigment molecules capture light energy and are thereby oxidized. A series of electron- transfer reactions which culminate with the reduction of NADP+ to NADPH, generate ATP from ADP + Pi. The oxidised pigment molecules are reduced to H2O, thereby generating O2.

2.The dark reactions use NADPH and ATP to reduce CO2 and incorporate it into the three-carbon precursors of carbohydrate.

The light reactions takes place in the thylakoid membrane of chloroplasts in leaves and green parts of plants. The inside of the thylakoid is referred to as the lumen. The light reactions are catalysed by enzymes located in the thylakoid membrane, whereas the dark reactions take place in the stroma. The principal photoreceptor of light is chlorophyll. These chlorophyll molecules do not participate directly in photochemical reactions but function to act as light harvesting antennas. The absorbed photons are transferred from molecule to molecule until it reaches the photosynthetic reaction centre.

In the respiratory chain, electrons flow from NADH + H+ to O2, with the production of water and energy. However in photosynthesis, electrons are taken up from water and transferred to NADP+, with an expenditure of energy. Photosynthetic electron transport is therefore energetically “uphill work.” To make this possible, the transport is stimulated at two points by the absorption of light energy. This occurs through two photo systems protein complexes that contain large numbers of chlorophyll molecules and other pigments Another component of the transport chain is the cytochrome bf complex, an aggregate of integral membrane proteins that includes two cytochromes (b563 and f). Plastoquinone, which is comparable to ubiquinone, and two soluble proteins, the copper containing plastocyanin and ferredoxin, function as mobile electron carriers. At the end of the chain, there is an enzyme that transfers the electrons to NADP+. Because photosystem II and the cytochrome b/f complex release protons from reduced plastoquinone into the lumen, photosynthetic electron transport establishes an electrochemical gradient across the thylakoid membrane, which is used for ATP synthesis by an ATP synthase.

ATP and NADPH + H+, which are both needed for the dark reactions, are formed in the stroma. (Voet et al., 2013).

1.3 Calvin cycle

The actual CO2 fixation i.e., the incorporation of CO2 into an organic compound is catalysed by ribulosebisphosphate carboxylase/oxygenase (“rubisco”). Rubisco, the most abundant enzyme on Earth, converts ribulose 1,5-bis-phosphate, CO2 and water into two molecules of 3-phosphoglycerate. These are then converted, via 1,3-bisphosphoglycerate and 3-phosphoglycerate, into glyceraldehyde 3-phosphate. In this way, 1,2-glyceraldehyde 3-phosphates are synthesized from six CO2. Two molecules of this intermediate are used by gluconeogenesis reactions to synthesize glucose 6-phosphate. From the remaining 10 molecules, six molecules of ribulose-1,5-bisphosphate are regenerated, and the cycle then starts over again. In the Calvin cycle, ATP is required for phosphorylation of 3-phosphoglycerate and ribulose-5-phosphate. NADPH + H+, the second product of the light reaction, is consumed in the reduction of 1,3-bisphosphoglycerate to glyceraldehyde-3- phosphate.

Carbohydrates are the most abundant biomolecules produced on earth; photosynthetic plants and algae convert over 100 billion metric tons of CO2 and water into sugars, starches, and cellulose like substance. Carbohydrates supply energy for the human body to function. They are the most abundant bulk nutrients and form the major source of biological energy through their oxidation in the tissues. They also furnish organic precursors for the biosynthesis of many cell components. Carbohydrates are not essential in the human diet, but because carbohydrate rich foods are abundant and cheap, compared with fats and protein, they naturally form a major part of the diet in most of the world. (Voet et al., 2013).

1.4 Proteins

The requirements for total protein, at various stages during the life cycle of humans, were reviewed and evaluated by a joint panel of the Food and Agriculture Organization, the World Health Organization, and the United Nations University (FAO/ WHO/UNO).

The requirement for dietary protein consists of two components:

  1. The requirement for the nutritionally indispensable amino acids (histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine) under all conditions and for conditionally indispensable amino acids (cysteine, tyrosine, taurine, glycine, arginine, glutamine, proline) under specific physiological and pathological conditions and
  2. The requirement for nonspecific nitrogen for the synthesis of the nutritionally dispensable amino acids (aspartic acid, asparagine, glutamic acid, alanine, serine) and other physiologically important nitrogen-containing compounds such as nucleic acids, creatine, and porphyrins.

With respect to the first component, it is usually accepted that the nutritive values of various food protein sources are to a large extent determined by the concentration and availability of the individual indispensable amino acids. Hence, the efficiency with which a given source of food protein is utilized in support of an adequate state of nutritional health depends both on the physiological requirements for the indispensable amino acids and total nitrogen and on the concentration of specific amino acids in the source of interest (Young and Pelett, 1994).Simarouba Glauca

Proteins, are synthesized from a complex series of steps which involves the transcription of DNA already present in each cell of an organism, and its consequent transcription into a polypeptide chain. This chain is modified by other inherent mechanisms in the cell to yield protein. Simarouba Glauca

1.5 Crude Ash

Ash is the inorganic residue remaining after the water and organic matter have been removed by heating in the presence of oxidizing agents, which provides a measure of the total amount of minerals within a food. The ash content is a measure of the total amount of minerals present within a food. Minerals are required for many purposes like forming the frame and rigid structure of the body, as part of the body/cell fluids and for number of cellular and sub cellular physiological functions (Chutani, 2008). The mineral content includes specific inorganic components present within a food, such as Ca, Na, K and Cl. Determination of the ash and mineral content of foods is important for a number of reasons. The most important reason in regards a plant like S. glauca is the nutritional importance. Some minerals are essential to a healthy diet (e.g. calcium, phosphorous, potassium and sodium) whereas others can be toxic (e.g. lead, mercury, cadmium and aluminum).Simarouba Glauca

1.6 Fat and Fibre

The importance of fat and fibre in nutrition cannot be underestimated. Crude fat contains fat, complex lipid, sterols, fatty acids and fat soluble dyes; while crude fibre contains cellulose, hemicellulose, and lignin. Simarouba Glauca

1.7 Aim of the Research Work

This study is designed to screen the proximate constituents of the leaf extracts of Simarouba glauca which includes moisture, protein, carbohydrates, ash fibre and lipid content of Simarouba glauca. In addition to this, to determine the quantitative carbohydrate and amino acid constituents of Simarouba glauca. Thus, the nutritional value of Simarouba roots and leaves with some emphasis on their possible use both as a medicinal and nutritional food for the sick or convalescent.

Chapter Five

Discussion and Conclusion

5.1 Discussion

This study investigated the proximate properties of the leaf and root extract of Simarouba glauca D.C. It also went on to investigate the glucose and amino acid constituent of the extracts.

5.1.1 Moisture Content

The moisture content of both the leaf and the root are 62 ± 2.70% and 65.89 ± 0.76% respectively. Though the moisture content is seldom of interest in terms of nutritional value, it is an important factor in determining the storability or dry substance content of the plant. The high moisture content of Simarouba glaucaleaves and roots indicates that it is prone to microbial growth and deterioration. Therefore, only fresh leaves and root extract will be safe and contain useful constituents for its nutritional or medicinal use.

5.1.2 Crude Protein and Amino Acids

The crude protein measured in Simarouba roots and leaves showed revealed relatively low values and therefore Simarouba glauca may not serve as an important source of nutritional protein in the diet of humans or in animal feed. Patil and Gaikwad (2011) reported that the protein content in the Simarouba Kernels were 18.2g/100g which increased to 47.7g/100g in defatted meal of Simarouba. This means that in use as animal feed, Simarouba glauca roots and leaves can be supplemented with Simarouba kernels as a protein source. Amino acids are constituents of proteins. The constituents of essential amino acids, Isoleucine, leucine, methionine, phenylalanine, and valine were determined. Also, assays were done to determine the constituents of other amino acids such as tyrosine, glycine, alanine, arginine, glutamate, and cysteine. Glycine, arginine and phenylalanine showed negative values, indicating their absence or that their presence is negligible in Simarouba glauca. The sulphur containing amino acids, methionine and cysteine were the highest in the roots and leaves (20.06 ± 0.12 and 176.21 ± 8.14) and (18.56 ± 0.04 and75.78 ± 2.72) respectively as opposed to the low values recorded in Simarouba seeds by Patil and Gaikwad. From the results the amino acid constituent of Simarouba root and leaves extracts were low, which is expected from their low crude protein values. Among the essential amino acids assayed for, methionine gave the highest result of 20.06 ± 0.12 in leaves and 18.56 ± 0.04 in the roots; while phenylalanine was the lowest, yielding negative values in both leaves and roots. Other essential amino acids, isoleucine, leucine and valine were within the range of 5.99 13.79 for the leaves and 3.51 -7.49 for the roots.

5.1.3 Crude Ash

The ash content of Simarouba glauca leaf was determined to be about 5%. This is a relatively high value. That of the root was found to be much lower at a value of about 1.5%, though still a considerable value in terms of nutrition. Minerals are required for many purposes like forming the frame and rigid structure of the body, as part of the body/cell fluids and for number of cellular and sub cellular physiological functions (Chutani, 2008). Simarouba glauca can leaves and root extract can therefore help replenish electrolyte and osmotic balance while being administered as a medicinal plant to the sick or convalescent.

5.1.4 Carbohydrate, Glucose and Crude Fibre

The carbohydrate content of the leaves of Simarouba is much higher when compared with the root, 17.7% and 0.69% respectively. The reverse is the case of crude fibre content, 14.0 ± 2.0% and 31.4 ± 4.4% respectively. This implies that the leaves of Simarouba is a more important energy giving source than the root. Its glucose content is also significantly higher, however, 0.851 ± 0.164% as against 0.235 ± 0.107 for the root. These values are however too low to prove problematic to a diabetic patient. While the leaves are a more suitable energy giving source, the roots are richer in fibre content. A generous intake of dietary fiber can be provided by the roots of Simarouba glauca. Increased consumption of dietary fiber improves serum lipid concentrations, lowers blood pressure, improves blood glucose control in diabetes, promotes regularity, aids in weight loss, and appears to improve immune function (Anderson et al., 2009).

From the analysis Simarouba glauca leads and roots has very minimal crude fat percentage, 0.32% and 0.24% respectively. Jeyarani and Reddy (2001) reported that the seeds contain 40% Kernels and the kernels contain 60% fat, which is edible. This again reveals the inferior nutritional value of the leaf and root of Simarouba glauca when compared with its seeds and kernel.

5.2 Conclusion

Conclusively, Simarouba glauca root and leaves have relatively low nutritional value, with the leaves being higher in protein, minerals, fat and carbohydrates; while the roots are higher in moisture and fibre. They should therefore not be used as a major source of nourishment, in additional to their medicinal value, for the sick or convalescent. Since the seeds and fruits of Simarouba have been found to have high nutritional constituents, it is recommended Simarouba leaves and root extract can be administered medicinally, while a meal of Simarouba seeds and kernel is provided along with other sources of nourishment for the sick and the convalescent. Also, Simarouba roots and leaves can serve as a basic source of animal feed, with supplements of proteins added to the feed.

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