Characterization Of Prebiotic Inulin Isolated Biology Essay

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Prebiotics are basically any non-digestible food ingredients that can benefit the human body by stimulating the growth and activity of one or more species of probiotics in the large intestine and produce short chain fatty acids(SCFAs) resulting in a decrease of the colon pH [1]. Lowering the pH of the colon assists in the reduction of pathogenic bacteria and viruses which cause gastro-intestinal diseases. A probiotic on the other hand is a live microorganism which, when administered in adequate amount, confers a health benefit to the host [2]. Probiotics are the names given collectively to all the beneficial bacteria in the human colon. These friendly bacteria ferment the prebiotic substances.

Prebiotics are mostly oligosaccharides in chemical nature, but they may also be non carbohydrates. These prebiotics are classed as soluble fibres which include various short chain fructo-oligosaccharide, inulin, gums and various lactose derivatives such as tagatose, malto-oligosaccharide, polydextrose, resistant starches and many others.[1] Although in present day literatures, a large number of macromolecules having chemical similarity with oligosaccharides are termed as prebiotics according to Roberfroid, only two particular fructo-oligosaccharides, namely, oligofructose and inulin fully meet the definition of prebiotics. These short-chain prebiotics after being fermented provide nourishment to the bacteria present in the right hand side of the colon. On the other hand long chain prebiotics viz. Inulin are fermented in a slower rate but their functions are same as that of the short chain prebiotics although they provide nourishment to the bacteria present in the left hand side of the colon. Full-spectrum prebiotics, e.g., oligofructose-enriched inulin (OEI) provide the full range of molecular link lengths from 2-64 links per molecule and nourish bacteria throughout the colon. Inulin has been defined as a polydisperse carbohydrate material consisting mainly, if not exclusively, of β-(2-1) fructosyl-fructose links[5]. Inulin producing plant species are found in several monocotyledonous and dicotyledonous families, including Liliaceae, Amaryllidaceae, Gramineae, and Compositae. So far, only one inulin-containing plant species (Chicory. Chichorium intybus) is used to produce inulin industrially. In chicory inulin, both Gpy-Fn(α- D-glucopyranosyl-[β-D- fructofuranosyl]n-1 β-D-fructofuranoside) compounds are considered to be included under the same nomenclature. Native inulin is processed by the food industry to produce either short chain fructans, specially oligofructose (degree of polymerization: 2-10; average:5) as a result of partial enzymatic hydrolysis (inulinase, Ec3.2.1.7) or long chain fructans by applying an industrial physical separation technique. Prebiotic carbohydrates are also found naturally in fruits and vegetables like bananas, berries, asparagus, garlic, wheat, oatmeal, barley ( other whole grains), flaxseed, tomatoes, Jerusalem-artichoke, onion, greens ( especially dandelion greens but also spinach, collard greens, chard, kale mustard greens and others) and legumes ( lentils, kidney beans, chickpeas, navy beans, black beans) [5]. Inulin supports larger total bacterial counts but lowers populations of bifidobacteria and lactobacilli than soybean oligosaccharides (SOS), isomalto-oligosaccharides (IMO) or lactulose.

Understanding the importance of isolation and purification of inulin from it's native source, in the present investigation an attempt has been made to isolate inulin from six candidate vegetable, fruit and cereal sources namely, onion, garlic, banana, asparagus, oat and wheat. The main objective of the proposed work is to characterize inulin so isolated according to the functional group present in the chemical structure using Fourier transformed infrared spectroscopy. A pure food grade sample of inulin ( Made-Himeia) has been used as the basis of comparison. It is expected that such study will lead to identify a suitable native source for proactive inulin prebiotic.

MATERIAL AND METHODS:

Materials: Onion, Garlic, Banana, Asparagus, Oat, Wheat were purchased from local market.

Food grade inulin was purchased from himedia, pune, India and this sample was used as a standard.

Instruments: FTIR Spectroscope, model- VERTEX 70

Method of isolation:

The candidate prebiotic native sources namely onion, garlic and asparagus after procuring from market were washed properly with distilled water. In case of onion, garlic and banana peeling of outer skin was done manually and the core portion of the individual item was taken for study. In case of asparagus, the root and bud were taken for study excluding the stem portion. All individual material mentioned above was taken in a mortar- pestle and manually crushed to the desired consistency level. 10 ml distilled water was added to this paste and with the help of a glass rod the contents were well stirred for 10 minutes at 300C. The resulting aqueous mixture was then filtered using whatman filter paper grade 47, dia 111mm, to isolate water soluble inulin from other associated water insoluble materials. The filtrate containing isolated inulin was subjected to FTIR spectroscopy for it's chemical characterisation.

Result and Discussion:

In order to identify the potential source of inulin amongst the six materials taken for study, chromatogram analysis has been carried out to arrive at a conclusive decision. For all the test cases comparison has been done with that of the pure food grade sample. Figure 1 shows the FTIR chromatogram of the standard food grade inulin sample and this has been used for comparison purpose. Figures 2 through 8 show the FTIR chromatograms of garlic, asparagus bud, asparagus root, banana, oat, onion and wheat respectively.

These FTIR chromatograms have been studied both qualitatively and quantitatively. A careful comparison qualitatively shows that chemical composition of inulin isolated from garlic is identical with the standard sample throughout the wavelength scaned. This indicates that the garlic is enriched with inulin and is a potential source of the same. Figures 3 and 4 clearly indicate that asparagus buds and it's root are also potential sources of inulin. Chromatogram of banana, wheat, onion and oat do not clearly indicate their inulin content.

In order to arrive at a decisive conclusion a quantitative study along with the qualitative observation is of utmost important.

A close comparison of chromatogram figure 1 and 2 evidently indicates that the centroid of the main maximum of pure inulin sample and the isolated one from the garlic fall on the same wavelength of approximately 3320 cm-1. Thus quantitatively it may be stated that the product isolated from garlic is highly enriched with inulin. It is also observed from chromatogram figures 3 and 4 that the centroid of main maximum in these two cases also laid in the same wavelength as that of the pure inulin. Thus asparagus stem and bud can also be considered as the potential source of inulin. When chromatograms of banana, oat, wheat and onion (figures 5-8) are compared with the pure sample of inulin, it is observed that the centroids of main maximum shift to about 40 cm-1almost in each individual case.

From the above observation it can be inferred that garlic and asparagus are potential sources of inulin which can be conveniently extracted by standard protocols. On the other hand the other materials studied for extraction of inulin()do not appear to be a potential source for commercial extraction of inulin. It may be noted that for tropical developing countries in Asia and Africa availability of asparagus is limited and for the obvious reason it's cost is prohibitively high. Thus garlic seems to be a potential source of inulin from industrial point of view.

FTIR Analysis:

Figure 1: FTIR chromatogram of Inulin

Figure 2: FTIR chromatogram of Garlic

Figure 3: FTIR chromatogram of Asparagus buds

Figure 4: FTIR chromatogram of Asparagus root

Figure 5: FTIR chromatogram of Banana

Figure 6: FTIR chromatogram of Oat

Figure 7: FTIR chromatogram of Onion

Figure 8: FTIR chromatogram of Wheat

Figure 25: FTIR chromatogram comparison between garlic, banana, oat, asparagus, wheat, onion

In order to ascertain whether inulin have been extracted from the candidate food sources in reality, FTIR chromatogram technique has been used in the present investigation. A sample of pure food grade inulin purchased from open market was subjected o FTIR. Result is shown in figure 16. Extracted inulin from the different candidate food sources were then subjected to the FTIR analysis in a similar manner as was done in the case of pure inulin(figure 16). Results so obtained for each individual samples are shown in figure 17 to 22. In order to compare the results the FTIR chromatogram results of the individual samples were superimposed on the FTIR diagram of pure inulin. This is shown in figure 23. On closure examination it reveals that isolated prebiotic in each case of the candidate food materials is substantially pure inulin.

7. CONCLUSION:

Preliminary study on isolation of prebiotic from different sources has been initiated and its effect on probiotics is also being studied. However, a programmed and systematic study is necessary to proceed further in this investigation. Experimental runs have been designed and all out efforts are being made to perform experiment as per plan.

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