PRODUCTION OF WINE FROM THE FERMENTATION

CHAPTER ONE INTRODUCTION An orange specifically, the sweet orange is the citrus (Citrus sinensis) and its fruit It is the most commonly grown tree fruit in the world The orange is a hybrid of ancient cultivated origin, possibly between pomelo (Citrus maxima) and mandarin (Citrus reticulata) It is an evergreen flowering tree generally growing to 9–10 m in height (although very old specimens have reached 15 m) The leaves are arranged alternately, are ovate in shape with crenulate margins and are 4–10 cm long The orange fruit is a hesperidium, a type of berry (Ramachandra and Arun, 2005) Orange trees are widely cultivated in tropical and subtropical climates for the sweet fruit, which is peeled or cut (to avoid the bitter rind) and eaten whole, or processed to extract orange juice, and also for the fragrant peel In 2008, 68.5 million tons of oranges were grown worldwide, primarily in Brazil and the US states California and Florida (Idise, 2012) The fruit of Citrus sinensis is called sweet orange to distinguish it from Citrus aurantium, the bitter orange The name is thought to derive ultimately from the Sanskrit for the orange tree, with its final form developing after passing through numerous intermediate languages In a number of languages, it is known as a "Chinese apple", e.g., Dutch sinaasappel ("China's apple") or appelsien, or northern German Apfelsine In English, however, "Chinese apple" generally refers to the pomegranate (Idise, 2012) Plate 1: Orange blossoms and oranges on tree Among the greatest and genius invention of man is the incessant production of vivacious and edible alcoholic beverages from fermentation of natural fruit juice such as grape in wine -making Wine has been part of human culture for over 6,000 years, serving dietary and socio-religious functions Its production takes place on every continent and has been enjoyed by many people from peasants to kings and its chemical composition is profoundly influenced by enological techniques (Nikhil et al., 2009).It is produced by fermentation of juice of ripe grapes using a microscopic single-celled organism called yeasts (Saccharomyces cerevisae), digest sugars found in fruit juice, producing alcohol and carbon dioxide gas in the process Other naturally occurring microorganisms may grow in the must or juice, affecting the flavors and aromas of the finished wine For example, lactic acid bacteria use the acids in wine as a source of energy, reducing the wine’s acidity These bacteria also produce other aromas and are responsible for the buttery smells that can be found in wine Sometimes the wine maker restricts the growth of lactic acid bacteria, especially if the wine is already low in acidity or if the buttery character would clash with other aromas of the wine Acetobacter, another type of bacteria, can spoil the wine by converting ethanol to acetic acid to make vinegar Wine naturally contains about 79 to 89 percent water, to 21 percent alcohol, less than percent fruit acids, and hundreds of aroma and flavor components in very small amounts Wines are categorized using a number of different methods Sometimes they are grouped into different categories by grape variety, region of origin, by color, by the name of the wine maker or viticulturist, or by production technique (Bisson et al., 2009).There are two types of wines namely Natural wines with 9-14% alcohol and Fortified (Dessert and appetizer) wines with 15 to 21% alcohol (Nduka, 2007) The nutritional role of wine is important since its average contribution to total energy intake is estimated to be 10 to20% in adult males (Macrae et al., 1993) Although grapes are the most common fruit used to make wine for the past few decades In lieu of this several studies have investigated the suitability of other fruits as substrates for the purpose of wine production (Okunowo et al., 2005) Moreover, the non-availability of grapes, which is usually the fruit of choice for wine production in the tropics has necessitated the search for alternative fruit source in tropical countries; Nigeria (Alobo and Offonry, 2009) Wine is also made from the fermented juice of pears, apples, berries, blackcurrants and even flowers such as dandelions are sometimes used However, are usually referred to as fruits or country wine (Bisson et al., 2009) Any fruit with good proportion of sugar may be used in producing wine and the resultant wine is normally named after the fruit The type of wine to be produced dictates the fruit and strain of yeast to be involved (Amerine and Kunkee, 2005) In contrast to most foods and beverages that spoil quickly or that can spread diseases, wine does not spoil if stored properly This implies that the production of wine from fruits is a form of extending the shelf life of the fruit hence, preservation Wine composition and quality are functions of many different intrinsic and extrinsic variables, many of which are microbiologically mediated A large diversity of microbes are inherent to winemaking including various yeasts, bacteria and fungi Prominent in this process are Saccharomyces species (predominantly Saccharomyces cerevisiae), which dominate the alcoholic fermentation, and the lactic acid bacteria (LAB), which carry out the malolactic conversion Efforts to determine the population size and potential impact of different microbes on the winemaking process are critical to production of a flavourful product Spoilage is considered growth of organisms that are unwanted at any particular place and time in the winemaking process (Sponholz, 1993) Thus the same microorganism can be both beneficial and detrimental to the winemaking process For example, growth of S cerevisiae is required during the alcoholic fermentation, but growth can be detrimental if it occurs in a finished, and bottled, wine Interestingly, both academics and winemakers have good reason to be interested in the microbial ecology of the winemaking process From an academic perspective wine represents an ideal landscape in which to study basic concepts of microbial ecology Several factors promote this view First and foremost, wine is a liquid medium that allows samples to be properly mixed prior to analysis, these contrasts with the situation for those studying microbial growth on solid surfaces (e.g., barrel stave, grape surface or vineyard soil) in which the microbial populations are heterogeneous and spatially distributed across the surfaces As a consequence, representative sampling of microbes on surfaces becomes a more statistically challenging process than sampling of a liquid medium like must or wine A second reason why wine is an attractive platform for microbial ecology is the diversity of microbes present which enables one to witness a range of microbial interactions from commensalisms and neutralisms to antagonisms From the winemaker perspective, close monitoring of the microbial changes occurring throughout the winemaking process is beneficial for several reasons: to promote and guide yeast during the alcoholic fermentation and ultimately to ensure the stability of the wine before bottling and storage (Delfini and Formica 2001) The evolution of undesired microbes during different stages of winemaking can produce volatile acidity, off-flavors and polysaccharide hazes, all of which can diminish the quality and acceptability of the final product (Sponholz, 1993) Prior to the onset of fermentation, the oranges themselves can be infected with moulds, yeasts and bacteria that can enter and alter the fermentation in a negative fashion Improper wine storage and handling post-fermentation can encourage microbiological faults, which can negatively impact wine quality As a result the winemaker must conduct basic physical, chemical, sensory and microbiological analyses of musts and wines to assure wine quality Whether it is for an investigation of basic ecological concepts or for the applied goal of predicting possible wine spoilage, one must have accurate and reproducible methods for enumeration of various microbial constituents at different stages in wine production Both indirect and direct approaches can be used to view these populations Fermentation of food for preservation, enhancement of nutritive values, improvement of flavor and preparation of beverages has been practiced probably since prehistoric times by people of nearly every civilization (Sofos, 1993).The word fermentation comes from the Latin verb fevere, which means to boil Which describes the appearance of the action of yeast on extracts of fruit or malted grain during the production of alcoholic beverages this appearance is due to production of carbon dioxide bubble caused by the anaerobic catabolism of sugar present in the extracts (Stanbury et al., 2003) during the process complex organic substances are broken down into simpler form? However, fermentation is interpreted differently by microbiologists and biochemists To a biochemist the word means process relating to the generation of energy by the catabolism of organic compound, whereas it meaning to a microbiologist tend to be much broader Which include the microbial physiology view, the industrial principle and it application in food processing; the microbial physiology involve the metabolism of organic carbon to generate energy by substrate level phosphorylation and in which organic molecule generated during catabolism of this carbon compound serve as the electron acceptor (reducing agent).while in industrial principle the culturing of microorganism on a large scale, irrespective of the reducing agent this may involve the aerobic phase of production i.e industrial production of antibiotics and powdered (dried) yeast cell, also those that involve the anaerobic phase of production include alcoholic beverages production and ethanol production both are physiologically refers to as fermentation The fermentation of food substances e.g milk, bread etc by microorganism such as lactic acid bacteria, has play an important role in the preservation of the fermented foods and as well as increased the flavor, nutrients, digestibility, security and other characteristics of the food(Barnum, 1998) Fermentation is a process of extracting energy from the oxidation of organic compounds such as carbohydrates using an endogenous electron acceptor, usually pyruvate, an organic compound Before fermentation takes place, one glucose molecule is broken down into two pyruvate molecules during glycolysis Fermentation is important in anaerobic conditions when there is no oxidative phosphorylation to maintain the production of Adenosine tri-phosphate (ATP) by glycolysis Duringalcoholic fermentation, usually carried out by yeasts, pyruvate is then converted into ethanol and carbon dioxide thus: C6H12O6 → 2C2H5OH+2CO2 During this process, the carboxylic carbon atom is released in the form of carbondioxide with the remaining components becoming acetaldehyde The acetaldehyde in the absence of oxygen will then be further reduced by alcohol dehydrogenase to form ethanol along with carbon-dioxide (Robinson, 2006).It is a way for a cell to obtain energy without using oxygen Tropical citrus fruits have been used as substrates for the production of wines and the alcohol profiles Various factors influence the fermentation process and determine the end products obtained These include substrate related factors such as cultivar types, cultivation conditions, conditions at harvest and post harvest handling Yeast species are used in many industrial fermentation processes including alcoholic beverages production The quality of wine produced greatly depends on the yeast strain; development of improved starter organisms for fermentation of citrus juice may offer a relative simple avenue for reducing post harvest wastage of citrus fruits in low utilization environment and in places where the production of citrus concentrates is low or nonexistent Juice concentrates are readily storable and can be used for production processes even when the fruit is out of season In an effort aimed at increasing the low industrial utilization and reducing the high wastage of orange (Citrus sinensis) fruits in the developing world, and investigated the possibility of exploiting the fermentative ability of yeasts to produce orange wines The quantity of alcohol present in orange wines produced from musts fermented by Saccharomyces cerevisae were determined Citrus is the most economically important tree fruit crop in the world The majority of citrus arrives to market in the form of processed products, such as single strength orange juice and frozen juice concentrates In Florida, one of the world’s top producing regions, more than 96% of all oranges are processed in to range juice In the1999–2000 season, Florida produced more than four billion litres of single strength orange juice and as such, the Florida citrus juice industry is considered an important food processing industry in the United States Economic losses due to juice spoilage are minimized by good sanitation procedures before and during citrus processing Pasteurization, concentration, or low temperature storage protocols help to reduce the number of microorganisms in the final product However, these products are not free of microbiological spoilage problems, especially non pasteurized single-strength juices Citrus juices are acidic beverages (pH to 4) with high Sugar content (15°Brix) Under these conditions, acidolactic bacteria, molds, and yeasts comprise the typical microbiota Present In citrus juices Potentiality of Rhizopus Oryzae to utilize orange peels under solid state fermentation conditions to produce macerating fluid with high cellulolytic and pectinolytic activities Tropical fruits have been used as substrates for the production of wines (Maldonado et al., 1975; Anuna et al., 1990; Ndip et al., 2001; Osho, 2005; Okunowo et al., 2005) Various factors influence the fermentation process and determine the end products obtained These include substrate related factors such as cultivar types, cultivation conditions, conditions at harvest and post harvest handling (Daudt and Ough, 1973; Bell et al., 1979; Liu, 2002; Jonathan and Errol, 2000; Joshi and Sandhu, 2000; Kourkoutas et al., 2005) Though the fermentation of fruit sugar usually yields ethanol as the predominant alcohol, small quantities of other higher alcohols (referred to as fusel oil) are also produced from the oxidative deamination, decarboxylation and reduction of amino acids and sugar degradation (Anuna and Akpapunam, 1995) The presence of pectin in some fruits may also result in methanol generation in the fermenting wort (Anuna and Akpapunam, 1995) The alcohol profile is a significant factor in the quality of wines (Drawert and Rapp, 1966; Anuna and Akpapunam, 1995) Yeast species are used in many industrial fermentation processes including alcoholic beverages production The quality of wine produced greatly depends on the yeast strain (Kunkee, 1984; Okunowo et al., 2005) Development of improved starter organisms for fermentation of citrus juice may offer a relative simple avenue for reducing post harvest wastage of citrus fruits in low utilization environment and in places where the production of citrus concentrates is low or nonexistent Juice concentrates are readily storable and can be used for production processes even when the fruit is out of season (Ramachandra and Arun, 2005; Siddik et al., 2006) For example, in India alone an estimated loss of around 35 000 million Indian Rupees (around 638 million US $) worth fruits and vegetables was recorded despite the fact that India produces around 60 to 65 million tons of fruits and vegetables (Ramachandra and 10 35 Plate 5: Orange wine 36 CHAPTER FOUR 4.0 DISCUSSION Nwachukwu et al., (2006) established that five species of Saccharomyces were isolated from aged palm wine; Saccharomyces cerevisiae, Saccharomyces ellipsoideus, Saccharomyces cheresanus, Saccharomyces oviformis, Saccharomyces logos The isolated species had many similarities; The Ability of the isolate to grow in 10% Sodium chloride + 50% glucose medium confirmed that the isolate is saccharomyces cerevisae The growth test distinguishes saccharomyces cerevisae from all other species Evaluating the result during orange wine production, the drop in pH and corresponding increase in titratable acidity of must during the aerobic phase and increase in pH with decrease in titratable acidity during anaerobic phase is attributable to yeast metabolism The increase in titratable acidity at early stage of fermentation was due to production of acid which accompanied the fermentation of sugar The decrease towards the end of the fermentation is as a result of the acid being used up by the fermenting yeast to bring about desirable flavour in the wine (omojasola et al., 2003) These also show acidification of the medium during the fermentation stages, which is crucial to wine production Acidity plays a vital role in determining wine quality by aiding the fermentation process and enhancing the overall characteristics and balance of the wine Lack of acidity will mean a poor fermentation (Berry, 2000) The pH obtained for the final products fall within acidity level of sweet and dry wines Usually, acidity of wines lies between pH and for dry wine and 3.5 to 4.5 for sweet wine 37 Higher acidities are sometime encountered with fortified and sparkling wines (Bisson and Butzkc, 2007) Studies have shown that during fermentation of fruits, low pH is inhibitory to the growth of spoilage organisms but creates conducive environment for the growth of desirable organisms Also, low pH and high acidity are known to give fermenting yeasts competitive advantage in natural environments (Reddy and Reddy, 2005) The increase in the total yeast count during aerobic fermentation can be attributed to the presence of utilizable sugar (sucrose) and yeast nutrient The daily aeration of the fermenting must will also have aided rapid multiplication of the yeast cells (Berry, 2000) In the aerobic phase of fermentation the yeast metabolize carbohydrate using Embden-Mayerof-parnas pathway (Okafor, 1987) During the anaerobic fermentation stage, yeast was noted to have stopped increasing rather decreasing towards the end of the fermentation This is likely because during this stage alcohol levels had increased in the fermenting medium During anaerobic growth the yeast utilizes intermediate products like acetaldehydes as hydrogen acceptors for alcohol production (Okafor, 1987; Prescott et al., 2008) Acetaldehyde serves as the terminal electron acceptor; pyruvate is first decarboxylated, by pyruvate decarboxylase, to yield CO and acetaldehyde Acetaldehyde is then reduced to ethanol by alcohol dehydrogenase (enzyme present in S.cerevisiae), thus regenerating NAD1+ (Horsey, 2007) S cerevisae is a facultative anaerobe which produces alcohol efficiently and tolerates higher levels of ethanol than other fungi In the aerobic phase of orange must fermentation which lasted for five (5) days, alcohol was produced This can be attributed to the high 38 concentration of yeast cells and high concentration of sugar in the must The fermenting vessel which was litre in volume, it was filled to the level of litres This provides little airspace in between the surface of the must and the muslin cloth for diffusion of oxygen It was observed that yeasts create a micro anaerobic environment around themselves as they submerge by using oxygen atoms trapped in the spaces between the molecules surrounding them(Jackeller, 2004) A similar observation was made by Crabtree (1929) In the evaluation of different yeast strains to produce different fruit wines The primary fermentation took place for 12 days with the vessel been constantly agitated to supply oxygen to the containing yeast cells Fruits tested produced substantial alcohol within the 12 days It was observed that, at high glucose concentrations, even under aerobic conditions, S.cerevisiae metabolism is fermentative rather than oxidative This is associated with morphological and biochemical alterations in the mitochondria, which resemble those in cells grown under anaerobic conditions It is the uptake of hexose sugars that is the rate-limiting step in glycolysis, and it has been shown that there is more than one type of glucose transport, depending primarily on substrate affinity These mechanisms are referred to as ‘low-affinity’ and ‘high affinity’ uptake In order to supplement the sugar content of the musts, sucrose was part of the additives Reports have shown that the major problems associated with the use of tropical Fruits in wine production are their low sugar content (Aloboand Offonry, 2009) Remarkable amount of alcohol was produced from the fruit wines during fermentation The specific gravity and total sugars decreased throughout the fermentation process This was as a result of activities of the 39 fermenting yeast The yeast converts the sucrose to glucose sugars which is being utilized for growth and subsequently produces ethanol and CO2 (Okafor, 1987) The final specific gravity obtained (0.980osp.gr) falls within the 1.000 and 0.990 osp.gr range for wine (Jack, 2007) The final alcohol content of the wine (9.6%) ranks it among good table wines According to Michael (2000) a good table wine must have alcohol content between and 14% 40 4.1 CONCLUSION AND RECOMMENDATION This study is based on the evaluation of orange as a substrate for wine production and the efficiency of locally isolated yeast (Saccharomyces cerevisiae) from palm wine for production of fruit wine The results obtained from the fermentation shows that acceptable wine could be produced from orange must The study has also given an insight efficiency and role of local yeast strains during alcoholic fermentation of fruits and orange as good and acceptable substrates for wine production This study has demonstrated that it is possible to produce wines from locally available fruit with good microbiological standard and high acceptability 41 REFERENCES Adaikan, P., Ganesan, A.A (2004) Mechanism of the Oxytoxic activity of Comosus proteinases Journals of Pharmaceutical Biology 42(8): 646-655 Alobo, A.P., Offonry, S.U (2009) Characteristics of Coloured Wine produced from Roselle (Hibiscus sabdariffa) calyx extract J Institute Brewing 115(2): 91-94 Amerine, M.A., Kunkee, R.E (2005) Microbiology of Winemaking Am Rev Microbiol 22: 232-258 Association of Analytical Communities International (2000) Official Methods of Analysis of AOAC International (17th edition) Vol II, Gaithenburg, MD, USA, Official Method 920.57 Awe, S., Sani, A and Ojo, F.T (2009) Microbiological quality of some selected spices (Thymus vulgaris, Murrayakoenigiand Piper nigrium) Niger J.Microbiol 23(1): 18721877 Awe, S (2011) Production and Microbiology of Pawpaw (Carica papaya L) Wine Current Research Journal of Biological Sciences 3(5): 443-447 Barnett, J.A., Payne, R.W and Yarrow, D (2000) Yeast Characteristics and Identification 3rd Edn., Cambridge University Press, Cambridge, pp: 1139 42 Bassir, O (1962) Observation on the fermentation of Palm wine West Afr J Biol Appl Chem 6: 20-25 Berry, C.J.J (2000) First Steps in Wine Making Published by G.W Kent, Inc 3667 Morgan Road, Ann Arbor M I 48108, pp: 235 Bisson, L.F and Butzkc, C.E (2007) History of Wine making.WINE, Microsoft Encarta Cabib, E., Silverman, S.J., Shaw, A., Das Gupta, S., Park, H., Mullins, J.T., Mol, P.C and Bowers, B (1991) Carbohydrates as structural constituents of yeast cell wall and septum Pure and Applied Chemistry Volume 63, p483-489 Crabtree H.G (1928) Biochemistry of alcoholic fermentation Biochem j 22(5):1289-98 Dubois,M., Gilles, K., Hamilton, J.K., Rebers, P.A and smith, F (1956) phenol-sulphuric acid calorimetric estimation of carbohydrates Analytical chemistry 28:300-350 Dupree, W.E., Woodruff, J.G and Siewert, S (1953) Watermelon rinds in food products Ga Agr Expt Stat Bul 285, 30pp Elijah, A.I., Ojimelukwe, P.C., Ekong, U.S., Asamudo, N.U (2010) Effect of Sacoglottisgabonensis and Alstoniaboonei on the kinetics of Saccharomyces cerevisiae isolated from palmwine Afri.J of Biotechnol 9(35): 5730-5734 Ezeronye, O.U ( 2004) Nutrient utilization profile of Saccharomyces cerevisiae from palm wine fruit fermentation Antonie van Leeuwenhoek 86 (3): 235-240 43 FAO (1994) Production Yearbook 1993 47th Edition.Food and Agric Org, United Nations, Rome Faparunsi, S.I and Bassir, O (1972) Factors affecting palm wine, Period of Tapping West Afr J Biol Appl Chem 15: 24-32 Fawole, M.O.and Oso, B.A (1995) Laboratory Manual in Microbiology.Spectrum books Limited Spectrum House, Ring Road, Ibadan, Nigeria, pp: 1-48 Fleet, G.H (1993) Wine: Microbiology and Biotechnology Harwood Academic Publishers, London.p 130 Fleet, G.H (2003).Yeast interactions and wine flavour International Journal of Food Microbiol 86: 11-22 Heller, K.J (2001) Probiotic bacteria in fermented foods: product characteristics and starter organisms Ame J Clin Nutr 73: 374S-379S Idise, O.E, Izuagbe, Y.S (1988) Microbial and chemical changes in bottled palm wine during storage Nigeria Journals of Microbiology 8(1): 175-184 Idise, O (2012) Studies of wine produce from pineapple (Ananascomosus) International journalfor biotechnology and molecular biology research 3(1): 1-7 Ihekoroye, A.I and Ngoddy, P.O (1985).Integrated Food Science and Technology for the Tropics, Macmillan Publisher, London 44 Isitua , C.C and Ibeh (2010) Novel method of wine production form banana (Musa acumunata) and pineapple (Ananascomosus) waste African journal of Biotechnology (7): 914-922 Jack, B.K (2007) Wine Making Retrieved from: http://www.Jackkeller.net, (Accessed on: September 06, 2010) Joshi, V.K and Bhutani, V.P (1991).The influence of enzymatic clarification in fermentation behaviour and qualities of apple wine Science of Aliments 11: 491-498 Kunkee, R.F., Amerine, M.A (2002) Yeast in Wine Making In: Rose, H.A and Harrison, J.S (Eds) The Yeast-Academic Press, London pp 5-71 Landry, C.R., Townsend, J.P., Hartl, D.L and Cavalieri, D (2006) "Ecological and evolutionary genomics of Saccharomyces cerevisiae".Molecular Ecology Volume 15 p59-575 Leal, F., Coppensd’Eeckenbrugge, G., and Holst, B.K (1998).Taxonomy of the genera Ananas and Pseudananas– a historical review Selbyana: 19: 227-235 Lourens-Hattingh, A., Wiljoen, B.C (2001) Growth and survival of a probiotic yeast in dairy products Food Resolution International.34 (9): 791-796 45 Macrae, R., Robinson, R.K and Sadler, M.J (1993) Alcohol fermentation Encyclopedia for food technology and Nutrition Academic Press Limited, London Harcourt Bracce Jovanovich Publishers, London, San Diego, New York, Boston, 3: 1735-1842 Martini, A (1993) "Origin and Domestication of the Wine Yeast Saccharomyces cerevisiae".Journal of Wine Research 4: 165-176 Michael, P (2000) Foods of the Gods: Part 1-Wine in Ancient Egypt Retrieved from: http://www.touregypt.net/Egypt-info/magazine-mag11012000-magf.htm, (Accessed on: May 12, 2012) Ndip, R.N., Akoachere, J.F., Dopgima, L.L and Ndip, L.M (2001).Characterisation of yeast strains for wine production: Effect of fermentation variables on quality of wine produced, Journal of Applied Biochemistry and Biotechnology 95(3): 209-220 Nester, E.W., Anderson, D.G., Roberts, C.E., Pearsall, N.N., Nester, M.T (2004) Microorganisms in Food and Beverage Production- Alcoholic fermentation by Yeast: In Microbiology-A Human Perspective 4th edition, McGraw Hill, New York, USA pp 151-153 Nwachukwu, I.N Ibekwe, V.I Nwabueze, R.N and Anyawu, B.N (2006) Characterization of palm wine yeast isolate for industrial utilization African journal of biotechnology Vol 5(19):1725-1728 46 Obire, O (2005) Activity of Zymomonas species in palm sap obtained from three areas in Edo state, Nigeria Journal of Applied Science and Environmental Management 9: 25-30 Okafor, N (1975a) Microbiology of Nigeria Palm wine with particular reference to bacteria Journals of Applied Bacteriology 38: 81-88 Okafor, N (1975b) Preliminary microbiological studies on the preservation of palm wine Journals of Applied Bacteriology 43: 159-161 Okafor, N (2007) Modern Industrial Microbiology and Biotechnology.Science Publishers, Enfield, NH, USA p530 Okafor, N (1975) Preliminary microbiological studies on the preservation of palm wine Journals of Applied Bacteriology 38: 1-7 Okunowo, W.O., Okotore, R.O.and Osuntoki, A.A (2005) The alcoholic fermentative efficiency of indigenous yeast strains of different origin on orange juice African journal of Biotechnology., 4: 1290-1296 Omojasola, P.F and Ademuyiwa, F.O (2003).Some characteristics of Roselle wine produced from fermentation of hibiscus sabdariffa using saccharomyces cerevisiae Nigeria Journal of Pure & Applied science 18: 1401 – 1406 47 Oyeleke, F.I and Olaniyan, A.M (2007).Extraction of juice from some tropical fruits using a small scale multi-fruit juice extractor African journals of Crop Science.Proc 8: 18031808 Polunin, I (1951).Pineapple dermatosis British Journal of Dermatology: 63 p441 Prescott, L.M., Harley, J.P and klein, D.A (2008) Microbiology 7th Edition.,McGraw-Hill Companies, Inc., 1221 Avenue of the Americas, New York Ramachandra, H.G.R., Arun, H.K (2005) Spray drying of mango juice Journal of Diary Science 85: 395-404 Reed, G and Nogodawahames, T.W (1991) Yeast Technology Journals of applied Science (2): 310-320 Robinson, J (2006) The Oxford Companion to Wine 3rd Ed Oxford University Press pp 268-780 Rohrbach, K.G Leal, F and Coppensd’Eeckenbrugge, G (2003) History, distribution and world production In: Bartholomew, D.P, Paull, R.E and Rohrbach, K.G (eds) The Pineapple:Botany, Production and Uses CABI Publishing, Oxon, UK pp 1-12 Siddiq, M., Harte, J., Dolan, K., Khan, A., Collins, J.K., Perkins Veazie, P.M (2006) The use of lycopene-rich water melon juice or juice concentrate in different food system International Food Technology Meeting Abstracts p223 48 Sofos, J.N (1993) Current microbiological considerations in food preservation International Journals of Food Microbiology 19: 87-108 Thais, M., Guimarães Danilo, G M., IaraP Cyntia, M T., Tania, M.B (2006) Isolation and characterization of Saccharomyces cerevisiae strains ofwinery interest Brazilian Journal of Pharmaceutical Sciences vol 42 Uzochukwuru, B.U.A., Balogh, F.E and Ngoddy, P.D (1991).Standard pure culture inoculum of natural fermented Palm sap Nigeria Journals of Microbiology 9:67-77 Webber, H.J., Walter, R Harry, W.L., Willard, H (1967) The Citrus Industry, Horticultural Varieties of Citrus Riverside CA: University of California Division of Agricultural Sciences 49