GRIN - The effect of temperature on yeast growth (2023)


1.1 Background of study
1.2 Statement of Problem
1.3 Objective of Study
1.4 Statement of Hypothesis
1.5 Scope of Study
1.6 Limitation of Research
1.7 Definition of Unfamiliar Terms

2.1 History
2.1.0 History of Yeast (Saccharomyces cerevisiae)
2.1.1 General consideration and taxonomy
2.1.2 Scientific classification:
2.1.3 Morphological Characteristics
2.1.4 Biochemical Characteristics
2.1.5 Yeast metabolism
2.1.6 Ecology and Natural Habitats of Yeast
2.2 Uses of yeast
2.3 Medicinal Value of Yeast on Human Health
2.4 Effect of Temperature on the Growth of Saccharomyces cerevisiae
2.5 How Temperature Affect Yeast
2.6 Temperature range that kills yeast
2.7 Fermentation of yeast
2.7.1 Effect of temperature on yeast fermentation
2.7.2 Other factors affecting yeast fermentation
2.8 Health benefit of fermentation
2.8.1 Improves Digestion
2.8.2 Suppresses Helicobacter pylori
2.8.3 Anticancer Effects
2.8.4 Enhances Bioavailability of Nutrients
2.8.5 Reduces Symptoms of Lactose Intolerance
2.9 Economic Importance of fermentation
2.10 Economic Importance of Yeast

3.1 Materials
3.1.1 Apparatus
3.1.2 Samples
3.2 Collection of Sample
3.3 Application of Temperature on Saccharomyces cerevisiae (yeast) using puff-puff production as a basal technique
3.3.1 Saccharomyces cerevisiae (Yeast) preparation of different water temperature but the same room storage effect on flour paste.
3.3.2 Saccharomyces cerevisiae (Yeast) preparation of the same water temperature but different room storage effect on flour paste.





Table 1 Natural yeast habitats

Table 2 Normal water temperature at 26oC

Table 3 Warm water temperature at 67oC

Table 4 Hot water temperature at 80oC

Table 5 The same water temperature (26oC) at different initial room temperature


Figure 1 Budding yeast

Figure 2 Fission in yeast

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Figure 3 Candida formation in yeast

Figure 4 Yeast cell


The objectives of this study are to evaluate to study the effect of temperature on the growth of yeast using puff-puff production as a basal techniques, to study how temperature affect the growth of yeast. Two methods were adopted in this study which includes yeast preparation of different water temperature but the same room storage effect on flour paste and yeast preparation of the same water temperature but different room storage effect on flour paste. Hygienically packed flour, sugar and yeast were purchased from a retailer at market and was taken to School Microbiology Laboratory for analysis, to determine how temperature affect yeast at different water temperature but the same room storage on flour paste and different water temperature of 26oC, 67oC and 80oC respectively, was poured into three different beakers labelled Sample A, Sample B and Sample C. In each beakers yeast was poured and stirred then left for thirty minutes before adding flour which was stirred smoothly to form a paste. The paste was transferred into three different 500ml measuring cylinder. The samples were later placed at the same room stored which was observed between the ranges of five minutes interval for thirty minutes to see the effect of the temperature on the flour paste. While, the second analysis was to determine how temperature effect yeast at same water temperature but different room storage on flour paste the same water temperature of 26oC, was poured into three different beakers labelled Sample A, Sample B and Sample C. Sample A was stored in the refrigerator, Sample B was stored at room temperature and Sample C was stored in the sun, after two hours it was observed. At higher temperature the cells became less reactive as the cell did not multiply. This could be as a result of temperature that alters the action of the yeast on the mixture, as high temperature denatures the enzymes and stop fermenting because it activates the particles to vibrate too fast and burst the enzymes made of protein molecules in the yeast. Saccharomyces cerevisiae (Yeast) grow much slower and finally cease growing when its temperature gets either too low or too high.

Keywords: Growth, Saccharomyces cerevisiae (Yeast) and Temperature.



1.1 Background of study

Yeasts are eukaryotic single-celled microorganisms classified as members of the fungus kingdom. The first yeast originated hundreds of millions of years ago, and 1,500 species are currently identified (Kurtzman, 2006). They are estimated to constitute 1% of all described fungal species (Kurtzman, 2006). Yeasts are unicellular organisms that evolved from multicellular ancestors, (Yong, 2012) with some species having the ability to develop multicellular characteristics by forming strings of connected budding cells known as pseudohyphae or false hyphae (Kurtzman, 2006). Yeast sizes vary greatly, depending on species and environment, typically measuring 3–4μm in diameter, although some yeasts can grow to 40 μm in size. (Walker, 2002) Most yeasts reproduce asexually by mitosis, and many do so by the asymmetric division process known as budding.

Yeasts, with their single-celled growth habit, can be contrasted with molds, which grow hyphae. Fungal species that can take both forms (depending on temperature or other conditions) are called dimorphic fungi ("dimorphic" means "having two forms"). By fermentation, the yeast species Saccharomyces cerevisiae converts carbohydrates to carbon dioxide and alcohols – for thousands of years the carbon dioxide has been used in baking and the alcohol in alcoholic beverages (Legras, 2007). It is also a centrally important model organism in modern cell biology research, and is one of the most thoroughly researched eukaryotic microorganisms. Researchers have used it to gather information about the biology of the eukaryotic cell and ultimately human biology (Ostergaard, 2000). Other species of yeasts, such as Candida albicans, are opportunistic pathogens and can cause infections in humans. Yeasts have recently been used to generate electricity in microbial fuel cells, (Ostergaard, 2000) and produce ethanol for the biofuel industry.

Temperature is one of the most important physical parameters which has a direct influence on yeast growth and fermentation performance (Watson, 1987). Although many types of yeast exploited for alcohol production, wines and baked products are mesophilic organisms which are capable of growth between 0oC and 48oC, the preferred temperature for Saccharomyces yeasts is between 25 to 35oC (Watson, 1987). It is known that yeast growth rate and metabolism increases when temperature is raised from sub-optimal to optimal temperatures and decreases when temperature is increased (Thevelein, 1984).

It is known that there are various stresses which occur prior to and during the fermentation processes including temperature, osmotic, pH, nutrient deprivation, and stresses associated with the accumulation of ethanol and carbon dioxide (Gibson et al., 2007). In order to maintain fermentation performance as well as survive, bioethanol yeast strains must cope with these environmental changes by relying on their physiological response mechanisms (Grosshans et al., 2006).

However, the optimum temperature for yeast growth is a narrow range and analysis of the metabolic response of Saccharomyces cerevisiae to continuous heat stress has demonstrated that when the temperature is increased to 43oC, yeast cells began to lose their viability (Miyazaki, 2005).

1.2 Statement of Problem

There are various types of yeast in the market. They range from the very expensive to the inexpensive, with various degrees of quality. A longer rise time has always been an issue due to a room that is a little too cold / hot or it could be that moist of the yeast was dead.

1.3 Objective of Study

The objectives are:

i. To study the effect of temperature on the growth of yeast using mixture.
ii. To study how temperature affect the growth of yeast and to have a basic knowledge on yeast consumption.
iii. To compare the growth of yeast in different temperature.
iv. To gather a background knowledge of their concentrations and to assess their potential nutritive and medicinal benefits

1.4 Statement of Hypothesis

H0: Temperature has no effect on the exponential growth rate of Saccharomyces cerevisiae.

HA: Temperature has an effect on the exponential growth rate of Saccharomyces cerevisiae.

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1.5 Scope of Study

The scope of this study was to examine the effect of temperature on yeast growth. Temperature range study was 26°C, 67°C and 80°C. Then, time series of yeast's growth was determined by initial water temperature and different room storage.

1.6 Limitation of Research

The major limitation of this project is that it can only be used to determine the effect of temperature between the ranges of 26oC to 80oC.

1.7 Definition of Unfamiliar Terms

Arbutin: Arbutin may be made synthetically or derived from plants, has antioxidant properties, and can help brighten an uneven skin tone.

Archaeologists: A person who studies human history and prehistory through the excavation of sites and the analysis of artefacts and other physical remains.

Aneuploidy: The condition of having an abnormal number of chromosomes in a haploid set.

Blastic: Having a given type or number of buds, cells, or cell layers,” or “undergoing a given type of development,” as specified by the initial element: holoblastic.

Fermentation: The chemical breakdown of a substance by bacteria, yeasts, or other microorganisms, typically involving effervescence and the giving off of heat.

Liquefy: To reduce to a liquid state.

Mesophilic: A mesophile is an organism that grows best in moderate temperature, neither too hot nor too cold, typically between 20 and 45 °C (68 and 113 °F).

Oxidatirely: The combination of a substance with oxygen.

Phylogenetic: relating to the evolutionary development and diversification of a species or group of organisms, or of a particular feature of an organism.

Physiological: relating to the branch of biology that deals with the normal functions of living organisms and their parts.

Teleomorph: sexually reproductive form in the life cycle of any fungus of the phyla Ascomycota and Basidiomycota. Anamorph: The asexual reproductive form in the life cycle of any fungus of the phyla Ascomycota and Basidiomycota.

Thallic: containing thallium, especially in the trivalent state.

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Viability: ability to survive or live successfully.

Recombinant: relating to or denoting an organism, cell, or genetic material formed by recombination.



2.1 History

2.1.0 History of Yeast (Saccharomyces cerevisiae)

The word "yeast" comes from Old English gist, gyst, and from the Indo-European root yes-, meaning "boil", "foam", or "bubble". Yeast microbes are probably one of the earliest domesticated organisms. Archaeologists digging in Egyptian ruins found early grinding stones and baking chambers for yeast-raised bread, as well as drawings of 4,000-year-old bakeries and breweries (Phillips, 2016). In 1680, Dutch naturalist Anton van Leeuwenhoek first microscopically observed yeast, but at the time did not consider them to be living organisms, but rather globular structures Huxley (1871) as researchers were doubtful whether yeasts were algae or fungi (Ainsworth, 1976). Theodor Schwann recognized them as fungi in 1837 (Barnett, 2004).

In 1857, French microbiologist Louis Pasteur showed that by bubbling oxygen into the yeast broth, cell growth could be increased, but fermentation was inhibited – an observation later called the "Pasteur effect". In the paper "Mémoire sur la fermentation alcoolique," Pasteur proved that alcoholic fermentation was conducted by living yeasts and not by a chemical catalyst (Barnett, 2003).

By the late 18th century two yeast strains used in brewing had been identified: Saccharomyces cerevisiae (top-fermenting yeast) and Saccharomyces carlsbergensis (bottom-fermenting yeast). Saccharomyces cerevisiae has been sold commercially by the Dutch for bread-making since 1780; while, around 1800, the Germans started producing Saccharomyces cerevisiae in the form of cream. In 1825, a method was developed to remove the liquid so the yeast could be prepared as solid blocks (Klieger, 2004). The industrial production of yeast blocks was enhanced by the introduction of the filter press in 1867. In 1872, Baron Max de Springer developed a manufacturing process to create granulated yeast, a technique that was used until the First World War In the United States, naturally occurring airborne yeasts were used almost exclusively until commercial yeast was marketed at the Centennial Exposition in 1876 in Philadelphia, where Charles L. Fleischmann exhibited the product and a process to use it, as well as serving the resultant baked bread (Snodgrass, 2004).

The mechanical refrigerator (first patented in the 1850s in Europe) liberated brewers and winemakers from seasonal constraints for the first time and allowed them to exit cellars and other earthen environments. For John Molson, who made his livelihood in Montreal prior to the development of the fridge, the brewing season lasted from September through to May. The same seasonal restrictions formerly governed the distiller's art (Denison, 1955).

2.1.1 General consideration and taxonomy

The yeasts described in this review are all members of the phylum Ascomycota and the class Saccharomycota. Phylogenetic analysis of the phylum Ascomycota has significantly changed yeast classification in recent years (Hibbett et al., 2007). Yeasts are eukaryotic microorganisms widespread in natural environments including the normal microbial flora of humans, on plants, on airborne particles, in water, in food products, and in many other ecological niches. Yeasts are important in many complex ecosystems, as frequent early colonizers of nutrient-rich substrates (Kurtzman et al., 2011). They are involved in many interactions with other microorganisms, including symbiosis, mutualism, parasitism, and competition. They also exhibit both asexual and sexual states. The asexual state of given yeast is called the anamorph, while the sexual state is the teleomorph. One result of this phenomenon is that there is a valid Latin name for each state, since no teleomorph has been found for many asexual forms or because the phylogenetic relationship between anamorph and teleomorph has not been confirmed.

The most common mode of vegetative growth of yeasts is by budding, which may be blastic or thallic. Anamorphic and teleomorphic genera may grow either as a “yeast-like” unicellular organism or as a “mold-like” filamentous organism, a phenomenon called dimorphism. Moreover, some species are able to form a true mycelium, while genera such as Candida produce a well-developed pseudomycelium, or both pseudo and true mycelium in the case of Candida tropicalis (Goldman, 2008).

Among the yeasts belonging to the phylum Ascomycota, the genus Saccharomyces is the most studied. Many of the approximately 20 species of this genus are of great biotechnological significance due to applications including alcoholic fermentation, bread-making, single cell protein, vitamin production, synthesis of recombinant proteins, and biological control (Webster and Weber, 2007). The most significant species is certainly Saccharomyces cerevisiae (baker’s and brewer’s yeast), due to its economic impact. Saccharomyces cerevisiae is used for the annual production of an estimated 60 million tons of beer, 30 million tons of wine, 800,000 tons of single cell protein, and 600,000 tons of baker’s yeast (Pretorius et al., 2003). The vegetative cells of Saccharomyces cerevisiae are normally diploid, but some strains have been reported as aneuploidy or tetraploid (Webster and Weber, 2007). Over the past four decades, a yeast first identified as Saccharomyces boulardii has been studied for its potential probiotic use (Buts, 2009).

2.1.2 Scientific classification:

- Kingdom: Fungi
- Phylum: Ascomycota
- Sub: Pezizomycotina
- Class: Saccharomycetes
- Order: Saccharomycetales
- Family: Saccharomycetaceae
- Genus: Saccharomyces
- Species: Saccharomyces cerevisiae (Walker, 2009).

Saccharomyces cerevisiae is classified as Ascomycetes, its ovum cell shape with a diameter of 5-10 micrometers (Walker, 2009).

This yeast can be cultivated easily, its generation time is short, it can multiply within 1.5-2 hours at 30 ° C, its production is fast and the maintenance of some specimens at low cost. Often used in industries such as beer, bread and fermented wine. The growth of yeast through the 4 phases is strongly influenced by the nutrients available in the medium that is the lag phase or also called the grace phase where bacteria still adapt to the surrounding environment, logarithmic phase (PK <50%), stationary phase (PK = 50%) and phase of death (PK> 50%) (Walker, 2009).

2.1.3 Morphological Characteristics

The characteristics of vegetative reproduction and vegetative cells can be used to classify yeast (De Becze, 1956). Vegetative reproduction is done either by fission or by budding or by formation of conidia. Reproduction by fission is a typical characteristic of Endomycetaceae and Schizosaccharomycoides (Figure 1). Genera Saccharomyces, Candida shows budding pattern (Figure 2). Reproduction by formation conidia is a characteristic of the genus Sterigmatomyces (Figure 3). Characteristics of vegetative cells are formation of pseudo and true mycelium. If yeast reproduces exclusively by budding it forms Pseudomycelium. Candida, Brettanomyces anomalus etc., show presence of Pseudomycelium. If yeast which reproduces solely by fission forms filamentous cells thus constitute true mycelium. Rhodosporidium, Leucosporidium etc. show true mycelium. The morphology of the vegetative cells grown in liquid and solid media is based on whether the cells are spherical, subglobus, ovoid, cylindrical etc. The apiculate cells of Nadsonia, the bottle-shaped cells of Pityrosporum, the triangular cells of Trignopsis are typical examples. Formation of chlamydosporem involves chlamydospore which has been defined as thick-walled, nondeciduos, intercalary or terminal, asexual spore formed by the rounding of a cell or cells. Chlamydospore formation is a characteristic feature of ascomycetous genus Metschnikowia and in cultures of genus Debaryomyces. Formation of endospores is the formation of vegetative cells delimited within cells or hyphae. This phenomenon is uncommon in yeast domain, but has been observed in genera Trichosporon, Cryptococcus, Syringospora and Oosporidium. Formation of ballistospores which are produced on sterigmata that protrude from vegetative cells and are ejaculated into the air by the drop mechanism.

Editorial note: This image was removed due to copyright issues.

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Figure 1: Budding Yeast.

Editorial note: This image was removed due to copyright issues.

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Figure 2: Fission in yeast.

Editorial note: This image was removed due to copyright issues.

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Figure 3: Candida formation in yeast.

2.1.4 Biochemical Characteristics

Yeast developing on a carbon source must either be able to ferment it or utilize it by respiration. It has been found that when yeast utilize carbon source fermentatively it is also able to utilize it oxidatively. But reverse does not occur. The ability or inability to ferment carbohydrates to ethanol and carbon dioxide is most important for differentiating species. Variety of sugars is fermented by a variety of yeast. Saccharomyces is known to show vigorous fermentation whereas genera like Rhodotorula and Lipomycesare strictly non-fermentative. Carbon assimilation tests are more sensitive than fermentative tests for detecting the presence of enzyme systems. Utilization of inositol is characteristic of genus Cryptococcus and inability to utilize lactose appears to be a feature of Saccharomyces. Splitting of arbutin is done to see ß-glucosidase activity in yeasts. If a yeast strain hydrolyzes arbutin, hydroxyquinone is formed, which gives a brown color with any soluble ferric salts incorporated in the medium (Reis, 2013).

Since nitrogen metabolism is a basic feature of growth, the ability or inability to utilize different sources of nitrogen can be made use of in classifying yeasts. Except for genus Saccharomyces which only grows in media containing certain yeast or protein hydrolysates all yeast can utilize a variety of nitrogen sources. The utilization of nitrate depends upon the sequential action of reductase enzymes which mediate reduction of nitrate to more reduced compounds. Potassium nitrate is generally used in medium as a nitrate source. Hansenula, Pachysolen and Citeromyces utilize nitrate. Species which utilize nitrate also utilize nitrite. In testing for assimilation, toxic effects due to nitrous acid maybe formed. So, testing of nitrite must be done at low concentrations. Sodium nitrite is generally used as a nitrite source. It has been found that aliphatic amine nitrogen can be utilized as a source of nitrogen. Kluyveromyces and Saccharomyces are known to utilize amini alkanes. Creatine is nitrogenous organic acid and creatinine is phosphorylated creatine. Utilization of creatine and creatinine by Cryptococcus neoformans and three species of genus Debaromyces is reported. Amino acids in general are a good source of nitrogen but they differ in their usefulness. L-amino acids (for example l-lysine) are utilized by Brettanomyces, Candida, Hansenula, Saccharomycodes, etc. (Reis, 2013).

The use of the ability or inability to grow in a synthetic medium devoid of vitamins like biotin, folic acid, niacin, inositol, riboflavin etc. was introduced by Wickerham. The value of the ability to grow in a vitamin free medium as a taxonomic criterion is variable. A variety of yeast species grow well in sugar concentration up to 40-70%. Saccharomyces, Dekkera can tolerate high sugar concentration. In general ability to grow above 37°C finds limited application. Certain species of genera Kluyveromyces and Hansenula are capable of growing at 45°C or even 48°C. Most yeast produce traces of volatile as well as non-volatile acids. It is seen when excess of acetic acid is produced. Some species of Kloeckera, Hansenula, Trichosporonare known to produce considerable amounts of acids. Under suitable cultural conditions several yeast strains elaborate extracellular polysaccharides. Bullera, Trichosporon produces certain amyloid compounds. Practically all yeast utilizes urea at low concentrations as a sole source of nitrogen provided that adequate amounts of vitamins are supplied. It is particularly marked in the genera Cryptococcus and Rhodotorula. Lipase activity has been demonstrated in several yeast species. It is usually employed as a confirmatory test. Candida lipolytica, C. rugosa and Trichosporon pullulans show lipase activity. The formation of distinctive pigments can be used for differentiation. Three main types of pigments maybe distinguished, carotenoids, pulcherrimin and rybosylaminoimidazole. Pigment is for instance highly characteristic of genera Rhodotorula and Sporobolomyces.

Fermentative yeasts form a variety of esters in varying amounts. Ethyl acetate has been found to be the commonest and most readily detectable ester formed by yeast. Hansenula and Kluyveromyces show strong ester formation. Yeast varies in sensitivity towards the antibiotic actidione. Saccharomyces cerevisiae is markedly sensitive (inhibited by 1 μg/ml), Schizosaccharomyces pombe is moderately sensitive (inhibited by 25 μg/ml), Kluyveromyces lactis is tolerant (not inhibited by conc. as high as 1000 μg/ml). It is accepted that the ability of yeast to liquefy gelatin is of very limited taxonomic value as very few yeasts are strongly proteolytic (Reis, 2013).



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