eChapters

Introduction

Urea is one of the common fertilizers used as a source of nitrogen. It has nitrogen in amide form (-NH₂) and this form of nitrogen is not utilized by plants. So it has to be hydrolysed. The enzyme, urease hydrolyses the urea to ammonia and ammoniacal nitrogen is taken up by plants or further nitrified.

The measurement of activity of this enzyme in soil is important from soil fertility point of view. There are two methods for determination of the urease activity of soils.

• First involves the measurement of end products of urea hydrolysis. i.e., NH₃ or CO₂.

Introduction

All biological reactions in soil are catalyzed by enzymes. Soil enzyme activities are believed to indicate the extent of specific processes in soil and in some cases, can act as indicators of soil fertility. Biological oxidation of organic compounds is generally a dehydrogenation process and there are many dehydrogenases that carry out the following reaction:

XH₂ is an organic compound and is a hydrogen donor and A is a hydrogen acceptor. These systems are integral part of the microorganisms, so dehydrogenase activity can be correlated with active population.

Principle

Measurement of dehydrogenase activity involves determination of Triphenyl formazan (TPF) produced by the reduction of Triphenyl tetrazolium chloride (TTC). TTC is a redox indicator, a yellow coloured water soluble salt which possesses the property of being easily transformed into, methanol soluble formazan (intensely coloured, water insoluble) by reduction. 

Introduction

The enzyme nitrogenase is produced in all N₂-fixing systems. The activity of nitrogenase enzyme relates to the biological nitrogen fixation, which can be determined directly by measurement of fixation product, ammonia and the further products of its assimilation. The activity can also be measured indirectly by measuring the ability of nitrogenase to reduce the number of substrates other than N₂. Substrates such as nitrous oxide (N₂O), azide ion (N₃^- ), cyanide (CN^-) and isocyanide (NC^-) are reduced and the requirements for ATP and reductant are similar to those of N₂ fixation. The reduction of protons to molecular hydrogen has been used to measure the nitrogenase activity.

Dilworth (1966) and Schollhorn and Burris (1966&1967) observed the inhibitory effect of Acetylene (C₂H₂) on N₂ fixation by extracts of Clostridium pasteurianum. Dilworth (1966) further observed by mass spectrometry that the C₂H₂ was reduced to C₂H₄. Both gases are readily measured at very low concentrations by gas chromatographic techniques.

Introduction

The major biogenic gases, which contribute to global warming, are carbon dioxide, methane and nitrous oxide. Out of these, methane is the most important gas because its thermal absorption is 30 times greater than carbon dioxide, though its atmospheric concentration is about 200 times less than carbon dioxide. Atmospheric measurements of greenhouse gases indicate that methane concentration had increased by 1 per cent annually to 1.7 ppm (by volume) during the last decade. The steady increase of atmospheric methane concentration in the past 200 years has attracted much attention.

Flooded rice soils provide an optimum environment for methane production and emission. Flooding a soil creates essential redox potential, anaerobic decomposition of organic matter and stabilizes the soil pH near neutral, thus makes the congenial environment for methane emission.

Rice is the world’s most important food crop

Introduction

The soil microbial biomass is an important component of the soil organic matter that regulates the transformation and storage of nutrients. It is a labile component of the soil organic fraction containing 1 to 3% of the soil carbon and up to 5% of the total soil N. The microbiologically mediated processes affect ecosystem functions associated with nutrient cycling, soil fertility, global C change and soil organic matter turnover. The size and activity of the soil microbial biomass must be assessed to fully understand nutrient flux in managed and natural ecosystems.

Soil microbial biomass estimations are useful in investigations that compare temporal nutrient fluctuations along natural and perturbed gradients. The effect of tillage, crop rotations and soil type on organic carbon and nutrient turnover can be assessed by following nutrient pools and activity associated with the soil microbial biomass.

Introduction

The balance between soil organic matter (SOM) levels and nutrient cycling are the key to sustained productivity of agricultural system. Both are closely related through microbially driven mobilisation/immobilisation processes. Three major models have been recognised for C pool viz. Labile C, non-labile C and recalcitrant. The models have been characterised on the bases of their half-life period. The labile C includes oxidisable C (with KMnO4), microbial biomass C (soil carbohydrates) and soil fauna exudates (polysaccharides). Its half-life period is from a few hours to a few days. The non-labile C includes aggregate C and its turn over time is from a few months to a few years while the recalcitrant (stabilised C) includes humic fractions and charcoal C having the turn over time of hundred years. Thus fractionation of SOM is carried out on the bases of humic substances, dissolved organic C, natural C13 abundance, microbial biomass C and ease of oxidation of C. The latter method provides qualitative characterisation of soil C.

Introduction

Organic matter in sediment consists of carbon and nutrients in the form of carbohydrates, proteins, fats and nucleic acids. Bacteria quickly eat the less resistant molecules, such as nucleic acids and many of the proteins. Sediment organic matter is derived from plant/animal detritus, bacteria or plankton formed in situ. Total organic carbon refers to the amount of organic matter preserved within sediment.

Materials

1. Sample
2. Crucibles
3. Furnace

Procedure

Take 1.0 g of sample in a crucible whose empty weight is already recorded. Place this crucible in a furnace. Switch on the furnace and set the temperature at 550°C. Keep the samples for 3 hours

Introduction

Sugars are commonly found in the form of biopolymers. Cellulose, a biopolymer of glucose is the predominant constituent of plant tissues, and may be shortly described as the structural bases of vegetable world. In this case, the glucose moieties are strung together by P-glycosidase linkages, which form the linear chain, making it resistant to chemical attack. Hemicellulose consists of short, highly branched, chains of sugars. It contains five carbon sugars (usually D-Xylose and L-Arabinose) and six carbon sugars (D-Galactose, D-Glucose and D-Mannose) and uronic acid. Its branched nature renders hemicellulose amorphous and relatively easy to hydrolyse to its constituent sugars. Lignin is a biopolymer rich in phenolic components, which provide structural integrity to plants. Acid and neutral detergent fraction methods (ADF and NDF) involve the gravimetric determination of residue previously treated with acid and neutral detergent solutions. These methods determine the insoluble fraction and their individual components. NDF represents the total plant fibre or cell wall including hemicellulose

Introduction

In recent years, the boom in industrialization has revolutionised the countries but has also added much of sufferings. There is tremendous increase in the water and air pollution. Large amounts of effluents from industries containing organic substances of different nature are discharged and enter the aquatic systems resulting in water pollution. Biological Oxygen Demand (BOD) is the amount of dissolved oxygen required by the aerobic microflora to oxidise the organic matter of dirty water. Increased organic matter results in more oxygen requirements by microorganisms to oxidise it and it results in enhancement of BOD of water. As the BOD of water rises, due to decrease in dissolved oxygen (DO), there is increase in death of fish due to suffocation. Determination of biological oxygen demand (BOD) only is insufficient to give a clear picture of organic content of water bodies and the presence of toxic chemicals in the sample so the chemical oxygen

Introduction

Bacteriological examination of milk indicates the hygienic condition of the milk i.e., quality of milk in relation to the health of consumer. Two main objectives of carrying out bacteriological examination are to evaluate the degree of cleanliness in the vicinity of production and handling of milk; and secondly to check the presence of infectious microorganism. Generally, number of bacteria present per milliliter is taken as an index in the routine examination of milk as other tests for identification and differentiation of pathogenic and non-pathogenic microorganisms are time consuming and expensive.

Direct Microscopic Count (DMC) consists of examining stained films of measured volume of milk or milk product, dried on a glass slide under a compound microscope. This method enables rapid estimation of total population of bacteria of a milk sample with relatively more accuracy as compared to other methods.

Introduction

Standard Plate Count (SPC) is used for determination of population of viable microbes. Enumeration procedures are based on the assumption that microbial cells present in a sample mixed with an agar medium form visible separated colonies. The bacterial colonies developed on the plates are counted. Each colony is presumed to have developed from one bacterium or clump of bacterial cells. Therefore, the results are expressed in colony counts per /ml of sample or colony forming units per /ml (cfu /ml). Total number of colonies counted on the plate multiplied by the dilution factor represents the number of viable organisms in a sample. SPC is a useful method for evaluating the microbiological quality at various stages of processing. It is also helpful in detecting the source of contamination in the product.

Introduction

Dye reduction tests show the comparative activity of the bacteria in milk, therefore are considered as rough indicator of the number of bacteria per millilitre of milk. These tests find wide application in dairy industry especially in grading of milk for manufacturing purposes. The small amount of equipment, materials and simplicity of method permit simultaneous testing of several samples. Since it combines speed and adaptability, this test may be used in milk plants, receiving stations, condensing plants, cheese factories and similar establishments to rapidly grade incoming milk.

Principle

It is based on the observation that the colour imparted to milk by a small amount of methylene blue disappears in a time, the length of which is dependant largely upon the number of microorganisms

J    :    Joule.

J    :    Joining segment.

J chain      :    A polypeptide present in polymeric IgM and IgA that links the subunits together.
    
J gene segments, joining gene segments     :    The J gene segments, or joining gene segments, are found some distance 5' to the C genes in immunoglobulin and T-cell receptor loci. A V and in the case of IgH chains, tcrâ chains and tcrä chains, a D gene segment must also rearrange to a J gene segment to form a complete V-region exon.

Introduction
Oil palm (Elaeis guineensis Jacq.), a tropical perennial tree native to West Africa, is the most productive oilseed crop for biodiesel, yielding up to 5 tons of oil per hectare (Mekhilef et al., 2011). Indonesia and Malaysia are the largest global producers, accounting for over 85% of the world’s palm oil (Atabani et al., 2012). Palm oil is extracted from the fruit of the oil palm and is widely used in both food and fuel applications. However, its cultivation has raised environmental concerns, including deforestation, habitat loss, and carbon emissions (Gui et al., 2008). Efforts are being made to promote sustainable palm oil production to mitigate these impacts (Mekhilef et al., 2011). Deforestation, habitat loss, and carbon emissions are only a few of the serious environmental issues brought on by its cultivation (Fargione et al., 2008). Oil palm plantation growth has been connected to a decline in biodiversity, especially in tropical rainforests that are home to endangered species like Sumatran tigers and orangutans (Fitzherbert et al., 2008). According to Carlson et al. (2013), the conversion of peatlands for the production of oil palm contributes to climate change by increasing greenhouse gas emissions.
History of Oil Palm as Biodiesel Feedstock
Oil palm has been used for cooking and lighting for thousands of years in Africa and Southeast Asia (Obahiagbon, 2012). Research into its use as a biodiesel feedstock began in the 1980s, with Malaysia pioneering biodiesel development (Zainal et al.,2017).
• 1982: The Palm Oil Research Institute of Malaysia (PORIM) successfully tested palm oil-based biodiesel in diesel engines (Yusoff, 2006).
• Early 2000s: Commercial production expanded in Indonesia and Malaysia, driven by government policies and renewable fuel demand (Mekhilef, 2011).
• Sustainability challenges have led to the establishment of certification schemes such as the Roundtable on Sustainable Palm Oil (RSPO) (Ruysschaert & Salles, 2014).

Microbiology

Study of microorganisms, a large and diverse group of microscopic organisms, that exist as single cell or cluster. It also includes virus, which are microscopic, but not cellular.

The Laboratory Worker is Surrounded by Many Dangers such as from..........

Handling of infectious materials.

Handling of broken glassware.

Accidental spill of corrosive reagents.

Swallowing of corrosive reagents such as concentrated Sulphuric acid,

Hydrochloric acid, sodium hydroxide, trichloroacetic acid etc.

Swallowing of infectious specimen.

Inhalation of poisonous fumes.

Infection in the conduct of necropsies.

Potential hazards in the form of inflammable chemicals and gas leakages.

1. Bacterial (Prokaryote) Genome

The chromosome of most bacteria is a single super coiled double stranded circular DNA molecule. A complex packaging mechanism is necessary in order to ensure that the entire DNA is folded within the bacterial cell in a manner, which will not inhibit transcription nor allow entanglement of the two daughter strands to occur during replication (Highly condensed >10 mg/ml in region less than l ìm in diameter). This is achieved by two main mechanisms.

• First the DNA is folded into between 40-100 loops
• Second each of the quasicircles is it self super coiled independently of the others each loop are abolished by Ribonuclease.

Deoxyribonucleic remove the super helical nature of the loops, though limited treatment with this enzyme will attack only a small number of the loops and therefore leads to removal of only some of the super coiling. The intact folded chromosome or nucleoid can be isolated in the presence of non ionic detergents and high molarities of salt in two possible forms free and membrane associated. The free form which sediments at 1600-1700s consists of about 60%DNA, 30%RNA and 10%protein the bulk of which is the enzyme RNA polymerase. The membrane bound form contains an additional 20% of membrane-attached protein. Bacterial genome size ranges from 580 to 9900kbp, & the average gene size is constant from 900 to 1100bp of which 87-94% are protein coding (Rickettisa’s 76% and Borelia Plasmid’s 41% are exceptions).

Introduction

The primary tools used by the molecular biologists in manipulating DNA are restriction enzymes and other DNA/RNA modifying enzymes. Over the years, the number and uses of these enzymes have increased as new molecules have been discovered. Restriction endonucleases are bacterial enzymes that cleave double-stranded DNA. They are present in bacteria presumably to destroy DNA from foreign sources. By cleaving the foreign DNA at specific sites. The host bacterial DNA os protected from cleavage because specific recognition sites are modified, usually by methylation form methylase at one of the bases in the site, making the site no longer a substrate for RE cleavage. Host bacteria used to propagate cloned DNA in the laboratory arte usually mutant in the host restriction genes, thus their intracellular enzyme activities will not destroy the foreign recombinant sequences. The endonuclaase with its accompanying methylase is called a restriction modification (R-M) system.

At least four different kinds of R-M systems exist characterized on the basis of the subunit composition, cofactor requirements and type of DNA cleavage. Most characterized enzymes belong to type II class, together with type IIS class. They comprise the commercially available restriction enzymes used for DNA analysis and manipulation. Type I and Type III enzymes are relatively uncommon and a few additional enzymes fit none of the classes.

Among more than 3000 different enzymes isolated from bacterial strains many share common specificities. Restriction enzymes that recognize identical sequences have been called as isoschizomers.

RAPD stands for Random Amplification of Polymorphic DNA. It is a type of PCR reaction, but the segments of DNA that are amplified are random. The scientist performing RAPD creates several arbitrary, short primers (8-12 nucleotides), then proceeds with the PCR using a large template of genomic DNA, hoping that fragments will amplify. By resolving the resulting patterns, a semi-unique profile can be gleaned from a RAPD reaction.

No knowledge of the DNA sequence for the targeted gene is required, as the primers will bind somewhere in the sequence, but it is not certain exactly where. This makes the method popular for comparing the DNA of biological systems that have not had the attention of the scientific community, or in a system in which relatively few DNA sequences are compared (it is not suitable for forming a DNA databank). Because it relies on a large, intact DNA template sequence, it has some limitations in the use of degraded DNA samples. Its resolving power is much lower than targeted, species specific DNA comparison methods, such as short tandem repeats. In recent years, RAPD is used to characterize, and trace, the phylogeny of diverse plant and animal species.