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PPT 206 Instrumentation, Measurement and Control SEM 2 (2012/2013). Microcalorimetric Characterization of Bacterial Inocula. Dr. Hayder Kh. Q. Ali hayderali@unimap.edu.my.
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PPT 206 Instrumentation, Measurement and Control SEM 2 (2012/2013) Microcalorimetric Characterization of Bacterial Inocula Dr. Hayder Kh. Q. Ali hayderali@unimap.edu.my
Microcalorimetric output is a power-time curve; the area under the curve measures the heat production of the culture.
Catabolic and Anabolic Reactions The sum of all chemical reactions within a living organism is known as metabolism. Metabolic pathways are a series of chemical reactions in which the product of one reaction is the substrate for the next reaction. The chemical reactions are catalyzed by enzymes, so the metabolic pathways present in an organism depend on the enzymes present in the organism. The enzymes present in an organism depend on the genes present in the organism.
Catabolism refers to chemical reactions that result in the breakdown of more complex organic molecules into simpler substances. • Catabolic reactions usually release energy that is used to drive chemical reactions. • Anabolism refers to chemical reactions in which simpler substances are combined to form more complex molecules. • Anabolic reactions usually require energy.
Anabolic reactions build new molecules and/or store energy. • The energy of catabolic reactions is used to drive anabolic reactions. • The energy for chemical reactions is stored in ATP.
Microbiological studies involving calorimetry are not of recent origin, although the present availability of commercial, sensitive microcalorimeters has given additional impetus to such studies. It has been established that calorimetric studies perceive only catabolic reactions and that observation of anabolic processes would require a much improved sensitivity.
Thus the major thermal events observed in the calorimetric study of microbial growth are attributed solely to catabolic activity. That the pattern of these thermal events as a function of time could be used for identification of microorganisms was implied by many researchers. Recent studies using more sensitive microcalorimeters have led to claims that such identification is possible.
It is claimed that features of the thermogram can be attributed to particular metabolic events for some 200 clinically significant bacteria. The studies reported to date have been in unstirred batch-type calorimeters without any control of the atmosphere above the incubation. It is a requirement of any microbial identification procedure that the immediate history of the organism should have no effect on the subsequent identification.
The causal relationship between specific metabolic events and details in the thermograms must be established by classical analytical procedures. The purely physical events which can occur (e.g. settling of cells, establishment of pH gradients, etc.) particularly in batch calorimetric procedures must be shown to have no resultant thermal effects.
The introduction of flow microcalorimeters presents a particularly simple and revealing means of studying metabolism in microbes. Furthermore from flow microcalorimetric experiments on the growth of the micro-organism, the possibility of identification under restricted conditions could be carried out. Indeed it may be possible to "characterise" strains of the micro-organism for their utility in specified industrial processes on the basis of the thermogram derived from flow microcalorimetric observation of growth.
Procedure of Experiment A flow microcalorimeter was operated at 310 K in a room maintained at 298 + 0.5 K. Incubations were performed in a fermentor vessel of approximately 200 ml capacity. The vessel was sealed by a neoprene ring and a threaded head-plate which contained 6 ports. These were used for inlet and outlet tubes to and from the calorimeter; thermometer; an oxygen electrode or addition port; and a port for a reflux condenser. The vessel was lagged and thermostatted on a magnetic stirring unit.
Microorgansimwas inoculated into the medium (100 ml) contained in the fermentor using a microlitre syringe at zero time, and the inlet tube to the calorimeter inserted 1 min later. After a further 3 rain the first response was detected in the calorimeter and the integration of the area under the thermogram initiated. Ten minutes after inoculation the effluent from the calorimeter was recycled into the fermentor.
