Heydarian, Seyed Mohammad; (1998) The influence of agitation on morphology, rheology and erythromycin production in Saccharopolyspora erythraea culture. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

In spite of the importance of filamentous actinomycetes, the number of published reports for engineering aspects of fermentation (e.g. effect of agitation) are rare compared with filamentous fungi. Intensive mechanical agitation is necessary for adequate oxygen transfer rate and bulk mixing. However, excessive physical damage due to high agitation on the filamentous mycelia during the fermentation is usually undesirable. This report investigates the effect of agitation, and in particular, shear on S. erythraea fermentation. Initially a relatively poor defined medium was used for S. erythraea fermentation at 7- L scale. It was shown that growth was inhibited at a low constant dissolved oxygen tension of 10% air saturation. However, the specific erythromycin production was virtually identical to that of a culture where DOT did not fall below 65%. In addition, at constant DOT (10%) a stirrer speed of 750 rpm caused serious mechanical damage to the mycelia compared with at 500 rpm. A bioassay was used for erythromycin measurements in this part of the study. A solid phase extraction method was developed for the purification and concentration of erythromycin for analysis by HPLC for future work. A complex medium was used to investigate the effects of shear on the S. erythraea fermentation at laboratory scale (7-L) and pilot scale (450-L) fermenters. Morphological measurements using image analysis showed that the major axis of the mycelia (both freely dispersed and clumps) decreased after the end of the rapid growth phase to a relatively constant value (equilibrium size) dependent on the stirrer speed. Variation of other morphological parameters are also discussed. A model of hyphal breakage was set up based on stress cycling (fatigue) resulting from continuous recirculation through the high shear zone of the impeller. The critical strain energy of failure was used, the mycelium was assumed to have fractal properties and the individual hypha was assumed to behave elastically with a thickness that increases linearly with the distance away from the growing tip. The comparison of the experimental data and the model showed reasonable agreement. The effect of shear on growth is discussed in terms of dry cell weight. Maximum dry cell weight was slightly lower at higher speeds. Maximum biomass was 11.1 ± 0.5 gL-1 at 1250 rpm (tip speed = 4.45 ms-1), whereas it was 12.7 ± 0.2 gL-1 at 350 rpm (tip speed =1.07 ms-1). Specific erythromycin production was not stirrer speed dependent in the range of 350 to 1000 rpm (tip speed = 3.56 ms-1) at 7-L scale, and it decreased by 10%, when the stirrer speed increased to 1250 rpm. The rheology of the culture changed dramatically during the fermentation. Power law model had a good agreement with the experimental data and the consistency index was generally lower at higher stirrer speeds. A correlation was recommended between the consistency index and biomass (Kalpha (DCW)2.1) and the relation between morphology and rheology is discussed. The mechanical properties of the cell wall of the S. erythraea were examined by disruption of fermentation broth in a homogeniser and it was shown that mechanical strength of the cell wall increased in a large extent during deceleration phase. These results were used to explain some aspect of mycelia breakage in the fermenter. This research has shown that shear affects morphology of the S. erythraea culture. These effects are generally less compared with filamentous fungi culture and are not detrimental to growth and erythromycin production, if enough nutrients are provided in the medium. A consequence of this is the ability to rise the agitation rate to increase oxygen transfer and bulk mixing without significant mechanical damage or loss of productivity.

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