Streptomyces thermoviolaceus was grown in a chemostat under conditions of glutamate limitation. The effects of growth rate on production of the antibiotic granaticin, extracellular protein and protease activity as components of secondary metabolism were studied at 37,45 and 50 "C. The amount of each secondary metabolite synthesized was highly dependent on growth rate and temperature. Granaticin yields were highest at growth rates of 0.1 to 0.15 h-l at 37 "C, 0.175 h-l at 45 "C and 0.045 h-l at 50 "C. Protease activity of culture supernatants responded to low nutrient concentration and/or low growth rate. Measurements of extracellular protein revealed complex changes in amount which were dependent on growth rate and temperature. At 45 "C and a growth rate of 0.15 h-l, biomass yield was highest between pH 5.5 to 6.5 whereas granaticin synthesis was low at pH 5-5 and rose to highest values at between pH 6.5 and 7-5.

The synthesis of granaticin, a polyketide-derived antibiotic synthesized as a secondary metabolite by Streptomyces thermoviolaceus strain NCIB 10076, was studied at different growth temperatures. Quantitative measurements of the antibiotic made during batch fermentations showed that the yield was greatest at 45 "C, whereas the rate of synthesis was most rapid at 37 "C. The timing of the appearance of granaticin in culture could not be assigned to any particular phase of growth or to de-repression due to depletion of any particular nutrient. However, at all temperatures, appearance of the antibiotic coincided with a rise in ammoniacal nitrogen presumably due to deamination of glutamate, the carbon source for growth. We have previously shown that production of the antibiotic is pH sensitive and that some carbon sources result in higher titres than others. This paper examines the effect of temperature on the physiology of growth and on antibiotic production in more detail under conditions that also allow an exact measurement of granaticin yield. I N T R O D U C T I O NTaxonomic studies have revealed that many Streptomyces are able to grow at 45 "C (Williams et al., 1983). However until recently there were relatively few reports concerning the ability of species from this group to grow at temperatures in excess of 50 "C. Goodfellow et al. (1987) surveyed 50 thermophilic streptomycetes from a range of habitats which they were able to assign to a number of clusters in a numerical phenetic study that identified some features of their physiology and metabolic versatility. Such work is valuable in view of the commercial advantages of thermophilic species. These include faster reaction times and reduced cooling costs for large fermentations which make the thermophilic streptomycetes potentially useful vehicles for antibiotic production at high temperatures. Streptomyces thermoviolaceus is a particularly useful model system for such studies because it grows over a temperature range of around 25 to 58 "C (Edwards & Ball, 1987) and produces granaticin, a pigmented and pH sensitive antibiotic (Maehr et al., 1979) which is synthesized via the polyketide pathway (Arnone et al., 1979; Snipes et al., 1979). Snipes et al. (1979), using 13C-labelled acetate and glucose, showed that granaticin is produced by mixed biogenesis from eight acetate units and one dideoxyglucose moiety. This agrees with other radiolabelling work that showed polyketide antibiotics to be acetate derived (Floss et al., 1985). Using an lSO2 atmosphere, Floss et al. (1985) also demonstrated that no molecular oxygen was incorporated into the molecule, indicating a direct cyclization of the polyacetyl chain rather than a classical hydroxylation using molecular oxygen, followed by a dehydration to the lactone. METHODSOrganism and culture conditions. Streptomyces thermoviolaceus subsp. thermoviolaceus NCIB 10076 was maintained on complex solid medium that contained (g 1-l): glucose, 4; malt extract, 10; yeast extract, 4; and after adjusting to pH 7.2, agar, 20....

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Granaticin, an isochromate quinone antibiotic is synthesized as a secondary metabolite by Streptomyces thermoviolaceus. Antibiotic productivity was investigated under a variety of cultural conditions, including complex and defined media, mesophilic and thermophilic temperatures and a variety of sole carbon sources. In a defined medium growth was supported, to varying extents, by different carbon sources and in most cases granaticin production was observed. Highest biomass and granaticin yields were obtained when cultures were grown in the presence of xylan, fructose, glutamate or proline as carbon source. Changes in pH during growth affected both the timing and extent of granaticin production.

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