Marcin Bizukojc

Physiological, morphological and kinetic aspects of lovastatin biosynthesis by Aspergillus terreus

This review focuses on selected aspects of lovastatin biosynthesis by Aspergillus terreus. Biochemical issues concerning this process are presented to introduce polyketide metabolites, in particular lovastatin. The formation of other than lovastatin polyketide metabolites by A. terreus is also shown, with special attention to (+)‐geodin and sulochrin. The core of this review discusses the physiology of A. terreus with regard to the influence of carbon and nitrogen sources, cultivation broth aeration and pH control strategies on fungal growth and product formation. Attention is paid to the supplementation of cultivation media with various compounds, namely vitamins, methionine, butyrolactone I. Next, the analysis of fungal morphology and differentiation of A. terreus mycelium in relation to both lovastatin and to (+)‐geodin formation is conferred. Finally, the kinetics of the process, in terms of associated metabolite formation with biomass growth is discussed in relation to published kinetic models. The review concludes with a list of the most important factors affecting lovastatin and (+)‐geodin biosynthesis.

Effect of pH on biosynthesis of lovastatin and other secondary metabolites by Aspergillus terreus ATCC 20542.

The effect of the initial pH value of the cultivation medium on lovastatin (mevinolinic acid) biosynthesis by Aspergillus terreus ATCC20542 was studied. It was found that if the pH value of the broth is acidic, the direct chromatographic assay of mevinolinic acid leads to the underestimated values. Thus, the equilibrium curve was determined for the transformation of β-hydroxy acid form of lovastatin (mevinolinic acid) into lovastatin lactone. The calculation of the equilibrium constant shows that when the pH value of the solution is 4.98, concentrations of both forms of lovastatin are equal to each other. This finding was next used to study mevinolinic acid formation at the various initial pH values of the medium. It occurs that even at pH lower than 5.5 mevinolinic acid is still, although inefficiently, produced and its presence remains unnoticed, unless the samples of the broth are alkalised prior to the assay. Mevinolinic acid is efficiently produced at the initial pH value of the medium equal to 7.5 and 8.5 and it correlates with the rapid utilisation of lactose by A. terreus. Additionally, other secondary metabolites were sought at the various initial pH values of the medium with the use of mass spectrometry. (+)-Geodin is only formed at pH 6.5, while monacolin L is found at the highest amount at pH 7.5.

Lovastatin biosynthesis by Aspergillus terreus with the simultaneous use of lactose and glycerol in a discontinuous fed-batch culture

The influence of various combinations of glycerol and lactose feed on the biosynthesis of two polyketide metabolites, lovastatin and (+)-geodin, by Aspergillus terreus ATCC20542 in a discontinuous fed-batch culture was presented. In these experiments lactose and/or glycerol were also used as the initial carbon substrates in the cultivation media. The application of glycerol feed, when lactose is the initial substrate, leads to the appreciable lovastatin concentration in the broth (122.4 mg l⁻¹), nevertheless the abundant (+)-geodin level is at the same time obtained (255.5 mg l⁻¹). The cultures with glycerol as the initial substrate and fed with lactose produce less lovastatin and (+)-geodin. The application of the various combined glycerol and/or lactose feeds allows for improving lovastatin production up to 161.8 mg l⁻¹ and decreases (+)-geodin concentration to 98.7 mg l⁻¹. The analysis of product formation rates and yield coefficients indicates that lovastatin is more efficiently produced on lactose, especially in the initial stages of the cultivation. Glycerol efficiently sustains fungal activity to form these polyketides in the late idiophase but it mainly favours (+)-geodin formation, if solely used in the feed. The feeds performed both with lactose and glycerol occur to be the most desired to maximise lovastatin and minimise (+)-geodin formation.

Kinetics of the aerobic biological degradation of shredded municipal solid waste in liquid phase

The organic fraction of municipal solid waste (OFMSW) should be utilised by means of biological methods. The biodegradation of solid wastes can be intensified owing to application of the bioreactors. Estimation of the optimum values of the organic load is one of the most important tasks for the aerobic biodegradation processes. The kinetic model of biological oxidation of the organic wastes has been presented in this paper. The experiments were carried out in batch 6-l working volume stirred tank bioreactors at constant temperature of 25 degrees C. Initial total solids have been at the levels of 15, 19, 34, 55 and 66 g l(-1). The kinetics of microbial decomposition of organic substances was described by means of an unstructured model. The satisfactory time courses for substrate chemical oxygen demand in the solid (CODs) and liquid phase (CODL) and biomass concentration (RNA) have been achieved. Also, the influence of the initial TS on the kinetics of the biodegradation process was investigated and the optimum value of initial TS for this type of processes was estimated at 34-55 g l(-1).

Lovastatin and (+)-geodin formation by Aspergillus terreus ATCC 20542 in a batch culture with the simultaneous use of lactose and glycerol as carbon sources

The influence of initial glycerol and lactose concentrations on lovastatin and (+)‐geodin formation in batch cultures of Aspergillus terreus ATCC 20542 was presented. At first the experiments comprised lovastatin biosynthesis on glycerol as the sole carbon source. Lovastatin titers below 40 mg/L were found under these conditions and they were lower than previously obtained results when lactose was used as the sole carbon source. However, the application of the mixture of glycerol and lactose allowed in achieving higher lovastatin concentration in the broth. It even exceeded 122 mg/L when 10 g lactose and 15 g glycerol per liter were used. The calculated lovastatin volumetric and specific formation rates on glycerol or lactose and on the mixture of these two showed that lovastatin was faster produced on lactose than on glycerol. In the trophophase, the maximum volumetric lovastatin formation rate on lactose was up to four times higher than on glycerol and so was the lovastatin specific formation rate. Similar relations for the accompanying (+)‐geodin biosynthesis were also studied. When the mixture of lactose and glycerol was used, the transformation of (+)‐geodin to other polyketide metabolites also took place.

Morphologically structured model for growth and citric acid accumulation by Aspergillus niger

A morphologically structured model for the batch process of biomass growth and citric acid accumulation by Aspergillus niger is presented in this paper. The model consists of ten ordinary differential equations, which balance biomass and four physiological zones of hyphae, and includes the most important medium components, such as carbon sources, nitrogen source and citric acid. Digital analysis of microscopic images was employed to determine the hyphal fractions within each physiological zone. All model parameters were determined on the basis of data obtained in four experiments carried out in two types of bioreactors and under various process conditions, specifically normal and increased pressure. Some parameters were directly calculated from the experimental data by means of linear and non-linear regression. This allowed minimisation of the number of parameters to be found by means of an optimisation procedure, down to 11 and thus, significantly reduced computation time. The simulated curves are largely consistent with the experimental data and some parameters appeared to be practically independent of process conditions. These findings contribute to the universality of the model proposed.

A new approach to determine the kinetic parameters for nitrifying microorganisms in the activated sludge systems

This work aims at establishing the methodology to determine kinetic parameters describing growth of autotrophs in the activated sludge systems. It is based upon the measurement of oxygen uptake rate (OUR) only. Two Monod kinetic parameters, namely maximum specific growth rate for autotrophic biomass (μ(max,A)) and half-saturation constant for ammonium ions (K(A)) were simply and rapidly determined within a single batch OUR test with the use of ammonified wastewater. The obtained mean values of the parameters are μ(max,A)=0.675 h(-1) and K(A)=0.912 mg NH(3)-Nl(-1) and they are close to the ones assumed in activated sludge models. It indicates that the methodology of the respiratory batch test elaborated in this study is reliable.