Zhang Cheng-wu

Accurate quantification of astaxanthin from Haematococcus crude extract spectrophotometrically

The influence of alkali on astaxanthin and the optimal working wave length for measurement of astaxanthin from Haematococcus crude extract were investigated, and a spectrophotometric method for precise quantification of the astaxanthin based on the method of Boussiba et al. was established. According to Boussiba’s method, alkali treatment destroys chlorophyll. However, we found that: 1) carotenoid content declined for about 25% in Haematococcus fresh cysts and up to 30% in dry powder of Haematococcus broken cysts after alkali treatment; and 2) dimethyl sulfoxide (DMSO)-extracted chlorophyll of green Haematococcus bares little absorption at 520–550 nm. Interestingly, a good linear relationship existed between absorbance at 530 nm and astaxanthin content, while an unknown interference at 540–550 nm was detected in our study. Therefore, with 530 nm as working wavelength, the alkali treatment to destroy chlorophyll was not necessary and the influence of chlorophyll, other carotenoids, and the unknown interference could be avoided. The astaxanthin contents of two samples were measured at 492 nm and 530 nm; the measured values at 530 nm were 2.617 g/100 g and 1.811 g/100 g. When compared with the measured values at 492 nm, the measured values at 530 nm decreased by 6.93% and 11.96%, respectively. The measured values at 530 nm are closer to the true astaxanthin contents in the samples. The data show that 530 nm is the most suitable wave length for spectrophotometric determination to the astaxanthin in Haematococcus crude extract.

Effect of lighting, temperature and pH on photosynthetic characters of Haematococcus pluvialis CG-11.

The effects of light intensity, temperature and pH on photosynthetic characters of Haematococcus pluvialis CG-11 were studied by measuring photosynthetic O2 evolution and fluorescence kinetics. The results showed that the saturation light intensity was 109.1 μmol/m2·s of H. pluvialis CG-11, with the maximum photosynthetic oxygen evolution rate of 75.9 μmol O2/mg Chla·h; the range of suitable growth temperature was between 25℃ to 30℃, the maximum photosynthetic rate was obtained at 25℃; the photosynthetic efficiency kept quite high at the pH range of 7.5—8.0,with the maximum photosynthetic oxygen evolution rate of 75.5 μmol O2/mg Chla·h at pH value of 7.5. Under experimental conditions of pH, a similar tendency manifested in the chlorophyll fluorescence kinetics parameters for H. pluvialis, at the pH range of 7.5—8.0, the Fv/ Fm, Fv/ F0, ΦPSⅡand ETR increased following the rising of pH, with their maximum values at pH 7.5, Fv/ Fm and Fv/ F0 decreased significantly over pH 7.5, while the decreasing trend didn’t change obviously in ΦPSⅡ and ETR.

Effects of carbon sources on growth and fatty acid composition of Pinguiococcus pyrenoidosus CCMP 2078.

Microalgae are potential sources of polyunsaturated fatty acids(PUFAs), mainly EPA(Eicosapentaenoic acid)and DHA(Docosahexaenoic acid). However, the EPA productivity of microalgae is lower compared with bacteria and fungi, and the reason is mainly the low growth rate and biomass under photoautotrophic conditions. To promote the EPA productivity, culture conditions such as culture temperature, irradiance, pH, nutrition sources and concentrations have been investigated. Nevertheless, some microalgae grow rapidly and have a higher biomass under mixotrophic conditions than which under photoautotrophic conditions. Pinguiococcus pyrenoidosus belongs to Pinguiophyceae, Pinguiococcus genara. The percentage of rude lipid content of the microalgae is 27.45%, among which EPA and saturated fatty acids (14∶0, 16∶0) are high, but the contents of other kinds of fatty acids are very low. So Pinguiococcus pyrenoidosus is a good material for producing PUFAs and saturated fatty acids. So far, there is little information about the mixotrophic growth on the growth rate and fatty acid composition of the microalgae. In order to determine the optimal culture conditions, EPA productivity and saturated fatty acids productivity, effects of different carbon resources (inorganic carbon compounds and organic carbon compounds) on the growth and fatty acid composition in P. pyrenoidosus CCMP 2078 were investigated. The experimental method included the following three parts: First, three different concentration levels of CO2 were set, namely 0.5%, 1.0% and 1.5%; Second, four different concentration levels of NaHCO3 were set, namely 5, 10, 20 and 25mmol/L; Third, four different concentration levels of glucose were set, namely 5, 10, 20 and 40g/L, respectively. Parameters of cell density and fatty acid composition were measured. The results showed that (1) Growth of P. pyrenoidosus CCMP 2078 was promoted by adding proper concentrations of carbon sources. The optimal concentrations of carbon sources for growth were 0.5% CO2, 5mmol/L NaHCO3 and 20g/L glucose, and the cell density of late logarithmic phage were 3.10-fold, 1.47-fold and 2.78-fold of that obtained under control; (2) When added the carbon sources except the low concentration of glucose, the percentages of TPUFA(Total polyunsaturated fatty acids)and EPA, and cell EPA and TSFA(Total saturated fatty acids)content were all lowered, but the percentage of TSFA was increased; (3) The yield of SFAs(Saturated fatty acids)and EPA were enhanced when added low concentrations of carbon sources. The yield of EPA and TSFA were 2.30-fold and 2.69-fold when added 0.5% CO2 of those obtained under control, respectively. The yield of TSFA was 1.85-fold when added 5mmol/L NaHCO3 of that obtained under control. The maximum yield of EPA and TSFA were 2.11-fold and 1.58-fold when added 5g/L and 10g/L glucose of those obtained under control, respectively. Therefore, low concentration of CO2 was the optimal culture condition for the growth of P. pyrenoidosus CCMP 2078, and also for the yield of EPA and SFAs. The increases of EPA and TSFA were resulted by the enhancement of the biomass concentration.