C-phycocyanin Production Research Articles

Production, isolation and characterization of C-phycocyanin from a new halo-tolerant Cyanobacterium aponinum using seawater

A halo-tolerant Cyanobacterium aponinum PCC 10605 was applied for the first time to produce high-level C-phycocyanin (C-PC). Combined with chemical extraction with sodium phosphate buffer and physical treatment using high pressure homogenization, a higher titer of C-PC was achieved. The culture conditions were optimized by mixing nitrate and ammonia ions, 2% carbon dioxide, and conditional light intensity. Thus, strain PCC10605 produced the highest titer C-PC of 0.652 g/g-DCW in the N1A2 medium with 10% light intensity and 16:8 light-period on day 7. PCC10605 accumulated 0.51 g-CPC/g-DCW at 20 g/L NaCl, while it grew normally in seawater with 30 g/L salinity, thus confirmed that PCC10605 was halo-tolerant strain. Besides, PCC10605 survived in 0.12 g/L phosphate medium that has never been reported. Finally, the purified C-PC exhibited DPPH, superoxide scavenging activity and antibacterial activity, which displayed 87.6%, and 18.7% removal of free radical, and 1.98 cm of inhibition zone for Escherichia coli.

Media optimization for C-phycocyanin production in Plectonema sp. using response surface methodology and central composite design

Cyanobacteria represent the richest sources of phycobiliproteins with especial reference to C-phycocyanin (C-PC), which in turn holds exhaustive therapeutic implications. Screening of several cyanobacterial strains namely Anabaena sp., Nostoc muscorum, Cylindrospermum sp., Plectonema sp., Scytonema sp., Spirulina sp., Synechococcous sp. and Tolypothrix sp. was carried out for their C-PC producing capacity, however the produced quantity of C-PC varies greatly among different strains. Owing to the crucial role of different media constituents on productivity of C-PC the current study was designed to optimize most appropriate media composition for augmented CPC production by selected superior producer. 36 factorial central composite design (CCD) dependent response surface methodology (RSM) was utilized to estimate the important medium components attributed with influencing C-PC productivity. RSM analysis of five independent coded factors including Na2CO3, K2HPO4, NaNO3, citric acid and EDTA were analyzed preceded by recognition of efficient variables for algal components production by Plectonema sp. Investigation of results revealed that the eminent medium components were NaCO3 (0.4 g/L); NaNO3 (0.5 g/L); K2HPO4 (2.8 g/L); citric acid (0.08 g/L) and EDTA (0.01 g/L) respectively. The optimized combination yielded 0.5536 mg/ml of C-PC. The increment of C-PC yield is R-Sq = 88.2%. Thus, our study led to the recognition of critical nutritional component that can be used further for enhanced productivity of C-PC.

Impact of incubation conditions on protein and C-Phycocyanin recovery from Arthrospira platensis post- pulsed electric field treatment

Pulsed electric field (PEF) was conducted for the extraction of proteins/C-Phycocyanins from Arthrospira platensis. The cyanobacterial suspension was treated with 1 µs long pulses at an electric field strength of 40 kV·cm−1 and a treatment energy of 114 kJ·kgsus−1 and 56 kJ·kgsus−1. For benchmarking, additional biomass was processed by high pressure homogenization. Homogeneity of the suspension prior to PEF-treatment influenced the protein/C-phycocyanin extraction efficiency. Stability of C-Phycocyanin during post-PEF incubation time was affected by incubation temperature and pH of the external medium. Biomass concentration severely affect proteins/C-Phycocyanins extraction yield via PEF-treatment. The optimum conditions for extraction of proteins/C-Phycocyanin was obtained at 23 °C while incubating in pH 8-buffer. The energy demand for PEF-treatment amounts to 0.56 MJ·kgdw−1 when processing biomass at 100 gdw·kgsus−1. PEF treatment enhances the protein/C-Phycocyanin extraction yield, thus, it can be suggested as preferential downstream processing method for the production of C-Phycocyanin from A. platensis biomass.

C-phycocyanin extraction from two freshwater cyanobacteria by freeze thaw and pulsed electric field techniques to improve extraction efficiency and purity

C-phycocyanin was extracted from Nostoc commune TUBT05 and Oscillatoria okeni TISTR8549 using freezing and thawing and pulsed electric field treatments. The results confirmed that freezing and thawing can be used to extract c-phycocyanin from both cyanobacterial strains, whereas pulsed electric field treatment can be used only with N. commune, whose cell structure allows extraction by this technology. The number of freeze/thaw cycles, the extraction solution used and the number of electric pulses had statistically significant influences on c-phycocyanin concentration, purity, yield and the total protein content in the crude extracts (p ≤ 0.005). Exposure of N. commune to three cycles of freeze/thaw in sodium-phosphate buffer yielded the highest c-phycocyanin content, purity, and yield. The results suggest that careful selection of both the number of freeze/thaw cycles and the extraction solution is required to ensure high productivity. Transmission electron microscopy micrographs confirmed that the cell wall structure and organization of the thylakoid were different in the two species, requiring a significantly different number of freeze/thaw cycles and pulse applications for optimal c-phycocyanin extraction. Both cyanobacterial strains were shown to be potential alternative strains for c-phycocyanin production. However, further work is needed to improve c-phycocyanin concentration and purity. We suggest that this will require refinement of the cultivation conditions and improved purification processes.

Sustainable phycocyanin production from Arthrospira platensis using solar-control thin film coated photobioreactor

Solar irradiance consists of photosynthetically-active photons that can be transformed to valuable biomolecules by microalgae. Light also has undesirable non-photosynthetic photons, such as ultraviolet and infrared wavelengths that heat up algal closed photobioreactors above optimum temperatures for growth. In this study, a solar control infrared blocking film (IRF) is applied to an algal flat plate photobioreactor to block excessive non-photosynthetic photons and regulate the temperature profile of Arthrospira platensis cultures for the production of C-phycocyanin (C-PC). The performance of the IRF is compared against other cooling mechanisms such as insulated-glazed photovoltaic (IGP), conventional water-jacket (CWJ) and a no heat control (NHC) photobioreactors. Experimental results show that the maximum temperature (30.94 ± 0.09 °C) in the IRF culture is only 5% higher than that in CWJ culture but 33% lower than that in NHC cultures. No significant differences were found in C-PC content or biomass productivity when Arthrospira is grown using IGP, CWJ or IRF but is significantly lower in NHC photobioreactors. Chlorophyll a fluorescence probing of A. platensis shows that IRF, IGP and CWJ cultures are not thermally stressed, however, NHCs cultures are highly stressed due to supraoptimal temperatures. Our results clearly indicate that solar control film is a potential tool for blocking non-photosynthetic photons and managing culture temperature in flat plate photobioreactors for sustainable C-phycocyanin production from A. platensis.

Physical extraction and extrusion entrapment of C-phycocyanin from Arthrospira platensis

C-phycocyanin (C-PC) is a blue photosynthetic pigment and the major phycobiliprotein in many blue-green algae with high nutritional values and antioxidant, antivirus, anti-cancer, and anti-inflammatory properties. It also boosts the immune system in various organisms and can be used in immunoassays to track target cells because it has fluorescent properties. This study focused on optimizing extraction conditions for C-phycocyanin encapsulation from Arthrospira platensis using ultrasonic and extrusion techniques. Parameters affecting C-phycocyanin production were type of biomass, buffer, and biomass-solvent ratio. Results indicated that the highest C-phycocyanin concentration of 6 mg ml−1 was obtained from freeze-dried samples extracted by 0.01 M sodium phosphate buffer (pH 7) and biomass-solvent ratio of 1:15, with extract purity and yield of 0.6 and 60 mg g−1, respectively. The optimized conditions of C-phycocyanin ultrasonic encapsulation at 98% were 3% sodium alginate, 2.5% calcium chloride, and 1 mg ml−1 initial C-phycocyanin concentration. C-phycocyanin extracted from A. platensis can be prepared in encapsulated form and utilized for various applications.

Elucidating the role of nutrients in C-phycocyanin production by the halophilic cyanobacterium Euhalothece sp.

In this study, a novel halophilic cyanobacterium was isolated and identified as Euhalothece sp. KZN. This fast-growing strain had the ability to synthesise high yields (12 mg g−1) of C-phycocyanin (C-PC), a highly fluorescent blue light-harvesting pigment with numerous potential uses in the biotechnology and commercial sectors. This study elucidated the individual and interactive role of different nutrients in BG11 growth medium for enhancing C-PC production in Euhalothece sp. KZN. Nine components of BG11 medium were screened for their effects via fractional factorial design (FFD). The results revealed a significant influence of nutrients, viz. MgSO4, NaNO3 and minor nutrients (citric acid, EDTA-iron citrate, CaCl2 and Na2CO3) on C-PC yield. These three components were further explored for their optimum concentration for enhancing C-PC production using a central composite design. The optimum values for these essential nutrients were found to be as follows: 0.10 g L−1 of MgSO4, 1.67 g L−1 of NaNO3 and 10 mL L−1 of minor nutrients which resulted in a 280% increase in C-PC yield with predicted and actual values of 43.97 and 45 mg g−1, respectively. Euhalothece sp. KZN is a strong potential candidate for C-PC production and can be further exploited to produce this industrially valuable compound.

A Bilevel Programming Approach to Optimize C-phycocyanin Bio-production under Uncertainty

High variability and unreliable expectations on product yields substantially hinder the industrialization of microorganism derived biochemicals as they present a risk to the profitability and safety of the underlying systems. Therefore, in this work, we propose an optimization approach to determine the lower and upper product yield expectations for the sustainable production of C-phycocyanin. Kinetic modeling is adopted in this study as it is an outstanding tool for fast prototyping, prediction and optimization of chemical and biochemical processes. On the upside, parameters in bioprocess kinetic models are used as a simplification of complex metabolic networks to enable the simulation, design and control of the process. On the downside, this conglomeration of parameters may result in significant model uncertainty. To address this shortcoming, we formulate a bilevel max-min optimization problem to obtain the worst-case scenario of our system given the uncertainty on the model parameters. By constructing parameter confidence ellipsoids, we determined the feasible region along which the parameters can minimize the system’s performance, while nutrient and light controls are used to maximize the biorenewable production. The inner minimization problem is embedded by means of the optimality conditions into the upper maximization problem and hence both are solved simultaneously. Through this approach, we determined pessimistic and optimistic scenarios for the bioproduction of C-phycocyanin and hence compute reliable expectations on the yield and profit of the process.

Phototrophic cultivation of NaCl-tolerant mutant of Spirulina platensis for enhanced C-phycocyanin production under optimized culture conditions and its dynamic modeling.

Commercial cultivation of Spirulina sp. is highly popular due to the presence of high amount of C-phycocyanin (C-PC) and other valuable chemicals like carotenoids and γ-linolenic acid. In this study, the pH and the concentrations of nitrogen and carbon source were manipulated to achieve improved cell growth and C-PC production in NaCl-tolerant mutant of Spirulina platensis. In this study,highest C-PC (147mg·L-1 ) and biomass (2.83g·L-1 ) production was achieved when a NaCl-tolerant mutant of S.platensis was cultivated in a nitrate and bicarbonate sufficient medium (40 and 60mM, respectively) at pH 9.0 under phototrophic conditions. Kinetic study of wildtype S.platensis and its NaCl-tolerant mutant was also done to determine optimum nitrate concentrations for maximum growth and C-PC production. Kinetic parameter of inhibition (Haldane model) was fitted to the relationship between specific growth rate and substrate concentration obtained from the growth curves. Results showed that the maximum specific growth rate (μmax ) for NaCl-tolerant mutant increased by 17.94% as compared to its wildtype counterpart, with a slight increase in half-saturation constant (Ks ), indicating that this strain could grow well at high concentration of NaNO3 . C-PC production rate (Cmax ) in mutant cells increased by 12.2% at almost half the value of Ks as compared to its wildtype counterpart. Moreover, the inhibition constant (Ki ) value was 207.85% higher in NaCl-tolerant mutant as compared to its wildtype strain, suggesting its ability to produce C-PC even at high concentrations of NaNO3 .

Optimization of production of C-phycocyanin and extracellular polymeric substances by Arthrospira sp.

The key factors influencing the production of C-phycocyanin (C-PC) and extracellular polymeric substances (EPS) by photoautotrophic culture of Arthrospira sp. were optimized using Taguchi method. Six factors were varied at either three or two levels as follows: light intensity at three levels; three initial culture pHs; two species of Arthrospira; three concentrations of Zarrouk's medium; three rates of aeration of the culture with air mixed with 2% v/v carbon dioxide; and two incubation temperatures. All cultures ran for 14days. The optimal conditions for the production of C-PC and EPS were different. For both products, the best cyanobacterium proved to be Arthrospira maxima IFRPD1183. The production of C-PC was maximized with the following conditions: a light intensity of 68µmol photons m-2s-1 (a diurnal cycle of 16-h photoperiod and 8-h dark period), an initial pH of 10, the full strength (100%) Zarrouk's culture medium, an aeration rate of 0.6 vvm (air mixed with 2% v/v CO2) and a culture temperature of 30°C. The concentration of Zarrouk's medium was the most important factor influencing the final concentration of C-PC. The optimal conditions for maximal production of EPS were as follows: a light intensity of 203µmol photons m-2s-1 with the earlier specified light-dark cycle; an initial pH of 9.5; a 50% strength of Zarrouk's medium; an aeration rate of 0.2 vvm (air mixed with 2% v/v CO2); and a temperature of 35°C. Production of C-PC and EPS in raceway ponds is discussed.

Higher production of C-phycocyanin by nitrogen-free (diazotrophic) cultivation of Nostoc sp. NK and simplified extraction by dark-cold shock

Nostoc sp. NK (KCTC 12772BP) was isolated and cultivated in a BG11 medium and a nitrate-free BG11 medium (BG110). To enhance C-phycocyanin (C-PC) content in the cells, different fluorescent lamps (white, plant, and red) were used as light sources for complementary chromatic adaptation (CCA). The maximum biomass productivity was 0.42g/L/d and 0.32g/L/d under BG11 and BG110 conditions, respectively. The maximum C-PC contents were 8.4% (w/w) under white lamps, 13.6% (w/w) under plant lamps, and 18% (w/w) under BG110 and the red light condition. The maximum C-PC productivity was 57.4mg/L/d in BG110 under the red lamp condition. These results indicate that a higher C-PC content could be obtained under a diazotrophic condition and a CCA reaction. The C-PC could be released naturally from cells without any extraction processes, when Nostoc sp. NK was cultivated in the BG110 medium with CO2 aeration and put in dark conditions at 5°C.

Using an innovative pH-stat CO2 feeding strategy to enhance cell growth and C-phycocyanin production from Spirulina platensis

Spirulina platensis can mitigate CO2 emissions and simultaneously produce an antioxidant called C-phycocyanin (C-PC), which is a high value nutraceutical product. An innovative pH control system was used to cultivate S. platensis, in which CO2 feeding, instead of acid/alkaline titration, was used to control the pH of the culture. The optimum culture pH was 9.5. Compared to continuous CO2 feeding system, the pH control system improved the CO2 removal efficiency from 13.6% to 62.3%. The C-PC content and productivity were also enhanced to 16.8% and 0.17g/L/d, respectively. Therefore, the proposed pH control system is economic and sustainable as it avoids the use of acid/alkaline and reduces the overall CO2 emissions. A two-stage C-PC purification process combining fractional precipitation and ion exchange chromatography could achieve the highest purity (A615/A280) of 4.33 with a 33% C-PC recovery.

Dynamic modeling and optimization of cyanobacterial C-phycocyanin production process by artificial neural network

Artificial neural networks have been widely applied in bioprocess simulation and control due to their advantageous properties. However, their feasibility in long-term photo-production process modeling and prediction as well as their efficiency on process optimization have not been well studied so far. In the current study, an artificial neural network was constructed to simulate a 15-day fed-batch process for cyanobacterial C-phycocyanin production, which to the best of our knowledge has never been conducted. To guarantee the accuracy of artificial neural network, two strategies were implemented. The first strategy is to generate artificial data sets by adding random noise to the original data set, and the second is to choose the change of state variables as training data output. In addition, the first strategy showed the distinctive advantage of reducing the experimental effort in generating training data. By comparing with current experimental results, it is concluded that both strategies give the network great modeling and predictive power to estimate the entire fed-batch process performance, even when few original experimental data are supplied. Furthermore, by optimizing the operating conditions of a 12-day fed-batch process, a significant increase of 85.6% on C-phycocyanin production was achieved compared to previous work, which suggests the high efficiency of artificial neural network on process optimization.

Optimization of C-phycocyanin production from S. platensis cultivated on mixotrophic condition by using response surface methodology

Abstract Production of phycocyanin from Spirulina platensis by mixotrophic condition was favored due to high growth rate result. The objective of this research was to obtain the statistical interaction models of molasses and urea to crude phycocyanin (CPC) production by applying response surface methology (RSM)associated central composite design approach. S. platensis was cultivated under mixotrophic condition by adding varied molasses and urea under continous illumination at 45.5 μmol m−2 s−1 for 6 days. By using 2nd order statistical models, optimum phycocyanin production was recorded on 114.74 mg L−1 and 0.196 L−1 of urea and molasses, respectively. Molasses could be the promising substrate for phycocyanin production of the microalgae.

Fed-batch strategy for enhancing cell growth and C-phycocyanin production of Arthrospira (Spirulina) platensis under phototrophic cultivation

The C-phycocyanin generated in blue-green algae Arthrospira platensis is gaining commercial interest due to its nutrition and healthcare value. In this study, the light intensity and initial biomass concentration were manipulated to improve cell growth and C-phycocyanin production of A.platensis in batch cultivation. The results show that low light intensity and high initial biomass concentration led to increased C-phycocyanin accumulation. The best C-phycocyanin productivity occurred when light intensity and initial biomass concentration were 300μmol/m2/s and 0.24g/L, respectively. The fed-batch cultivation proved to be an effective strategy to further enhance C-phycocyanin production of A.platensis. The results indicate that C-phycocyanin accumulation not only requires nitrogen-sufficient condition, but also needs other nutrients. The highest C-phycocyanin content (16.1%), production (1034mg/L) and productivity (94.8mg/L/d) were obtained when using fed-batch strategy with 5mM medium feeding.

Recent Developments in Production and Biotechnological Applications of C-Phycocyanin

An extensive range of pigments including phycobiliproteins are present in algae. C-phycocyanin (C-PC), a phycobiliprotein, is one of the key pigments of Spirulina, a microalgae used in many countries as a dietary supplement. Algal pigments have massive commercial value as natural colorants in nutraceutical, cosmetics, and pharmaceutical industries, besides their health benefits. At present, increasing awareness of harmful effects of synthetic compounds and inclination of community towards the usage of natural products have led to the exploitation of microalgae as a source of natural pigments/colors. This review describes recent findings about the sources and production of C-PC, with emphasis on specific techniques for extraction and purification, along with potential industrial applications in diagnostics, foods, cosmetics, and pharmaceutical industries.

Optimization of medium components for increased production of C-phycocyanin from Phormidium ceylanicum and its purification by single step process

Phycocyanin is a major protein produced by cyanobacteria, but very few phycocyanin-producing strains have been reported. In the present study, response surface methodology (RSM) involving a central composite design for four factors was successfully employed to optimize medium components for increased production of phycocyanin from Phormidium ceylanicum. The production of phycocyanin and interactions between sodium nitrate, calcium chloride, trace metal mix and citric acid stock were investigated and modeled. Under optimized condition P. ceylanicum was able to give 2.3-fold increase in phycocyanin production in comparison to commonly used BG 11 medium in 32 days. We have demonstrated the extraction, purification and characterization of C-phycocyanin using novel method based on filtration and single step chromatography. The protein was extracted by repeated freeze–thaw cycles and the crude extract was filtered and concentrated in stirred ultrafiltration cell (UFC). The UFC concentrate was then subjected to a single ion exchange chromatographic step. A purity ratio of 4.15 was achieved from a starting value of 1.05. The recovery efficiency of C-phycocyanin from crude extract was 63.50%. The purity was checked by electrophoresis and UV–Vis spectroscopy.