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Abstract
Raphidocelis subcapitata and Chlorella vulgaris are bioassay microalgae with rigid cellulosic cell wall which can hinder the release of intracellular proteins often studied as toxicity biomarkers. Since cell disruption is necessary for recovering intracellular biomolecules in these organisms, this study investigated the efficiency of ultrasonication bath; ultrasonication probe; vortexer; and bead mill in disintegrating the microalgae for anti-oxidative enzyme extraction. The extent of cell disruption was evaluated and quantified using bright field microscopy. Disrupted algae appeared as ghosts. The greatest disintegration of the microalgae (83-99.6 %) was achieved using bead mill with 0.42-0.6 mm glass beads while the other methods induced little or no disruption. The degree of cell disruption using bead mill increased with exposure time, beads-solution ratio and agitation speed while larger beads caused less disruption. Findings revealed that bead milling, with specific parameters optimized, is one of the most effective methods of disintegrating the robust algal cells.
Highlights
The use of the unicellular microalgae, Raphidocelis subcapitata and Chlorella vulgaris as bioassay test organisms is significant as they play a vital role in the structure and functioning of the aquatic ecosystems and any adverse effect on them will affect the upper trophic levels (Li et al, 2006)
Since cell disruption is necessary for recovering intracellular biomolecules in these organisms, this study investigated the efficiency of ultrasonication bath; ultrasonication probe; vortexer; and bead mill in disintegrating the microalgae for anti-oxidative enzyme extraction
The microalgae, R. subcapitata formerly known as Selenastrum capricornutum and C. vulgaris were obtained from Culture Collection of Algae and Protozoa (CCAP), UK, and grown in 500-ml Erlenmeyer flasks containing sterile Jaworski’s medium (JM)
Summary
The use of the unicellular microalgae, Raphidocelis subcapitata and Chlorella vulgaris as bioassay test organisms is significant as they play a vital role in the structure and functioning of the aquatic ecosystems and any adverse effect on them will affect the upper trophic levels (Li et al, 2006). A range of treatment methods have been suggested and used for breaking algal cells These include freezing, alkali and organic solvents, sonication, high pressure homogenization (French press), bead milling, microwaving, enzymatic lysis and manual grinding (Wimpenny, 1967; Chisti and Moo-young, 1986; Molina Grima et al, 2004; Zheng et al, 2011). The French press, sonication and bead beating are considered the most effective and widely used methods for the homogenization of single cells, cell suspensions and microorganisms for the recovery of intracellular products (Burden, 2012). High frequency sonic waves created by the expansion and contraction of a crystallized probe or bath, bring about implosive collapse of the gas-filled cavitation bubbles and generate intense microscopic shock waves, which cause cell wall disruption (Zheng et al, 2011)
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