The physical methods include comminution (ball milling, hammer milling), irradia­tion (electron beam, microwave), steam exploration (high pressure steam) and hydrothermolysis (liquid hot water). These methods have been successfully used to pre-treat other biomass for bioethanol production [58]. Physical methods of bio­mass pre-treatment mostly result in decreasing biomass particle size and degree of depolymerisation of hemicellulose and cellulose, as well as in reducing cellulose crystallinity [29]. However, physical methods are energy intensive, and the rate of hydrolysis is slow.

6.1.1 Chemical Method

The chemicals involved in the pre-treatment of biomass can be divided into two types; acidic (sulphuric, hydrochloric, phosphoric) and alkaline (lime, sodium hydroxide ammonium sulfite). Nguyen et al. [47] reported that acid pre-treatment of

Chlamydomonas reinhardtii biomass yielded around 58% (w/w) of glucose and resulted in an ethanol yield of 29.2% (w/w) at the end of the fermentation process. Aside the promising results obtained for microalgae, the efficiency of this method has been widely verified for other feedstocks [4, 9]. The chemical methods are, by far, the most prevalent biomass pre-treatment method. This is due to the low cost of the process, and its effectiveness in hydrolysing the hemicelluloses and celluloses to sugars. They are often used in conjunction with an initial degree of physical pre­treatment to reduce the biomass particle size. Also, the chemical methods are preferred due to their low operating costs when compared to the energy-intensive physical methods [29]. Since microalgal biomass does not contain lignin, the method is efficient in converting complex carbohydrates to fermentable sugars. However, the utilization of hazardous chemicals in the process can cause severe corrosion to the fermentation vessel and the resulting irrecoverable salts may form part of the biomass, thus changing the fine and specific biomass structure and its biochemical composition.

6.1.2 Biological Method

This method involves the utilization of microorganisms such as fungi to reduce the cellulose and hemicellulose crystallinity of the biomass. It is an environmentally and energetically efficient process because the pre-treatment occurs under mild con­ditions (low temperature and pH around 6-7). However, the rate of hydrolysis using this method is too slow for industrial application, and some fermentable materials are consumed in the process. Whilst this method is unlikely to be used as a sole treatment of microalgal biomass, it could serve as the first step of a multi-step pre­treatment method [24] .