Bioprocessing of acorn kernel with Lactobacillus plantarum to reduce its tannin

Acorn kernel contains a high percentage of starch that can be used as a source of energy in animal’s diet, but it has a high percentage of tannin, which can have deleterious effects on the animals. Tannins are bitter phenolic compounds with a diverse structure that are divided into two groups of condensed and hydrolyzable. Condensed tannins are flavonoid polymers of the proanthocyanidins compounds. Hydrolyzable tannins have a polyhydric alcohol at their core, the hydroxyl groups of which are esterifies with gallic acid. They may have long chains of gallic acid coming from the central glucose core (Reed 1995). Tannins have a high tendency to bind to proteins such as enzymes (Osawa et al. 2000). The harmful consequences of the high percentage of tannin in the diet of ruminants can reduce feed intake, have a negative effect on the activity of rumen bacteria, reduce the efficiency of digestive enzymes in the livestock and also reduce the availability of nutrients, which ultimately reduce growth and production of these animals (Salinger et al. 1996). Also, the high levels of tannin compounds in the poultry diet can reduce feed intake, growth rate, digestion of protein and amino acids, intestinal and pancreatic activity (Armstrong et al. 1974; Brand et al. 1989; Mahmood et al. 1997, 2008). Various chemical and physical methods such as sodium bicarbonate, polyethylene glycol, soaking in water, cooking or steaming, have been reported to reduce the amount of tannin and its harmful side effects (Frutos et al. 2004). Now a day, bioprocessing has vast applies in livestock and poultry feed, using the enzymatic capabilities of microorganisms. Furthermore, Lactobacillus plantarum is a safe microorganism that has the potential for high enzymes to break down and eliminate tannin compounds (Curiel et al. 2009). This bacterium has vast applications for food processing in the industries (Siezen et al. 2011). Material and

The Experiment was done in a completely randomized design with 7 treatments and 3 replicates. Experimental groups were treatments with or without bacteria, aerobic or anaerobic condition and 5 or 10 days processing periods. The moisture content of all samples was adjusted to 50% by sterile water. For each gram of experimental sample, 107 CFU of Lactobacillus plantarum bacteria were added and then, placed into the incubator at 37 ° C. In this experiment, phenolic and tannin compounds, pH, soluble carbohydrate and volatile fatty acids (acetic acid, propionic acid and butyric acid) were measured. Phenolic and tannin compounds were measured based on Makkar (2000) method. Soluble carbohydrate was measured by spectrophotometric method with standard raffinose solution (Dubois 1956) and volatile fatty acids (acetic, propionic and butyric acid) were measured by gas chromatography. Soluble carbohydrate in four treatments (zero day and ten days without bacteria, ten days with bacteria in aerobic and anaerobic conditions) and volatile fatty acids in three treatments (10 days without bacteria and 10 days with bacteria in aerobic and anaerobic conditions) measured. These treatments were selected to interpret and justify the main events of fermentation processes. Data were analyzed by SAS software (2011). The comparison of the means was done using Duncan's multiple range test at P<0. 01.

The findings of the experiment indicate that the highest reductions in the content of total phenol, non-tannin phenol, total tannin, condensed tannin and hydrolyzable tannin (39, 24, 49, 49. 5 and 59 %, respectively, see table 1 and 2) were observed after 5 days of processing and there was no significant difference between 5 and 10 days of processing. It is assumed that significant reduction in phenol and tannin compounds was the consequence of Lactobacillus plantarum enzymatic activity. A research showed that Lactobacillus plantarum were reduced phenolic compounds of pomegranate juice (Mousavi et al. , 2013). Stopping the decline in phenolic compounds after the 5 days could result in lowering the pH to below 4 (Table 3) and cessation the biological activity of the Lactobacillus plantarum (Hunget 1957), or decreasing the efficiency of the enzyme tanase (tannin aceyl hydrolase). Also, there was no significant difference between the total phenol, non-tannin phenol (Table 1), total tannin, condensed tannin and hydrolyzable tannin percentage (Table 2) of aerobic and anaerobic treatments, which showed that the biological processes reduced phenolic compounds in aerobic or anaerobic conditions with the same efficiency. It has been reported that the enzymatic processes of breaking tannins compounds by bacteria in aerobic conditions are slightly different from the anaerobic conditions, resulting in different end products (Bhat et al. 1998). Our research findings show that the acidity of experimental samples in aerobic treatments was more than anaerobic in the 5 or 10 days of bioprocessing period (P<0. 01). Smetankova et al. (2012) showed that Lactobacillus plantarum grows faster in aerobic conditions and therefore, the pH decreases more quickly. After the processing, the amount of fatty acids (acetate, propionate and butyrate) increased, but the percentage of soluble carbohydrate decreased (P<0. 01). Table 4 shows that acorn kernel contains high percentage of soluble carbohydrate (on day zero), which is required for the growth and biological activity of Lactobacillus plantarum bacteria. Table 5 presents the results of statistical analysis of the treatments for the percentage of volatile fatty acids. The results showed that the percentage of acetic acid, propionic acid and butyric acid after 10 days of aerobic or anaerobic bacterial processing increased significantly, in comparison with non-bacterial treatment, but no significant difference was observed between aerobic and aerobic conditions, which indicates that in this study the activity of bacteria for the production of volatile fatty acids was not different in both aerobic and anaerobic conditions after ten days.

For the first time, this study has shown that in both aerobic and anaerobic conditions, the tannins compounds were broken down and reduced in equal proportion. The results of the experiment have shown that Lactobacillus plantarum bacteria can be used for bioprocessing of acorn kernel to reduce its tannins.