Enhanced β-mannanase production from alternative sources by recombinant Aspergillus sojae

Abstract

β-mannanases can degrade galactomannans to mannose and it has been used in various application areas. The aim of this study was to produce the β-mannanase from carob pod extract including different nitrogen sources. The best operation values for fermentation were determined to be 8% initial sugar concentration with 0.5% yeast extract, 100 r.p.m., and 7% inoculation rate, which yielded the maximum β-mannanase activity as 423.60 U ml -1 . Effects of nitrogen sources on β-mannanase activity were also studied and it reached 695.6 U ml -1 by using 0.5% of ammonium nitrate as the nitrogen source at the determined optimum conditions. Results also showed that meat bone meal and soybean meal could be used as cost effective nitrogen sources based on achieved β-mannanase activity.Keywords: β-mannanase, recombinant Aspergillus sojae, carbon sources, nitrogen sourcesMicrobial mannanase has gained signifi cant attention, since it degrades the complex polysaccharides of plant tissues into simple small molecules such as manno-oligosaccharides and mannose. For example, the increased availability and high specifi city of β-mannanases have been used for numerous applications, such as production of manno-oligosaccharides, pharmaceutical applications, instant coffee production, animal feeds, paper and pulp production, detergent formulations etc. ( VAN ZYL et al., 2010;LU et al., 2014). β-Mannanase could be generally produced by various Aspergillus species ( VAN ZYL et al., 2010) and the activity of the produced enzymes can differ from species to species. Also, other microorganisms have been evaluated for β-mannanase productions. CHEN and coworkers (2007) expressed Aspergillus sulphures β-mannanase gene in Pichia pastoris and the specifi c activity was found to be 366 U ml -1 in locust bean gum. OZTURK and co-workers (2010) used recombinant Aspergillus sojae ATCC11906 (AsT1) for β-mannanase production from molasses and found the highest activity value as 363 U ml -1. WANG and co-workers (2010) have found that Pantoea agglomerans A021 could produce β-mannanase from konjac powder and the maximum activity of purifi ed Man26P was 514 U mg -1. WU and co-workers (2011) used Aspergillus niger E-30 to produce the β-mannanase from peeled potato and the maximum β-mannanase activity value was 1067.5 U mg -1 by purifi ed enzyme solution

β‐Mannanase production and kinetic modeling from carob extract by using recombinant Aspergillus sojae

“…The first condition was 8% initial substrate concentration, 147 rpm agitation speed, and 7% inoculum size; the second condition was 8% initial substrate concentration, 100 rpm agitation speed, and 7% inoculum size, which yielded to be 339.66 and 350.82 U/mL, respectively. It was also calculated that the sugar consumed, fungal growth, maximum substrate consumption rate, and maximum β‐mannanase production rate were 64.73 g/L, 14.16 g/L (gravimetrically), 8.00 g L –1 day –1 , and 29.03 g L –1 day –1 for the first and 57.51 g/L, 14.12 g/L (gravimetrically), 7.52 g L –1 day –1 , and 26.48 U mL –1 day –1 for the second, respectively . On the other hand, the effects of different nitrogen sources (pure nitrogen sources: yeast extract, beef extract, and ammonium nitrate; animal by‐products: meat bone meal, feather meal, and fish meal; and vegetable by‐products: red lentil, cotton seed meal, maize zein, and soybean meal) on β‐mannanase fermentation were also examined by Yatmaz et al, who reported that the highest β‐mannanase activities were found to be 695.6 U/mL with 0.5% of ammonium nitrate that is a pure nitrogen source, 597.31 U/mL with 0.75% of meat bone meal that is an animal by‐product, and 525.93 U/mL with 0.75% of soybean meal that is a vegetable nitrogen source .…”

“…Besides, it was found that the highest β‐mannanase activity reached to 482 U/mL at 6th day of fermentation, which was 1.40‐fold higher than glucose‐based medium . In another study, β‐mannanase fermentation conditions by recombinant A. sojae were also optimized by using Box–Behnken RSM in terms of initial sugar concentration, agitation speed, and inoculum size in the medium containing 0.4% (w/v) yeast extract, 0.05% MgSO 4 7H 2 O, and 0.1% K 2 HPO 4 per liter, which was reported under two optimal conditions. The first condition was 8% initial substrate concentration, 147 rpm agitation speed, and 7% inoculum size; the second condition was 8% initial substrate concentration, 100 rpm agitation speed, and 7% inoculum size, which yielded to be 339.66 and 350.82 U/mL, respectively.…”

“…For inoculation, the spore suspension (containing at least 1 × 10 7 spores/mL) was prepared by washing slants with 10 mL of sterile saline solution (50 mL 0.05% Tween‐80 plus 50 mL 0.8% NaCl). Then, 100 mL of the prepared medium was aseptically inoculated with the spore solution . The carob extract used as sole carbon source in fermentation was obtained at 80°C, 25% solid loading [w/v] for 2 hr …”

“…Then, 100 mL of the prepared medium was aseptically inoculated with the spore solution. 10,23,24 The carob extract used as sole carbon source in fermentation was obtained at 80 C, 25% solid loading [w/v] for 2 hr. 25…”

“…24 On the other hand, the effects of different nitrogen sources (pure nitrogen sources: yeast extract, beef extract, and ammonium nitrate; animal by-products: meat bone meal, feather meal, and fish meal; and vegetable by-products: red lentil, cotton seed meal, maize zein, and soybean meal) on β-mannanase fermentation were also examined by Yatmaz et al, 24 who reported that the highest β-mannanase activities were found to be 695.6 U/mL with 0.5% of ammonium nitrate that is a pure nitrogen source, 597.31 U/mL with 0.75% of meat bone meal that is an animal by-product, and 525.93 U/mL with 0.75% of soybean meal that is a vegetable nitrogen source. 24 As a different study, β-mannanase fermentations in glucose-and carob extract-based media were performed by adding talcum and aluminum oxide microparticles into the fermentation medium to control the fungal morphology. 2 When talcum microparticles were inserted into the glucose-and carob extract-based media, the enzyme activity was negatively affected in the glucose-based media and the maximum activity was 199.3 U/mL with control fermentation (no-added talcum) while it was reached to maximum (568.7 U/mL) by adding 5 g/L of talcum in carob extract-based media (which was 1.83-fold higher than control fermentation).…”