Fresh juice from C grade longan (mg/g fresh fruit; total sugars 51.6 ± 0.5, nitrogen 0.021 ± 0.004; gallic acid 0.025 ± 0.001; ellagic acid 0.016 ± 0.001) with addition of 8.52 g ammonium sulfate/l was the optimal medium for cultivation processes of Saccharomyces cerevisiae TISTR 5606 and Candida tropicalis TISTR 5306 capable of producing ethanol and phenylacetylcarbinol (PAC) among five longan grades. S. cerevisiae TISTR 5606 produced ethanol (33.4 ± 3.2 g/L) at a significantly higher level (p ≤ 0.05) than C. tropicalis TISTR 5306 (22.3 ± 1.1 g/L) with similar ethanol yields (Yp/s) between 0.21 and 0.22 g ethanol produced/g sugars consumed. Whole cells of C. tropicalis TISTR 5306 produced a significantly higher (p ≤ 0.05) PAC level (27.2 ± 0.7 mM) than S. cerevisiae TISTR 5606 (3.59 ± 1.33 mM) after 6 hr in an equivalent volume biphasic biotransformation system. Practical applications Longan is one of the important economic fruit of Thailand with production volume in 2017 reaching one million tons. Less than 10% of these are domestically consumed while more than 90% are exported to several countries whose population frequently consume longan as a nutritious food supplement. This study can help solving overproduction problem of longan fruit by processing C grade fresh longan, which is accounted for 5% of the overall production volume, in the form of juice extract with the relatively high sugars content. Microbial fermentation of this juice and subsequent whole cells biotransformation process result in ethanol and PAC, respectively. Ethanol can be used as an alternative biofuel or important industrial solvent while PAC is a precursor for production of commercial nasal decongestant (ephedrine) or anti‐asthmatic compound (pseudoephedrine).
Open burning of agricultural residues causes numerous complications including particulate matter pollution in the air, soil degradation, global warming and many more. Since they possess bio-conversion potential, agro-industrial residues including sugarcane bagasse (SCB), rice straw (RS), corncob (CC) and sweet sorghum bagasse (SSB) were chosen for the study. Yeast strains, Candida tropicalis, C. shehatae, Saccharomyces cerevisiae, and Kluyveromyces marxianus var. marxianus were compared for their production potential of bioethanol and phenylacetylcarbinol (PAC), an intermediate in the manufacture of crucial pharmaceuticals, namely, ephedrine, and pseudoephedrine. Among the substrates and yeasts evaluated, RS cultivated with C. tropicalis produced significantly (p ≤ 0.05) higher ethanol concentration at 15.3 g L−1 after 24 h cultivation. The product per substrate yield (Yeth/s) was 0.38 g g-1 with the volumetric productivity (Qp) of 0.64 g L−1 h−1 and fermentation efficiency of 73.6% based on a theoretical yield of 0.51 g ethanol/g glucose. C. tropicalis grown in RS medium produced 0.303 U mL−1 pyruvate decarboxylase (PDC), a key enzyme that catalyzes the production of PAC, with a specific activity of 0.400 U mg−1 protein after 24 h cultivation. This present study also compared the whole cells biomass of C. tropicalis with its partially purified PDC preparation for PAC biotransformation. The whole cells C. tropicalis PDC at 1.29 U mL−1 produced an overall concentration of 62.3 mM PAC, which was 68.4% higher when compared to partially purified enzyme preparation. The results suggest that the valorization of lignocellulosic residues into bioethanol and PAC will not only aid in mitigating the environmental challenge posed by their surroundings but also has the potential to improve the bioeconomy.
The (R)-phenylacetylcarbinol (PAC) batch biotransformation kinetics for partially purified Candida tropicalis TISTR 5350 pyruvate decarboxylase (PDC) were determined to validate a comprehensive mathematical model in 250 mL scale with 250 mM phosphate buffer/pH 7.0. PDC could convert initial 100/120 mM benzaldehyde/pyruvate substrates to the statistical significantly highest (p ≤ 0.05) maximum PAC concentration (95.8 ± 0.1 mM) and production rate (0.639 ± 0.001 mM min−1). A parameter search strategy aimed at minimizing overall residual sum of square (RSST) based on a system of six ordinary differential equations was applied to PAC biotransformation profiles with initial benzaldehyde/pyruvate concentration of 100/120 and 30/36 mM. Ten important biotransformation kinetic parameters were then elucidated including the zeroth order activation rate constant due to phosphate buffer species (ka) of (9.38 ± < 0.01) × 10–6% relative PDC activity min−1 mM−1. The validation of this model to independent biotransformation kinetics with initial benzaldehyde/pyruvate concentration of 50/60 mM resulted in relatively good fitting with RSST, mean sum of square error (MSE), and coefficient of determination (R2) values of 662, 17.4, and 0.9863, respectively.
The carbon to nitrogen (C/N) molar ratio of 21.88 ± 0.20 in fresh juice from C‐grade longan (FLJ‐C) supplemented with ammonium sulfate (AMS) was optimal for cultivation of Candida tropicalis TISTR 5306 producing ethanol concentration of 24.0 ± 1.1 g/L and specific pyruvate decarboxylase (PDC) activity of 0.138 ± 0.001 U/mg protein. The highest ethanol concentration in a 10 L continuous process (34.3 ± 0.5 g/L, dilution rate [D] = 0.0492 hr−1) was significantly higher (p ≤ .05) than that in 100 L batch process (13.2 ± 0.2 g/L at 108 hr). The highest volumetric PDC activities of 0.081 ± 0.001 U/ml (D = 0.0070 hr−1) and 0.107 ± 0.001 U/ml (48 hr) were attained for continuous and batch processes, respectively. The affinity constants and growth‐related weighting factors on sucrose and monosaccharides (glucose and fructose with the same values) utilization were (123 ± 1 g/L, 0.14 ± 0.01) and (34.1 ± 1.5 g/L, 0.43 ± 0.01) with MS and R2 of 92.2 and .9552, respectively. Practical applications FLJ‐C is an example of agricultural produces with relatively high production quantity (50,000 t/year in Thailand) and high sugars content but low marketing demands due to the smallest size of pulp. The complex nitrogen source contains costly components (yeast extract—78.3 USD/kg, malt extract—148 USD/kg, and peptone—151 USD/kg) which can be replaced with an inexpensive AMS (1.05 USD/kg). Further consideration of C/N molar ratio in FLJ‐C can optimize growth, ethanol (alternative biofuel/industrial solvent) production, and specific PDC (biocatalyst for phenylacetylcarbinol—PAC, anti‐asthmatic and nasal decongestant precursors) activity of C. tropicalis TISTR 5306 whole cells which can be used as a case study for other related yeasts. Batch and continuous production at 100 L and 10 L also illustrate practical utilization of FLJ‐C with AMS supplement at larger scales for development of subsequent zero waste process.
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