Batch and continuous cultivation processes of Candida tropicalis TISTR 5306 for ethanol and pyruvate decarboxylase production in fresh longan juice with optimal carbon to nitrogen molar ratio

Abstract

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.

Validation of mathematical model with phosphate activation effect by batch (R)-phenylacetylcarbinol biotransformation process utilizing Candida tropicalis pyruvate decarboxylase in phosphate buffer

“…Although chemocatalysts can offer the relatively high catalytic activity and selectivity for some reactions 6,7 , a number of organic compounds transformation processes still rely heavily on biocatalysts to achieve the desired level of enantioselectivity [8][9][10] . Thus, biocatalysts including enzymes, cells organelles, and whole cells in either native or artificially constructed forms 11 have been widely used in the production of both high-volume/low-value compounds such as ethanol [12][13][14][15][16][17] and low-volume/high-value chemical species including (R)-phenylacetylcarbinol (PAC) 2,15,[18][19][20][21][22][23][24] .…”

Valorization of rice straw, sugarcane bagasse and sweet sorghum bagasse for the production of bioethanol and phenylacetylcarbinol

“…Sugars, ethanol and acetic acid concentration were analyzed using HPLC (Agilent Technologies, Santa Clara, California) as described by Khemacheewakul et al 43 with the following conditions: 5 mM sulphuric acid in distilled water as mobile phase with Aminex ® HPX-87H Ion Exclusive column and refractive index detector (RID) at 0.75 mL/min ow rate. The temperature of the column oven was set at 40 o C. Kinetic parameters like volumetric productivity of ethanol (Q p ), speci c total sugars consumption rate (q s,tot ), speci c growth rate (µ), yield of ethanol produced over total sugars consumed (Y eth/s ), yield of acetic acid produced over total sugars consumed (Y ace/s ), and yield of dried biomass produced over total sugars consumed (Y x/s ) were calculated based on previously published work 34,37 . Similarly, the fermentation e ciency (FE%) was calculated using the established formula based on glucose 44 and shown in supplementary section.…”

“…marxianus TISTR 5057. The stock was maintained in 80% (v/v) glycerol at -20 o C 34 . Yeasts were cultured on Yeast extract Peptone Dextrose (YPD) agar (yeast extract 10 g/L, peptone 10 g/L, agar 20 g/L and glucose 20 g/L) and subsequently grown in YPD broth at 30°C, 250 rpm for 24 h and assessed for cells viability using a haemocytometer (Count Chamber, Model: PM MFR 650030, Haryana, India).…”

Mathematical Model With Phosphate Activation Effect Validates Batch (R)-phenylacetylcarbinol Biotransformation Process Utilizing Candida Tropicalis Pyruvate Decarboxylase in Phosphate Buffer 

“…Although chemocatalysts can offer the relatively high catalytic activity and selectivity for some reactions 6,7 , a number of organic compounds transformation processes still rely heavily on biocatalysts to achieve the desired level of enantioselectivity [8][9][10] . Thus, biocatalysts including enzymes, cells organelles, and whole cells in either native or artificially constructed forms 11 have been widely used in the production of both high-volume / low-value compounds such as ethanol [12][13][14][15][16][17] and low-volume / high-value chemical species including (R)-phenylacetylcarbinol (PAC) 2,15,[18][19][20][21][22][23][24] .…”

“…PAC could be produced through in vivo direct microbial transformation process with some strategies of benzaldehyde feeding using growing cells of yeasts, fungi, and bacteria [28][29][30] . This biotransformation process can be conducted in vitro by using non-viable whole cells 15,17,24 and partially purified pyruvate decarboxylase (PDC) enzyme 18,20,28,29,[31][32][33][34] . The detailed reaction mechanism of PAC biotransformation was clearly elucidated 18 .…”