Enhanced solid‐state citric acid bio‐production using apple pomace waste through surface response methodology

Aims: To evaluate the potential of apple pomace (AP) supplemented with rice husk for hyper citric acid production through solid‐state fermentation by Aspergillus niger NRRL‐567. Optimization of two key parameters, such as moisture content and inducer (ethanol and methanol) concentration was carried out by response surface methodology. Methods and Results: In this study, the effect of two crucial process parameters for solid‐state citric acid fermentation by A. niger using AP waste supplemented with rice husk were thoroughly investigated in Erlenmeyer flasks through response surface methodology. Moisture and methanol had significant positive effect on citric acid production by A. niger grown on AP (P < 0·05). Higher values of citric acid on AP by A. niger (342·41 g kg^?1 and 248·42 g kg^?1 dry substrate) were obtained with 75% (v/w) moisture along with two inducers [3% (v/w) methanol and 3% (v/w) ethanol] with fermentation efficiency of 93·90% and 66·42%, respectively depending upon the total carbon utilized after 144 h of incubation period. With the same optimized parameters, conventional tray fermentation was conducted. The citric acid concentration of 187·96 g kg^?1 dry substrate with 3% (v/w) ethanol and 303·34 g kg^?1 dry substrate with 3% (v/w) methanol were achieved representing fermentation efficiency of 50·80% and 82·89% in tray fermentation depending upon carbon utilization after 120 h of incubation period. Conclusions: Apple pomace proved to be the promising substrate for the hyper production of citric acid through solid‐state tray fermentation, which is an economical technique and does not require any sophisticated instrumentation. Significance and Impact of the Study: The study established that the utilization of agro‐industrial wastes have positive repercussions on the economy and will help to meet the increasing demands of citric acid and moreover will help to alleviate the environmental problems resulting from the disposal of agro‐industrial wastes.     

Grape pomace: A novel substrate for microbial production of citric acid

Summary Grape pomace was used as substrate for microbial production of citric acid. Of the five cultures examined,Aspergillus niger NRRL 567 was found to produce the greatest amount of citric acid from grape pomace in the presence of methanol at a concentration of 3% (vol/wt). The yield was 60% based on the amount of fermentable sugar consumed.     

Effect of substrate moisture content on fungal production of citric acid in a solid state fermentation system

Summary Under solid state fermentation conditions, the production of citric acid byAspergillus niger varied considerably with the initial moisture content of apple pomace. The stimulating effect of methanol on fungal production of citric acid decreased markedly as the substrate moisture decreased.     

Apple pomace: A potential substrate for citric acid production by Aspergillus niger

Summary Apple pomace was used as a fsubstrate for citric acid production by five strains of Aspergillus niger. A. niger NRRL 567 produced the greatest amount of citric acid from apple pomace in the presence of 4% methanol. The yield was 88% based on the amount of sugar consumed.     

Solid state fermentation of apple pomace for citric acid production

Summary A solid state fermentation process is described for the production of citric acid from apple pomace byAspergillus niger NRRL 567. The yields of citric acid varied with the pomace varieties, and were dependent on (1) the amount of methanol present in the pomace, and (2) the fermentation time and temperature. The process yielded as much as 90 g citric acid/kg apple pomace fermented in the presence of 3–4% methanol at 30°C in five days.

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Resumen Se describe un proceso de fermentación en estado sólido para la producción de ácido cítrico a partir de residuos de manzana, utilizandoAspergillus niger NRRL 567. La producción de ácido cítrico varió con el tipo de residuo utilizado y dependió de (1) la cantidad de metanol presente en los residuos y (2) del tiempo y la temperatura del proceso. La fermentación puede producir hasta 90 g de ácido cítrico/kg de residuo de manzana, fermentado en presencia de 3–4% de metanol a 30°C de temperatura durante 5 días.

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Résumé Le procédé de fermentation en milieu solide qui est décrit est déstiné à la production d'acide citrique à partir du marc de pommes parAspergillus niger NRRL 567. Les rendements en acide citrique varient suivant les variétés de pommes et dépendent: (1) de la quantité de méthanol présente dans le marc, et (2) du temps de fermentation et de la température. On peut obtenir jusqu'à 90 g d'acide citrique par kg de marc de pommes fermenté, en présence de 3 à 4% de méthanol, à 30°C et en 5 jours.

     

Pineapple waste - a novel substrate for citric acid production by solid-state fermentation

Summary Citric acid production in solid-state fermentation by Aspergillus foetidus ACM3996 was better on pineapple waste than on apple pomace, wheat bran or rice bran. The highest citric acid content achieved on pineapple waste was 16.1 g per 100 g dried pineapple waste, with a moisture content of 70% and in the presence of 3% methanol. This represents a yield of 62.4% based on the sugar consumed.     

The kinetic basis of the role of Ca++ ions for higher yield of citric acid in a repeated-batch cultivation system

The present investigation deals with role of Ca⁺⁺ ions in increasing the yield of citric acid in a repeated-batch cultivation system (working volume 9-1) and its kinetic basis. Five different hyper-producing strains of Aspergillus niger were evaluated for citric acid production using clarified cane-molasses as basal substrate. Among the cultures, NG^GCB101 (developed by u.v./chemical mutation in our labs) gave maximum production of citric acid i.e., 87.98 g/1, 6 days after mycelial inoculation. The addition of CaCl₂ to the culture medium promoted the formation of small rounded fluffy pellets (1.55 mm, diameter), which were desirable for citric acid productivity. CaCl₂ at a level of 2.0 M, added during inoculation time, was optimized for commercial exploitation of molasses. During repeated-batch culturing, a yield of citric acid monohydrate of 128.68 g/1 was obtained when the sampling vs. substrate feeding was maintained at 4-1 (44.50% working volume). The incubation period was reduced from 6 to only 2 days. The values of kinetic parameters such as substrate consumption and product formation rates revealed the hyperproducibility of citric acid by the selected Aspergillus niger NG^GCB101 (LSD = 0.456a, HS). Case studies are highly economical because of higher yield of product, lower energy consumption and the use of raw substrate without any additional supplementation.     

Bioconversion of palm oil mill effluent for citric acid production: statistical optimization of fermentation media and time by central composite design

A laboratory-scale study was conducted to evaluate the feasibility of using palm oil mill effluent (POME) as a major substrate and other nutrients for maximum production of citric acid using the potential fungal strain Aspergillus niger (A103). Statistical optimization of medium composition (substrate–POME, co-substrates–wheat flour and glucose, and nitrogen source–ammonium nitrate) and fermentation time was carried out by central composite design (CCD) to develop a polynomial regression model through the effects of linear, quadratic, and interaction of the factors. The statistical analysis of the results showed that, in the range studied, ammonium nitrate had no significant effect whereas substrate, co-substrates and fermentation time had significant effects on citric acid production. The optimized medium containing 2% (w/w) of substrate concentration (POME), 4% (w/w) of wheat flour concentration, 4% (w/w) of glucose concentration, 0% (w/v) of ammonium nitrate and 5 days fermentation time gave the maximum predicted citric acid of 5.37 g/l which was found to be 1.5 g/l in the experimental run. The determination of coefficient (R ²) from the analysis observed was 0.964, indicating a satisfactory adjustment of the model with the response. The analysis showed that the major substrate POME (P < 0.05), glucose (P < 0.01), nutrient (P < 0.05), and fermentation time (P < 0.01) was more significant for citric acid production. The bioconversion of POME for citric acid production using optimal conditions showed the higher removal of chemical oxygen demand (82%) with the production of citric acid (5.2 g/l) on the final day of fermentation process (7 days). The pH and biosolids accumulation were observed during the bioconversion process.     

Citric and gluconic acid production from fig by Aspergillus niger using solid-state fermentation

The production of citric and gluconic acids from fig by Aspergillus niger ATCC 10577 in solid-state fermentation was investigated. The maximal citric and gluconic acids concentration (64 and 490 g/kg dry figs, respectively), citric acid yield (8%), and gluconic acid yield (63%) were obtained at a moisture level of 75%, initial pH 7.0, temperature 30°C, and fermentation time in 15 days. However, the highest biomass dry weight (40 g/kg wet substrate) and sugar utilization (90%) were obtained in cultures grown at 35°C. The addition of 6% (w/w) methanol into substrate increased the concentration of citric and gluconic acid from 64 and 490 to 96 and 685 g/kg dry fig, respectively. Journal of Industrial Microbiology & Biotechnology (2000) 25, 298–304. Received 15 April 2000/ Accepted in revised form 11 August 2000     

Citric acid production from sugar cane molasses by 2-deoxyglucose-resistant mutant strain ofAspergillus niger

Citric acid production from sugar cane molasses byAspergillus niger NIAB 280 was studied in a batch cultivation process. A maximum of 90 g/L total sugar was utilized in citric acid production medium. From the parental strainA. niger, mutant strains showing resistance to 2-deoxyglucose in Vogal's medium containing molasses as a carbon source were induced by γ-irradiation. Among the new series of mutant strains, strain RP7 produced 120 g/L while the parental strain produced 80 g/L citric acid (1.5-fold improvement) from 150 g/L of molasses sugars. The period of citric acid production was shortened from 10 d for the wild-type strain to 6–7 d for the mutant strain. The efficiency of substrate uptake rate with respect to total volume substrate consumption rate,Q _s (g per L per h) and specific substrate consumption rate,q _s (g substrate per g cells per h) revealed that the mutant grew faster than its parent. This indicated that the selected mutant is insensitive to catabolite repression by higher concentrations of sugars for citric acid production. With respect to the product yield coefficient (Y _p/x), volume productivity (Q _p) and specific product yields (q _p), the mutant strain is significantly (p≤0.05) improved over the parental strain.     

Citric acid production from beet molasses by cell recycle ofAspergillus niger

Summary Production of citric acid from beet molasses at a varying pH profile using cell recycle ofAspergillus niger was investigated. Best results in terms of citric acid concentration, yield, productivity and specific citric acid productivity were obtained with a substrate pH of 3.0.     

Mutation of Aspergillus niger for hyperproduction of citric acid from black strap molasses

Spore suspensions of Aspergillus niger GCB 75, which produced 31.1 g/l citric acid from 15% sugars in molasses, were subjected to u.v.-induced mutagenesis. Among three variants, GCM 45 was found to be the best citric acid producer and was further improved by chemical mutagenesis using NTG. Out of 3 deoxy-D-glucose-resistant variants, GCM 7 was selected as the best mutant which produced 86.1 ± 1.5 g/l citric acid after 168 h of fermentation of potassium ferricyanide + H₂SO₄-pretreated black strap molasses (containing 150 g sugars/l) in Vogel's medium. On the basis of comparison of kinetic parameters, namely the volumetric substrate uptake rate (Q _s), and specific substrate uptake rate (q _s), the volumetric productivity, theoretical yield and specific product formation rate, it was observed that the mutants were faster growing organisms and had the ability to overproduce citric acid.     

A new process for simultaneous production of tannase and phytase by <Emphasis Type="Italic">Paecilomyces variotii</Emphasis> in solid-state fermentation of orange pomace

The production of enzymes such as tannases and phytases by solid-state fermentation and their use in animal feed have become a subject of great interest. In the present work, Paecilomyces variotii was used to produce tannase and phytase simultaneously. Solid-state fermentation, a process initially designed for tannase production, was implemented here using orange pomace as substrate. Orange pomace is the waste product of the large orange juice industry in Brazil, and it has also been used as an ingredient in animal feed. In addition to enzymatic production, biotransformation of the phenolic content and antioxidant capacity of the orange pomace were analyzed after fermentation. Fermentation conditions, namely moisture level and tannic acid concentration rate, were studied using CCD methodology. The response surface obtained indicated that the highest tannase activity was 5,000 U/gds after 96 h at 59% (v/w) and 3% (w/w) and that of phytase was 350 U/gds after 72 h at 66% (v/w) and 5.8% (w/w) of moisture level and tannic acid concentration, respectively. The amount of tannase production was similar to the levels achieved in previous studies, but this was accomplished with a 7% (w/w) reduction in the amount of supplemental tannic acid required. These results are the first to show that P. variotii is capable of producing phytase at significant levels. Moreover, the antioxidant capacity of orange pomace when tested against the free radical ABTS was increased by approximately tenfold as a result of the fermentation process.     

Optimizing medium constituents and fermentation conditions for citric acid production from palmyra jaggery using response surface method

The quantitative effects of pH, temperature, time of fermentation, sugar concentration, nitrogen concentration and potassium ferrocyanide on citric acid production were investigated using a statistical experimental design. It was found that palmyra jaggery (sugar syrup from the palmyra palm) is a suitable substrate for increasing the yield of citric acid using Aspergillus niger MTCC 281 by submerged fermentation. Regression equations were used to model the fermentation in order to determine optimum fermentation conditions. Higher yields were obtained after optimizing media components and conditions of fermentation. Maximum citric acid production was obtained at pH 5.35, 29.76 °C, 5.7 days of fermentation with 221.66 g of substrate/l, 0.479 g of ammonium nitrate/l and 2.33 g of potassium ferrocyanide/l.     

Dissimilation of citric acid byStreptococcus paracitrovorus

Summary and conclusions S.paracitrovorus does not readily dissimilate citric acid in the absence of sugar but does attack citric acid relatively vigorously in the presence of small quantities of glucose or lactose. The effect of glucose and lactose in initiating the dissimilation of citric acid is catalytic.The sugars which act catalytically are themselves fermented to approximately equimolar quantities of carbon dioxide, ethyl alcohol and lactic acid. The dissimilation of a combined substrate of citrate and glucose forms, in addition, acetic acid, acetylmethylcarbinol, 2,3-butylene glycol and under certain conditions, pyruvic acid which acts as an intermediate compound. Pyruvate is dissimilated to products similar to those from a fermentation of citrate plus glucose.The reactions ofKrebs' citric acid cycle apparently do not apply to the dissimilation of citric acid byS.paracitrovorus because the fermentation of citric acid proceeds anaerobically, consumes little oxygen aerobically and is not inhibited by arsenite.Inasmuch as milk contains lactose, the fermentation of citric acid in milk byS.paracitrovorus may be catalyzed as shown in these studies.Journal paper No.J711 of the Iowa Agricultural Experiment Station, Project 451.     

苹果渣乙醇提取物抗菌活性及防腐作用研究

为提高苹果渣资源利用率,探究苹果渣乙醇提取物的抗菌活性和防腐性能,该文采用微波辅助提取法制取苹果渣乙醇提取物,用抑菌圈实验测定其抗菌活性,并研究了其防腐作用。结果表明:(1)苹果渣乙醇提取物对酵母菌抑制作用不明显(抑菌圈直径<1 mm),对金黄色葡萄球菌和大肠杆菌的抑菌作用较明显(抑菌圈直径为6～9 mm),最佳抑菌浓度为4.0 g·L^-1。(2)pH值和盐浓度对其抑菌效果有影响,pH值为6～7,盐浓度为5.0 g·L^-1,抑菌效果最好。(3)对百香果有较好的保鲜防腐效果,最佳使用浓度为0.2%。在该浓度下贮藏后的百香果腐烂率为6.7%(对照组为67%),失重率为5.5%(对照组为36.3%),可溶性固形物、总酸含量均与贮藏前差异不显著(P> 0.05)(对照组P< 0.05),且果实较饱满,硬度较高,鲜艳有光泽,酸甜适中。综上所述,苹果渣乙醇提取物对大肠杆菌和金黄色葡萄球菌具有良好的抑制作用,对百香果的保鲜防腐效果佳,可应用到天然食品的保鲜防腐。     

Comparison of citric acid production from glycerol and glucose by different strains of <Emphasis Type="Italic">Yarrowia lipolytica</Emphasis>

A wild type strain A-101 of Y. lipolytica and its three acetate-negative mutants (Wratislavia 1.31, Wratislavia AWG7, and Wratislavia K1) were compared for the production of citric acid from glucose and glycerol (pure and crude) in batch cultures. The substrates were used either as single carbon sources or as mixtures of glucose and pure or crude glycerol. The kinetic parameters, i.e., the volumetric citric acid production rate and yield obtained in the study show that the Wratislavia 1.31 and Wratislavia AWG7 strains produced the highest amount of citric acid from glycerol, with a yield from 0.40 to 0.53 g g⁻¹. This substrate was found to be a better carbon source for the biosynthesis of citric acid than glucose. The results obtained with the same strains have shown low content of isocitric acid and polyols, such as erythritol and mannitol. Y. lipolytica A-101 strain produced the highest amount of isocitric acid, from 13.8 to 21% isocitric acid in the sum of citric acids. However, the highest concentrations of erythritol were found in cultures with Y. lipolytica Wratislavia K1, from 18.1 to 30 g l⁻¹, for glucose and pure glycerol, respectively.     

Production of citric acid from starch-hydrolysate by Aspergillus niger

The present investigation explored the possible use of a rarely used agro-industrial by-product, maize starch-hydrolysate, for economic production of citric acid. To achieve this, seventeen strains of Aspergillus niger were screened for their capacity to produce citric acid using starch-hydrolysate as a substrate. The most efficient strain, ITCC-605 was selected for further improvement in citric acid content by mutation. Mutants developed by treatment with EMS and UV, singly and in combination, produced citric acid in the range of 0.51-64.7 g kg(-1) of glucose consumed. The mutant UE-1 produced the maximum citric acid which was about 130 times more than that produced by the parent strain, ITCC-605. For further increase in citric acid production from this substrate, the cultural conditions were optimized: concentration of starch-hydrolysate, 15% (glucose equivalent); ammonium nitrate, 0.25%; KH2PO4, 0.15%; nicotinic acid, 0.0001% and initial pH of 2.0. Under these conditions, the mutant strain UE-1 yielded 490 g citric acid kg(-1) of glucose consumed in 8 days of incubation at 30 degrees C. The productivity of 341 mgl(-1)h(-1) corresponded to 49% substrate conversion to citric acid.     

固态发酵技术提高苹果渣附加值的应用研究

能源危机和苹果渣带来的环境问题促进了固态发酵技术的发展.相对于液体深层发酵,固态发酵在生产中具有处理量大,操作简单,成本低等独特的优势.综述了以苹果渣为原料利用固态发酵技术生产酶制剂、柠檬酸、酒精和蛋白饲料等的研究应用进展,以及存在的问题和前景展望.     

Sugarcane-pressmud as a novel substrate for production of citric acid by solid-state fermentation

Sugarcane-pressmud, a by-product of cane-sugar manufacture, was used as a substrate for production of citric acid by Aspergillus niger CFTRI 30, in a solid-state fermentation system. Of the 170 g of sugar supplied, 131 g were consumed, with a 79% yield of citric acid over 120 h. Potassium ferrocyanide improved the conversion to about 88% and lowered the fermentation time by 24 h. Enrichment with sugar and NH₄NO₃ was essential to improve productivity. About 174 g citric acid/kg dry sugarcane-pressmud were produced after 120 h in ferrocyanid-treated medium which initially contained 12.5% (w/w) effective sugar and 0.1% (w/w) NH₄NO₃. About 3% (w/w) of the original sugar present in the sugarcane-pressmud was non-utilizable. This is the first report on the potential of sugarcane-pressmud for citric acid production.V.S. Shankaranand and B.K. Lonsane are with the Fermentation Technology and Bioengineering Discipline, Central Food Technological Research Institute, Mysore-570 013, India