The production of bioflocculants by Bacillus licheniformis using molasses and its application in the sugarcane industry

A bioflocculant produced by B. licheniformis was investigated with regard to a low-cost culture medium and its industrial application. Molasses replaced sucrose as the sole carbon source in bioflocculant fermentation. The optimum low-cost culture medium was determined to be composed of 20 g/L molasses, 0.4 g/L urea, 0.4 g/L NaCl, 0.2 g/L KH₂PO₄, 1.6 g/L K₂HPO₄, and 0.2 g/L MgSO₄. The bioflocculant from B. licheniformis was then applied to treat sugarcane-neutralizing juice to remove colloids, suspended particles, and coloring matters in a sugar refinery factory. The optimal operation conditions were a bioflocculant dosage of 21 U/mL, pH 7.3 and a heating temperature of 100°C. The color and turbidity of the sugarcane juice reached IU 1267 and IU 206, respectively, after clarification with the bioflocculant; these values were almost the same as those acquired following treatment with polyacrylamide (PAM), the most widely applied flocculant in sugar industries. These results suggest the great potential for use of bioflocculants in the sugar refinery process.     

Study on the flocculability of the Arthrobacter sp., an actinomycete resuscitated from the VBNC state

A bioflocculant with high flocculating activity, LC13-SF, produced by strain LC13^T which was in a viable but nonculturable (VBNC) state, and which was woken up by Rpf (resuscitation promoting factor), was systematically investigated with regard to its fermentation conditions and flocculating activity. The key parameters influencing the bioflocculant LC13-SF were investigated through measuring the optical density at 660 (OD₆₆₀) of the fermentation liquid and the optical density at 550 (OD₅₅₀) of the centrifugal supernatant. The flocculating efficiency and the Zeta potentials were chosen as the response variables for the study of the flocculating activity. The results showed that the optimal conditions for bioflocculant LC13-SF production were a fermentation time of 72 h, an initial pH of 7.0, a fermentation temperature of 30°C and a shaking speed of 150 r/min. The optimized flocculating process was as follows: a final volume percentage of bioflocculant LC13-SF and 0.5% (w/w) CaCl₂ were 1.5 and 5%, respectively in a 4 g/L Kaolin suspension, and the system pH was adjusted to 8.0. Under these conditions, the flocculating efficiency and the absolute value of the Zeta potential reached 94.83% and 4.37, respectively.     

Evaluation of the flocculation potential and characterization of bioflocculant produced by Micrococcus sp. Leo

Bioflocculants are safe, biodegradable and environmentally friendly biopolymeric materials. These merits portend it as preferred alternative to inorganic and organic synthetic polymeric flocculants. The culture conditions optimal for the production of bioflocculant by Micrococcus sp. Leo with subsequent evaluation of the properties of the produced compound were investigated. Optimum culture conditions for bioflocculant production included 2% (vol/vol) inoculum size, incubation temperature of 28°C, agitation speed of 160 rpm and initial pH of 4.0. Glucose and (NH₄)₂SO₄ and Al³⁺ were the best as sole carbon, nitrogen and cation sources, respectively. The purified bioflocculant flocculated kaolin suspension optimally at a dosage of 0.2 mg/mL following jar test, and flocculating activity of about 70% was retained after heat treatment of 100°C. Chemical analysis showed that the bioflocculant was composed of 28.4% polysaccharide, 2.6% protein and 9.7%. uronic acid. Thermogravimetric analysis demonstrated that the bioflocculant could not decompose completely at 400°C. FTIR spectra revealed the presence of hydroxyl, carboxyl and amino groups as the main functional groups. The bioflocculant produced by Micrococcus sp. Leo appears to hold promise as an alternative to conventional flocculants commonly used in water/wastewater treatment.     

Characterization and application of bioflocculant prepared by Rhodococcus erythropolis using sludge and livestock wastewater as cheap culture media

A new bioflocculant was produced by culturing Rhodococcus erythropolis in a cheap medium. When culture pH was 7.0, inoculum size was 2 % (v/v), Na₂HPO₄ concentration was 0.5 g L^?1, and the ratio of sludge/livestock wastewater was 7:1 (v/v), a maximum flocculating rate of 87.6 % could be achieved. Among 13 different kinds of pretreatments for sludge, the optimal one was the thermal-alkaline pretreatment. Different from a bioflocculant produced in a standard medium, this bioflocculant was effective over a wide pH range from 2 to 12 with flocculating rates higher than 98 %. Approximately, 1.6 g L^?1 of crude bioflocculant could be harvested using cold ethanol for extraction. This bioflocculant showed color removal rates up to 80 % when applied to direct and disperse dye solutions, but only 23.0 % for reactive dye solutions. Infrared spectrum showed that the bioflocculant contained functional groups such as –OH, –NH₂, and –CONH₂. Components in the bioflocculant consisted of 91.2 % of polysaccharides, 7.6 % of proteins, and 1.2 % of DNA. When the bioflocculant and copper sulfate (CuSO₄) were used together for decolorization in actual dye wastewater, the optimum decolorization conditions were specified by the response surface methodology as pH 11, bioflocculant dosage of 40 mg/L, and CuSO₄ 80 mg/L, under which a decolorization rate of 93.9 % could be reached.     

Bioconversion of lignocellulose and simultaneous production of cellulase,ligninase and bioflocculants by Alcaligenes faecalis-X3

Lignocelluloses have been used as carbon sources for bioflocculant production. However, the low bioconversion efficiency of lignocellulose to bioflocculants is a major challenge. In this study, a lignocellulolytic strain of Alcaligenes faecalis-X3 was cultivated in ramie bio-degumming wastewater. Optimal production of ligninase, cellulase and bioflocculants (MBF-X3) was evaluated. The highest activity of MBF-X3 under the optimal conditions of pH 6.0 at 48 h of fermentation was 95.44%, with the maximum production of ligninase and cellulase (0.27 and 0.12 U/mL, respectively). The crude ligninase and cellulase had optimum activities at pH 5.0 and 40 °C and pH 6.0 and 50 °C, respectively. The cellulase activity was increased by Mn²⁺, Ca²⁺, Zn²⁺, and Mg²⁺ at 1 mM. The ligninase activity was significantly enhanced in the presence of Zn²⁺ at 10 mM. The flocculating activity of MBF-X3 was not changed by the addition of any metal cation. The results demonstrated that A. faecalis possesses an excellent enzyme system for the efficient bioconversion of lignocellulose into MBF-X3. Additionally, MBF-X3 has a high flocculating efficiency of Disperse Blue-2BLN (85.7%) at a dose of 1.0 g/L.     

一株维氏气单胞菌发酵培养基优化及其制备疫苗免疫效力比较

采用单因素试验、响应面试验法对维氏气单胞菌(Aeromonas veronii)发酵培养基的氮源、碳源、无机盐和磷酸盐成分及用量进行优化组合,确定优化培养基组成：胰蛋白胨10.8 g/L,葡萄糖5.0 g/L,牛肉膏3.0 g/L,磷酸二氢钾2.0 g/L,硫酸镁0.4 g/L,NaCl 5.0 g/L。并与基础培养基的发酵活菌数、制备的灭活疫苗免疫效力进行比较,经过验证试验绘制维氏气单胞菌在优化培养基条件下的7 L发酵罐生长曲线。在优化发酵培养基条件下,维氏气单胞菌活菌数为5.94×10⁹ cfu/mL,比基础培养基增幅43.13%;制备的灭活疫苗相对保护率为77.78%,比基础培养基提高了14.81%。7 L发酵罐发酵培养10 h,活菌数达到最大8.85×10⁹ cfu/mL。通过对发酵培养基的优化,可以获得低成本、优质高效的维氏气单胞菌发酵菌液,为今后维氏气单胞菌灭活疫苗规模化发酵培养提供参考。     

Production of a novel bioflocculant MNXY1 by Klebsiella pneumoniae strain NY1 and application in precipitation of cyanobacteria and municipal wastewater treatment

Aims: To isolate and characterize the novel bioflocculant‐producing bacteria, to optimize the bioflocculant production and to evaluate its potential applications. Methods and Results: Klebsiella pneumoniae strain NY1, a bacterium that produces a novel bioflocculant (MNXY1), was selected on the chemically defined media. It was classified according to the 16S rRNA gene sequence, morphological and microscopic characteristics. MNXY1 was characterized to contain 26% protein and 66% total sugar. The constituent sugar monomers of MNXY1, revealed by NMR analysis, are glucose, galactose and quinovose. Favourable culture conditions for MNXY1 production were determined. Strain NY1 produces a high level (14.9 g l^?1) of MNXY1. MNXY1 is thermostable and tolerant to the extreme pH. It precipitated 54% of cyanobacteria from laboratory culture and 72% of the total suspended solids from raw wastewater. Conclusions: Strain NY1 was identified to produce a novel bioflocculant MNXY1. The outstanding performance of MNXY1 in practical applications and its availability in copious amounts make it attractive for further investigation and development for industrial scale applications. Significance and Impact of the Study: This is first report for the identification of a quinovose‐containing bioflocculant and application of a protein–polysaccharide complex bioflocculant in precipitation of cyanobacteria. These findings suggest that MNXY1 holds great potential for use in management of harmful algae and city wastewater treatment.     

Production and characterization of an intracellular bioflocculant by Chryseobacterium daeguense W6 cultured in low nutrition medium

A novel intracellular bioflocculant (named MBF-W6) produced by Chryseobacterium daeguense W6 cultured in low nutrition medium was investigated in this study. The effects of carbon source, nitrogen source, C/N ratio, initial pH, inoculum size, culture temperature and shaking speed on MBF-W6 production were studied. Chemical analysis showed that the purified MBF-W6 was mainly composed of 32.4% protein, 13.1% polysaccharide and 6.8% nucleic acid. Fourier-transform infrared (FTIR) spectroscopy indicated the presence of carboxyl, hydroxyl, and methoxyl groups. The elemental analysis of purified MBF-W6 revealed that the mass proportion of C, H, O, N and S was 40.92:6.53:44.01:8.53:1.01 (w/w) correspondingly. MBF-W6 had good flocculating rate in Kaolin suspension without any cation addition. The highest flocculating rate of 96.9% was achieved under the optimal conditions (bioflocculant dosage 1.2 mg l⁻¹, pH 5.6 and temperature 15 °C).     

共存碳源对克雷伯氏菌NIII2发酵蔗糖产絮凝剂的影响

研究了共存碳源对克雷伯氏菌NIII2以蔗糖为主要碳源发酵产絮凝剂的影响.实验结果表明:柠檬酸为共存碳源时,克雷伯氏菌NIII2分泌絮凝剂过程中容易产酸,使得絮凝剂的产量和碳源转化率都较低.当丁二酸、乙酸、乳酸为共存碳源时,发酵液pH均高于7.5,絮凝剂产量有所提高,最高可达10.87g/L,碳源转化率也较高,为43.48%.与柠檬酸为共存碳源相比,当投加丁二酸时,克雷伯氏菌NIII2所产微生物絮凝剂中蛋白质与糖含量比值提高了33%,絮凝剂的Zeta电位值由-60.00 mV升高至-28.07 mV,絮凝剂分子粒径广泛分布在0~300μm之间且大粒径分子所占比例增加,聚合度加大,絮凝剂表面形貌呈现结块团状无定型结构,从而提高絮凝剂的活性和性能.该微生物絮凝剂投加量为4.0 mg/L,对2 g/L高岭土的SS去除率可达97.3%.     

响应面法优化褐藻胶降解菌Cobetia sp.20发酵培养基

为了提高褐藻胶降解菌株Cobetia sp.20产褐藻胶裂解酶的能力,利用响应面法优化其发酵产褐藻胶裂解酶的培养基。首先利用单因素法分别对发酵培养基中的不同碳源、碳源添加量、不同氮源、氮源添加量以及氯化钠添加量、磷酸二氢钾添加量、硫酸镁添加量和pH进行探究,研究各因素对产酶的影响。在单因素实验的基础上,通过Plackett-Burman试验确定Cobetia sp.20发酵培养基中影响产酶的主要因素。通过响应面试验建立回归方程。研究结果表明,Cobetia sp.20最优发酵培养基配方为褐藻胶15.00 g/L、硫酸铵7.50 g/L、氯化钠15.00 g/L、硫酸镁0.50 g/L、磷酸二氢钾5.30 g/L、硫酸亚铁0.01 g/L、pH值7.58。优化后酶活为142.79 U/mL,比优化前提高了26.36%。褐藻胶裂解酶活的提高,为褐藻胶裂解酶的工业化生产提供了参考。     

混合白腐真菌的固定化及其在治理铅污染废水中的应用

【目的】为缓解重金属废水污染对全球食品安全和人类健康的威胁,降低铅(plumbum, Pb)在土壤及动植物体内的积累,借助固定化技术提高菌株的重金属去除效率。【方法】以白腐真菌(white rot fungi)为实验材料,通过混菌兼容性及铅离子(Pb²⁺)去除能力筛选出吸附效果好且兼容性优的复合菌种,探究最优混菌类型及其比例,优化菌球最佳固定化助剂配方,在此基础上深入探究菌球在实际应用中的最优吸附条件。【结果】黄孢原毛平革菌(Phanerochaete chrysosporium)、云芝(Coriolus versicolor)、凤尾菇(Lentinus sajor-caju)和平菇(Pleurotus ostreatus) 4种菌株兼容效果佳,可进行后续实验;其中云芝和凤尾菇以体积1:1混合后对Pb²⁺去除效果显著优于各单菌作用;固定化条件优化实验中,20.0 g/L海藻酸钠、15.0 g/L生物炭和2.0×10⁶个/mL白腐真菌组成混菌体系,辅以二氧化硅及沸石制得的固定化菌球在96 h Pb²⁺...     

Food-Grade Expression and Characterization of a Dextranase from Chaetomium gracile Suitable for Sugarcane Juice Clarification

The microbial production of dextranase using cheap carbon sources is beneficial to solve the economic loss caused by the accumulation of dextran in syrup. A food-grade microbial cell factory was constructed by introducing the dextranase encoding gene DEX from Chaetomium gracile to the chromosome of Bacillus subtilis, and the antibiotic resistance marker gene was subsequently deleted via the Cre/loxP strategy. The dual-promoter system with a sequentially arranged constitutive P43 promoter resulted in an 85 % increase in DEX expression. Under the optimal fermentation conditions of 10 g/L maltose, 15 g/L casein, 1 g/L Na₂HPO₄, 1 g/L FeSO₄ and 8 g/L NaCl, DEX activity was increased from 2.625 to 64.34 U/mL. Recombinant DEX was purified 5.98-fold with a recovery ratio of 26.67 % and specific activity of 3935.02 U/mg. Enzyme activity was optimal at 55 °C and pH 5.0 and remained 80.34 % and 71.36 % of the initial activity at 55 °C and pH 4.0 after 60 min, respectively. The enzyme possessed high activity in the presence of Co²⁺, while Ag⁺ showed the strongest inhibition ability. The optimal substrate was 20 g/L dextran T-2000. The findings could facilitate the low-cost, large-scale production of food-grade DEX for use in the sugar industry.     

Optimization of Mixed Cultivation of the Moderate Thermophilic Bioleaching Microorganisms for High Cell Density Using Statistical Methodology

The low functional microbial population density in the industrial bioleaching process has been a limiting factor for the high leaching efficiency, making the microbial cultivation and continuous inoculation an alternative for sustaining the microbial activity. In the present experiment, the defined mixed cultivation of Leptospirillum ferriphilum YSK, Sulfobacillus acidophilus TPY, Acidithiobacillus caldus S2, and Ferroplasma thermophilum L1 was evaluated and optimized by Statistical Methodology. Going through the Plackett–Burman experimental design, pH value, temperature, and c(MgSO₄·7H₂O) were considered as the most significant factors in the defined range. Then, the relationships were analyzed using the steepest ascent design, the central composite design, and finally the response surface methodology. It was suggested that the optimum parameters were pH 1.38, MgSO₄·7H₂O 0.552?g/L, temperature 44?°C, FeSO₄·7H₂O 40?g/L, sulfur 8?g/L, yeast 0.02% w/v, (NH₄)₂SO₄ 3g/L, K₂HPO₄ 0.5g/L, KCl 0.1g/L, Ca(NO₃)₂ 0.01?g/L, in which allowed total cell density of the microbial community to reach 7.63?×?10⁸ cells/mL in the cultivation period. The lab experiments were routinely undertaken with the expectation that the L. ferriphilum YSK, S. acidophilus TPY, A. caldus S2, F. thermophilum L1 could rapid grown from initial cell density of 0.25?×?10⁷ cells/mL to 2.82?×?10⁸ cells/mL, 1.68?×?10⁸ cells/mL, 2.76?×?10⁸ cells/mL, 2.51?×?10⁷ cells/mL, respectively in 58?h. It demonstrates a possibility to co-culture these microbes in a single reactor, providing an efficient way to regenerate of inoculation for biomining process.     

Statistical optimization of culture media for production of phycobiliprotein by Synechocystis sp. PCC 6701

Synechocystis sp. PCC 6701 has a brilliantly colored pigment, phycobiliprotein containing phycoerythrin. Culture medium was optimized by sequential designs in order to maximize phycobiliprotein production. The observed fresh weights after 6 days were 0.58 g/L in BG-11, 0.83 g/L in medium for Scenedesmus sp. and 0.03∼0.52 g/L in the other tested media. Medium for Scenedesmus sp. was selected to be optimized by fractional factorial design and central composite design since the medium maintained a more stable pH within a desirable range due to higher contents of phosphate. The fractional factorial design had seven factors with two levels: KNO₃, NaNO₃, NaH₂PO₄, Na₂HPO₄, Ca(NO₃)₂, FeEDTA, and MgSO₄. From the result of fractional factorial design, nitrate and phosphate were identified as significant factors. A central composite design was then applied with four variables at five levels each: nitrate, phosphate, pH, and light intensity. Parameters such as fresh weight and phycobiliprotein contents were used to determine the optimum value of the four variables. The proposed optimum media contains 0.88 g/L of nitrate, 0.32 g/L of phosphate under 25 μE·m⁻²·s⁻¹ of light intensity. The maximum phycobiliprotein contents have been increased over 400%, from 4.9 to 25.9 mg/L after optimization.     

Optimized production of 2,3-butanediol by a lactate dehydrogenase-deficient mutant of Klebsiella pneumoniae

To obtain high-yield production of 2,3-butanediol (2,3-BD) from glucose, we optimized the culture conditions for a lactate dehydrogenase-deficient mutant (ΔldhA) of Klebsiella pneumoniae using response surface methodology. 2,3-BD production was successfully improved by optimizing pH (5.6), aeration (3.50 vvm) and concentration of corn steep liquor (45.0 mL/L) as a nitrogen source, resulting in a maximum level of 2,3-BD production of 148.8 g/L and productivity of 2.48 g/L/h. 2,3-BD was also obtained with high concentration (76.24 g/L) and productivity (2.31 g/L/h) from the K. pneumoniae mutant strain using sugarcane molasses as a carbon source.     

重组大肠杆菌产γ-环糊精葡萄糖基转移酶的摇瓶发酵优化

为了实现来源于碱性芽孢杆菌Alkalophilic Bacillus clarkii 7364的γ-环糊精葡萄糖基转移酶的高效胞外表达,对OmpA信号肽介导的E.coli BL21(DE3)/pET20b(+)-γcgt基因工程菌进行发酵培养基及发酵条件的优化,并进行正交试验,获得最优培养基:甘油5g/L、蛋白胨6g/L、酵母膏24g/L、钙离子6mmol/L、镁离子2mmol/L、甘氨酸0.75%、PO₄^3- 0.1mol/L;在此基础上最适发酵条件:pH6.5、25℃培养、装液量30ml/250ml、转速220r/min、0.02%SDS、在发酵10h时利用5g/L乳糖进行诱导,使得酶活从初始的5189.2U/ml提高到20268.8U/ml。研究结果得到高效表达的培养条件,为实现该酶的工业化应用打下了基础。     

MEDIUM OPTIMIZATION OF ANTIFUNGAL ACTIVITY PRODUCTION BY Bacillus amyloliquefaciens USING STATISTICAL EXPERIMENTAL DESIGN

In order to overproduce biofungicides agents by Bacillus amyloliquefaciens BLB371, a suitable culture medium was optimized using response surface methodology. Plackett–Burman design and central composite design were employed for experimental design and analysis of the results. Peptone, sucrose, and yeast extract were found to significantly influence antifungal activity production and their optimal concentrations were, respectively, 20 g/L, 25 g/L, and 4.5 g/L. The corresponding biofungicide production was 250 AU/mL, corresponding to 56% improvement in antifungal components production over a previously used medium (160 AU/mL). Moreover, our results indicated that a deficiency of the minerals CuSO₄, FeCl₃ · 6H₂O, Na₂MoO₄, KI, ZnSO₄ · 7H₂O, H₃BO₃, and C₆H₈O₇ in the optimized culture medium was not crucial for biofungicides production by Bacillus amyloliquefaciens BLB371, which is interesting from a practical point of view, particularly for low-cost production and use of the biofungicide for the control of agricultural fungal pests.     

Optimization of culture conditions for 1,3-propanediol production from crude glycerol by <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis> using response surface methodology

To produce 1,3-propanediol (1,3-PD) from crude glycerol, cultivation conditions were optimized by response surface methodology (RSM) based on a 2⁵ factorial central composite design (CCD). RSM was adopted to derive a statistical model for the individual and interactive effects of crude glycerol, (NH₄)₂SO₄, pH, cultivation time and temperature on the production of 1,3-PD. Optimal conditions for maximum 1,3-PD production were as follows: crude glycerol, 35 g/L; (NH₄)₂SO₄, 8 g/L; pH, 7.37; cultivation time, 10.8 h; temperature, 36.88°C. Under these optimal conditions, the design expert presented the maximal numerical solution with a predicted 1,3-PD production level of up to 13.74 g/L. The experimental production of 1,3-PD yielded 13.8 g/L, which was in close agreement with the model prediction.     

Industrial wastewaters and dewatered sludge: rich nutrient source for production and formulation of biocontrol agent, <Emphasis Type="Italic">Trichoderma viride</Emphasis>

Axenic cultivation of biocontrol fungus Trichoderma viride was conducted on a synthetic medium and different wastewaters and wastewater sludges in shake flasks to search for a suitable raw material resulting in higher biocontrol activity. Soluble starch based synthetic medium, dewatered municipal sludge, cheese industry wastewater sludge, pre-treated and untreated pulp and paper industry wastewater and slaughter house wastewater (SHW) were tested for T. viride conidia and protease enzyme production. The maximum conidia production followed the order, soluble starch medium (>10⁹ c.f.u./mL), untreated pulp and paper industry wastewater (4.9 × 10⁷ c.f.u./mL) > cheese industry wastewater (1.88 × 10⁷ c.f.u./mL) ≈ SHW (1.63 × 10⁷ c.f.u./mL) > dewatered municipal sludge (3.5 × 10⁶ c.f.u./mL) > pre-treated pulp and paper industry wastewater (1.55 × 10⁶ c.f.u./mL). The protease activity of T. viride was particularly higher in slaughterhouse wastewater (2.14 IU/mL) and dewatered municipal sludge (1.94 IU/mL). The entomotoxicity of soluble starch based synthetic medium was lower (≈6090 SBU/μL) in contrast to other raw materials. The entomotoxicity inversely decreased with carbon to nitrogen ratio in the growth medium and the conidia concentration and protease activity also contributed to the entomotoxicity. The residual c.f.u./g formulation of T. viride conidia were up to approximately, 90% after 1 month at 4 ± 1 °C and about 70% after 6 months at 25 ± 1 °C. Thus, production of T. viride conidia would help in marketability of low cost biopesticide from the sludge and safe reduction of pollution load.     

Effect of culture medium on hydrogen production by sulfur-deprived marine green algae Platymonas subcordiformis

To study the effect of culture medium on hydrogen production by the marine green algae, Platymonas subcordiformis under sulfur deprivation, cell growth, hydrogen production, and starch and protein catabolism was investigated in the work. Algae cells cultured only in optimized medium required 6～8 days to reach the late logarithmic at the approximate density of (2.00 ± 0.18) × 10⁶ cells/mL, which in traditional medium needed 18～22 days to reach (1.85 ± 0.20) × 10⁶ cells/mL. Increased levels of Chlorophyll (10.74 ± 0.20 μg/mL), starch (149.50 ± 6.15 μg/mL), and protein (213.00 ± 7.36 μg/mL) were accumulated in optimized medium, which were 1.06, 1.47, and 1.87-fold of the algae cells cultured in traditional medium, respectively. The sealed culture of algae cells in sulfur-deprived optimized medium shifted to anaerobic conditions after 96 h of light illumination and produced 0.45 ± 0.12 mL H₂, but in traditional medium maintained aerobic condition and no hydrogen was produced. In addition, changes in starch and protein content during continuous light illumination indicated that more endogenous substrate was consumed in the sulfur-deprived optimized medium than that in the sulfur-deprived traditional medium.