Control of agitation and aeration rates in the production of surfactin in foam overflowing fed-batch culture with industrial fermentation

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Castellari, María G. Cendoya, Facundo J. Marcos Valle, Viviana Barrera, Ana M. Pacin" "autores" => array:5 [ 0 => array:2 [ "nombre" => "Claudia C." "apellidos" => "Castellari" ] 1 => array:2 [ "nombre" => "María G." "apellidos" => "Cendoya" ] 2 => array:2 [ "nombre" => "Facundo J." "apellidos" => "Marcos Valle" ] 3 => array:2 [ "nombre" => "Viviana" "apellidos" => "Barrera" ] 4 => array:2 [ "nombre" => "Ana M." "apellidos" => "Pacin" ] ] ] ] ] "idiomaDefecto" => "es" "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S0325754115001145?idApp=UINPBA00004N" "url" => "/03257541/0000004700000004/v2_201602050024/S0325754115001145/v2_201602050024/es/main.assets" ] "itemAnterior" => array:19 [ "pii" => "S032575411500125X" "issn" => "03257541" "doi" => "10.1016/j.ram.2015.09.004" "estado" => "S300" "fechaPublicacion" => "2015-10-01" "aid" => "65" "copyright" => "Asociación Argentina de Microbiología" "documento" => "article" "crossmark" => 1 "licencia" => "http://creativecommons.org/licenses/by-nc-nd/4.0/" "subdocumento" => "fla" "cita" => "Rev Argent Microbiol. 2015;47:335-43" "abierto" => array:3 [ "ES" => true "ES2" => true "LATM" => true ] "gratuito" => true "lecturas" => array:2 [ "total" => 1983 "formatos" => array:3 [ "EPUB" => 35 "HTML" => 1391 "PDF" => 557 ] ] "en" => array:13 [ "idiomaDefecto" => true "cabecera" => "Original article" "titulo" => "The decrease in the population of Gluconacetobacter diazotrophicus in sugarcane after nitrogen fertilization is related to plant physiology in split root experiments" "tienePdf" => "en" "tieneTextoCompleto" => "en" "tieneResumen" => array:2 [ 0 => "en" 1 => "es" ] "paginas" => array:1 [ 0 => array:2 [ "paginaInicial" => "335" "paginaFinal" => "343" ] ] "titulosAlternativos" => array:1 [ "es" => array:1 [ "titulo" => "La disminución de la población de Gluconacetobacter diazotrophicus en caña de azúcar, después de la fertilización nitrogenada, está relacionada con la fisiología de las plantas en experimentos de raíz dividida" ] ] "contieneResumen" => array:2 [ "en" => true "es" => true ] "contieneTextoCompleto" => array:1 [ "en" => true ] "contienePdf" => array:1 [ "en" => true ] "resumenGrafico" => array:2 [ "original" => 0 "multimedia" => array:7 [ "identificador" => "fig0015" "etiqueta" => "Figure 3" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr3.jpeg" "Alto" => 3411 "Ancho" => 2383 "Tamanyo" => 255487 ] ] "descripcion" => array:1 [ "en" => "Bacterial population inside roots in split root experiments. Each value represents the media of data for five independent plants (Log of cell number/g root) with the respective standard deviation. Mean values with equal letters are not statistically different at p≤0.05, using the t-Student test. dpi: days post inoculation; dpf: days post fertilization; ND: not detected; N+: addition of 180mg of nitrogen/plant." ] ] ] "autores" => array:1 [ 0 => array:2 [ "autoresLista" => "Osvaldo Rodríguez-Andrade, Luis E. Fuentes-Ramírez, Yolanda E. Morales-García, Dalia Molina-Romero, María R. Bustillos-Cristales, Rebeca D. Martínez-Contreras, Jesús Muñoz-Rojas" "autores" => array:7 [ 0 => array:2 [ "nombre" => "Osvaldo" "apellidos" => "Rodríguez-Andrade" ] 1 => array:2 [ "nombre" => "Luis E." "apellidos" => "Fuentes-Ramírez" ] 2 => array:2 [ "nombre" => "Yolanda E." "apellidos" => "Morales-García" ] 3 => array:2 [ "nombre" => "Dalia" "apellidos" => "Molina-Romero" ] 4 => array:2 [ "nombre" => "María R." "apellidos" => "Bustillos-Cristales" ] 5 => array:2 [ "nombre" => "Rebeca D." "apellidos" => "Martínez-Contreras" ] 6 => array:2 [ "nombre" => "Jesús" "apellidos" => "Muñoz-Rojas" ] ] ] ] ] "idiomaDefecto" => "en" "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S032575411500125X?idApp=UINPBA00004N" "url" => "/03257541/0000004700000004/v2_201602050024/S032575411500125X/v2_201602050024/en/main.assets" ] "en" => array:20 [ "idiomaDefecto" => true "cabecera" => "Original article" "titulo" => "Control of agitation and aeration rates in the production of surfactin in foam overflowing fed-batch culture with industrial fermentation" "tieneTextoCompleto" => true "paginas" => array:1 [ 0 => array:2 [ "paginaInicial" => "344" "paginaFinal" => "349" ] ] "autores" => array:1 [ 0 => array:4 [ "autoresLista" => "Shulin Yao, Shengming Zhao, Zhaoxin Lu, Yuqi Gao, Fengxia Lv, Xiaomei Bie" "autores" => array:6 [ 0 => array:2 [ "nombre" => "Shulin" "apellidos" => "Yao" ] 1 => array:2 [ "nombre" => "Shengming" "apellidos" => "Zhao" ] 2 => array:2 [ "nombre" => "Zhaoxin" "apellidos" => "Lu" ] 3 => array:2 [ "nombre" => "Yuqi" "apellidos" => "Gao" ] 4 => array:2 [ "nombre" => "Fengxia" "apellidos" => "Lv" ] 5 => array:4 [ "nombre" => "Xiaomei" "apellidos" => "Bie" "email" => array:1 [ 0 => "bxm43@jiau.edu.cn" ] "referencia" => array:1 [ 0 => array:2 [ "etiqueta" => "*" "identificador" => "cor0005" ] ] ] ] "afiliaciones" => array:1 [ 0 => array:2 [ "entidad" => "College of Food Science and Technology, Nanjing Agricultural University, Key Laboratory of Food Processing and Quality Control, Ministry of Agriculture of China, 1 Weigang, Nanjing 210095, PR China" "identificador" => "aff0005" ] ] "correspondencia" => array:1 [ 0 => array:3 [ "identificador" => "cor0005" "etiqueta" => "⁎" "correspondencia" => "Corresponding author." ] ] ] ] "titulosAlternativos" => array:1 [ "es" => array:1 [ "titulo" => "Control de las tasas de aireación y agitación en la producción de surfactina en un cultivo alimentado (fed-batch) en espuma desbordante con fermentación industrial" ] ] "resumenGrafico" => array:2 [ "original" => 0 "multimedia" => array:7 [ "identificador" => "fig0015" "etiqueta" => "Figure 3" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr3.jpeg" "Alto" => 1884 "Ancho" => 3259 "Tamanyo" => 281994 ] ] "descripcion" => array:1 [ "en" => "Effect of agitation and aeration rate on foam overflowing flow rate (up) and surfactin concentration in the overflowing foam (down) at 15h (A and D), 21h (B and E), and 33h (C and F) by Bacillus amyloliquefaciens fmb50 in batch culture." ] ] ] "textoCompleto" => "IntroductionBiosurfactants are amphiphilic molecules widely produced by a variety of microorganisms, which have been considered as an alternative to chemical surfactants. Lipopeptides are one of the major types of biosurfactants<a class="elsevierStyleCrossRef" href="#bib0165">14</a>. As the most effective biosurfactant that has been found so far, surfactin can lower the surface tension of water from 72 to 27mN/m<a class="elsevierStyleCrossRefs" href="#bib0120">5,12</a>. Surfactin exhibits antibacterial and antiviral properties and biodegradability, and appears to be promising for applications in areas such as bioremediation and oil recovery<a class="elsevierStyleCrossRefs" href="#bib0095">1,2,15</a>. However, after almost 50 years, surfactin is not yet a viable alternative to chemical surfactants because of the low yield in bioreactors<a class="elsevierStyleCrossRef" href="#bib0160">13</a>, its relatively high medium cost, and severe foaming in aerated and stirred bioreactors<a class="elsevierStyleCrossRef" href="#bib0175">16</a>. Therefore, the production of surfactin is still limited to laboratory scale<a class="elsevierStyleCrossRef" href="#bib0120">5</a>. To cope with these problems, renewable substrates and foam overflow fermentation were used in recent years<a class="elsevierStyleCrossRefs" href="#bib0115">4,9</a>.Some studies on fed-batch culture of surfactin have been undertaken in the past few years<a class="elsevierStyleCrossRefs" href="#bib0115">4,5,16</a>. In foam overflowing fed-batch culture (FOFC), the flow rates of the feed and of the overflow foam were equal, and thus the broth volume in the bioreactor was kept constant, and continuous enrichment of surfactin in foam overflow was accomplished<a class="elsevierStyleCrossRef" href="#bib0125">6</a>. It has been shown that by controlling the agitation and aeration rates, maximum surfactin productivity could be achieved when the oxygen volumetric mass transfer coefficient (kLa) value was 0.0132/s<a class="elsevierStyleCrossRef" href="#bib0175">16</a>. However, the effect of agitation and aeration rates on surfactin enrichment in the foam has not been reported.In our previous studies, we identified a strain of Bacillus amyloliquefaciens as producer of five surfactin homologues by using high performance liquid chromatography (HPLC) and electrospray ionization mass spectrometry methods<a class="elsevierStyleCrossRef" href="#bib0160">13</a>. A high yield of surfactin from strain B. amyloliquefaciens fmb50 was obtained by genome shuffling, and a method for surfactin determination was established<a class="elsevierStyleCrossRef" href="#bib0180">17</a>.In this work, a semi-defined medium, which is of low cost and high yield compared with the commonly used Landy medium, was initially combined with foam overflow batch culture. The effect of agitation and aeration rates on the foam overflowing rate (fout) and surfactin enrichment in the batch culture process were studied. By using fed-batch culture, a continuous, high concentration enrichment of surfactin could be achieved. This novel fermentation technology has a potential for the industrial application of surfactin production.Materials and methodsMicroorganism and culture mediaB. amyloliquefaciens fmb50 (CGMCC No. 6249), the surfactin producer used in this study, is registered by the China Committee for Culture Collection of Microorganisms.The seed medium (BPY) consisted of: beef extract 5.0g/l, peptone 10.0g/l, yeast extract 5.0g/l, glucose 10.0g/l and NaCl 5.0g/l (pH 7.0).The semi-defined medium (IBM) used for fermentation was optimized by the Taguchi method in our previous work, and consisted of: corn powder 35g/l, ammonium nitrate 15g/l, urea 6g/l, KCl 1.47g/l, NaH2PO4 20mmol/l, MnSO4 0.5mmol/l, MgSO4 0.1mmol/l, CuSO4 12.8μmol/l, FeSO4 1μmol/l and CaCl2 0.5μmol/l (pH 7.0).Flask and bioreactor culture conditionsIn the primary inoculum, a loop of colonies from a fresh potato dextrose agar-slant was transferred into 50ml BPY medium in shaken flasks (250ml) and cultured at 37°C and 180rpm for 12h. For the secondary inoculation, 10ml of the primary culture was inoculated into 200ml BPY medium in shaken flasks<a class="elsevierStyleCrossRef" href="#bib0150">11</a>, and cultured in the same conditions as those in the primary inoculum.Bioreactor cultivation was performed in a 19 l bioreactor containing 12l of medium (L1523, Bioengineering AG, Switzerland); the temperature was controlled at 32°C and the pH was maintained at 7.0 with the automatic addition of 4.0mol/l NaOH. Three control strategies were adopted in batch cultivation to investigate the effects of agitation and aeration rates on fout and surfactin enrichment in foam from overflowing cultures. The agitation and aeration rates were controlled as shown in <a class="elsevierStyleCrossRef" href="#tbl0005">Table 1</a>. In control 1, the agitation and aeration rates were controlled at 200rpm and at 0.66vvm respectively. In control 2, the agitation and aeration rates were controlled at a relatively high level; the agitation rate was 300rpm and the aeration was increased from 1.2 to 2.66vvm as the foam overflowed and the medium volume in the bioreactor was reduced.<elsevierMultimedia ident="tbl0005"></elsevierMultimedia>Analytical methodsThe colony forming units (CFU) were calculated using plate colony-counting methods. Dry cell weight (DCW) was obtained by collecting the fermentation broth with different incubation times in the IBM medium. After centrifugation at 1000×g for 15min, the collected pellets were dried for 8h at 90°C and the dry cells weighed<a class="elsevierStyleCrossRef" href="#bib0135">8</a>. Then, the dry cell weight (DCW) was determined using a pre-determined standard curve relating the number of colony forming units (CFU) to the dry weight: DCW=2.43×log CFU/ml−18.63, with an R2 value of 0.92. Results were represented as CFU per milliliter.Culture samples were precipitated and then extracted with methanol as described by Cooper et al.<a class="elsevierStyleCrossRef" href="#bib0110">3</a> in the extraction of surfactin. Surfactin concentration in crude samples was determined by reverse phase HPLC (U-3000, Dionex, United States) equipped with an Agilent C18 column (4.5mm×250mm, Agilent, United States) and a UV detector. About 20μl of the surfactin sample was injected into the column and then eluted with acetonitrile with 0.1% TFA at a flow rate of 0.84ml/min. Eluent absorbance was monitored at 210nm<a class="elsevierStyleCrossRef" href="#bib0180">17</a>. Quantization was performed based on a standard curve using a standard sample of surfactin (Sigma–Aldrich Co., United Sates). The standard curve equation is: y=17.771x−38.742 (R2=0.999). The equation for the calculation of the specific growth rate (μ/h) was: μ=dX/(X·dt); the equation for the calculation of the specific growth rate (qp/g/(g*h)) was: qp=dP/(X·dt), X: cell yield (g/l); P: product yield (U/ml); t: fermentation time (h).Results and discussionEffect of the control strategy on cell specific growth rateThe effect of agitation and aeration rates on cell growth in the three control strategies adopted in this study in batch culture is shown in <a class="elsevierStyleCrossRef" href="#fig0005">Figure 1</a>. In controls 1 and 2, B. amyloliquefaciens fmb50 in the bioreactor reached up to 109CFU/ml after 15±0.2h culture, and then the cell growth went into stationary phase (<a class="elsevierStyleCrossRef" href="#fig0005">Fig. 1</a>A). However, in control 3, cell growth declined from 12h of cultivation. That may be due to the agitation rate changing between 150 and 250rpm; normal cell growth seemed to be inhibited and the biomass decreased due to the simultaneous increase of the overflow. B. amyloliquefaciens fmb50 reached its maximum specific cell growth rate at 6h of cultivation in control 1, which was 4h earlier than in control 2 (<a class="elsevierStyleCrossRef" href="#fig0005">Fig. 1</a>B). In control 3, where cell growth was disturbed, a lower specific cell growth rate was observed. The maximum specific cell growth rate was lowered nearly by half with respect to controls 1 and 2. Although the increase in the agitation rate promoted quadratic cell growth, this one was not delayed.<elsevierMultimedia ident="fig0005"></elsevierMultimedia>Surfactin production is essentially associated with cell growth<a class="elsevierStyleCrossRefs" href="#bib0130">7,16</a>, although high biomass does not necessarily mean high surfactin production. It seems that a lower specific growth rate is more conducive to the production of biosurfactants such as mycosubtilin<a class="elsevierStyleCrossRef" href="#bib0115">4</a>. The effect of agitation and aeration rates on cell growth and cell specific growth rate was studied in this paper and the best control strategy was chosen to enhance surfactin production.Effect of the control strategy on fout and the concentration and production of enriched surfactin in foamFoam began to overflow outside the bioreactor after 6h of cultivation (<a class="elsevierStyleCrossRef" href="#fig0010">Fig. 2</a>A). The maximum fout in control 1 was 0.3l/h while in control 2 it was 1.3l/h, which indicated that the foam overflow was significantly affected by the agitation and aeration rates used in different strategies. The surfactin concentration in the overflow foam also changed with time. As shown in <a class="elsevierStyleCrossRef" href="#fig0010">Figure 2</a>B, different trends were observed in the three control strategies. In control 3, surfactin concentration increased linearly and showed the highest specific production rate, reaching a maximum of 0.426l/h (<a class="elsevierStyleCrossRef" href="#fig0010">Fig. 2</a>C).<elsevierMultimedia ident="fig0010"></elsevierMultimedia>The linear increase of surfactin concentration and the relatively high specific production rate in control 3 (<a class="elsevierStyleCrossRef" href="#fig0010">Fig. 2</a>B and C) indicated that high surfactin enrichment in foam may be achieved by a combined control of the agitation and aeration rates at a reasonable level. As shown in <a class="elsevierStyleCrossRef" href="#fig0010">Figure 2</a>B, the maximum surfactin concentration in foam reached 3.342g/ml and the maximum surfactin production in foam reached 2.525g (approximately overflowing 7.6l foam) in control 3. However, maximum surfactin concentration and maximum surfactin production were 1.8g/ml and 8.46g (approximately overflowing 4.7l of foam) in control 1, respectively. These results in control 3, which show the linear increase of surfactin concentration and the relatively high specific production rate, are similar to those in the study by Davis et al.<a class="elsevierStyleCrossRef" href="#bib0125">6</a>, the enrichment of surfactin in the foam was enhanced to 0.44g/l with the time going by supplementing culture broth in the stationary phase<a class="elsevierStyleCrossRef" href="#bib0125">6</a>. Foam stopped overflowing at 22h in control 1 and at 26h in controls 2 and 3. This was partly due to the medium level in the bioreactor dropping as overflowing foam: however; the surfactin in the foam was thus separated from the broth almost completely<a class="elsevierStyleCrossRef" href="#bib0125">6</a>. Apparently, continuous surfactin production in the bioreactor ensured an increasing trend of surfactin concentration in the foam. With the agitation and aeration rate controlled under different strategies, the surfactin production presented a different kinetic trend. Control 3, under which fout was below 0.7l/h, indicated that surfactin enrichment may be improved through stir speed and aeration control.Influence of agitation and aeration rates on fout and concentration of enriched surfactinThe influence of agitation and aeration rates on fout and surfactin enrichment in the overflowing foam were investigated in batch culture experiments conducted using a series of combinations of aeration and agitation rates. Foam overflow began after 6h of cultivation (<a class="elsevierStyleCrossRef" href="#fig0010">Fig. 2</a>A). As indicated in <a class="elsevierStyleCrossRef" href="#fig0015">Fig. 3</a>(A–C), fout tended to increase with an increased aeration rate from 1 to 1.5vvm and agitation rate from 150 to 250rpm. At 33h, fout was extremely low, only a maximum 0.14l per hour (<a class="elsevierStyleCrossRef" href="#fig0015">Fig. 3</a>C); however, raising the aeration rate to 250rpm, the fout could increase to a maximum 1.76 per hour.<elsevierMultimedia ident="fig0015"></elsevierMultimedia>Agitation and aeration rates significantly affected the concentration of surfactin in the foam. In the early stage of fermentation (i.e. at 15h; <a class="elsevierStyleCrossRef" href="#fig0015">Fig. 3</a>A), an increase in agitation rate increased fout; however, the surfactin concentration was at a relatively low level and decreased from 1.6g/l to 1g/l with the agitation increase. At 21h, the maximum surfactin concentration of 4.7g/l occurred at 150rpm agitation and 1vvm aeration. With the increase of agitation to 250rpm, the surfactin concentration was reduced to 1.5g/l. Interestingly, the change in the agitation rate had no effect on the surfactin concentration at 1.5vvm aeration (<a class="elsevierStyleCrossRef" href="#fig0015">Fig. 3</a>E). At 33h, the surfactin concentration in the foam was maintained at >2.5g/l with an aeration rate of 1.5vvm and it did not drop with an increasing agitation rate. However, under the condition of aeration rate at 1vvm, the surfactin concentration in the foam was 2.5g/l only with the agitation rate of 250rpm. Apparently, the agitation and aeration rates significantly affected the concentration of surfactin enriched in the foam.The agitation and aeration rates in the aerobic fermentation greatly affected the foam formation rate and the surfactin concentration in the foam. Similar to our case, a previous study observed that a higher stirring speed or aeration rate led to a higher fout<a class="elsevierStyleCrossRef" href="#bib0125">6</a>. However, surfactin enrichment in this study presented a different kinetic relationship with fout compared to that in the basic study by Davis et al.<a class="elsevierStyleCrossRef" href="#bib0125">6</a>, in which the foam continuously overflowed under a constant agitation rate. In the late stages of batch FOFC, such as at 33h (<a class="elsevierStyleCrossRef" href="#fig0015">Fig. 3</a>F), the surfactin concentration in the foam was lower than at an earlier stage (21h), because > 50% of the culture broth in the bioreactor was gone. Thus, the broth level in the bioreactor was too low to maintain the overflowing foam and further foam formation was also limited because almost all the surfactin had already been drained away in the foam. Both the surfactin concentration in the foam and fout were determined by the surfactin produced in the broth. Furthermore, the fout level affected the surfactin enrichment, and at the same time, fout was influenced by the agitation and aeration rates.Fed-batch culture with controlled agitation and aeration ratesA fed-batch culture process was also adopted in this study. <a class="elsevierStyleCrossRef" href="#fig0020">Figure 4</a> shows that foam began to overflow after 6h of cultivation. Afterwards, fout gradually dropped from 0.6l/h to 0.2l/h by real-time control of the agitation and aeration rates and was maintained at this rate. From 15h, the IBM medium was fed into the bioreactor at a 0.2l/h feed rate (matching fout). In this culture, the biomass was kept above 109CFU/ml. Foam continued to overflow until the cultivation terminated and surfactin was enriched in the foam with a concentration >4g/l; the maximum concentration reached 4.7g/l. At the end of the fermentation, the surfactin concentration in the culture broth in the bioreactor was very low (data not shown), which means that almost all the surfactin was transferred into the foam.<elsevierMultimedia ident="fig0020"></elsevierMultimedia>In other studies of foam overflowing cultures, although the product was enriched and separated in foam, the foam overflowed spontaneously, and thus the surfactin concentration in it fluctuated<a class="elsevierStyleCrossRefs" href="#bib0125">6,10,16</a>. For this reason, surfactin in the foam was not enriched to a very high level; in previous studies, in which surfactin was produced by potato process effluent, it was 1.67g/l and 0.9g/l<a class="elsevierStyleCrossRefs" href="#bib0125">6,10</a>. Thus, controlled foam overflow in this work achieved almost 3 and 4 times the surfactin concentration compared with the previous studies, respectively<a class="elsevierStyleCrossRefs" href="#bib0125">6,10</a>. Furthermore, the medium feeding in our study overcame disruption of cell growth, which ensured that surfactin was continuously produced. Combined with control of fout at a low level, the surfactin concentration in the foam was maintained above 4g/l, which will significantly reduce production costs and improve industrial production capacity.ConclusionsOur study focused on the control of agitation and aeration rates in foam overflowing fermentation to improve surfactin enrichment and continuous high-level production. In batch mode, the agitation and aeration rates were found to have a close relationship with cell growth and surfactin production. In further research, our study revealed that fout does not always negatively impact on the surfactin concentration in the foam. The broth level in the bioreactor and the surfactin residue in the broth also affect both fout and the surfactin enrichment of the foam.Through feeding the medium into the bioreactor, a fed-batch fermentation process was successfully established in which foam overflowed at a controlled flow rate of 0.2l/h, and surfactin in the enriched foam was kept at a level above 4g/l.Ethical disclosuresProtection of human and animal subjectsThe authors declare that no experiments were performed on humans or animals for this study.Confidentiality of dataThe authors declare that no patient data appear in this article.Right to privacy and informed consentThe authors declare that no patient data appear in this article.Conflict of interestThe authors declare that they have no conflicts of interest." "textoCompletoSecciones" => array:1 [ "secciones" => array:13 [ 0 => array:3 [ "identificador" => "xres602698" "titulo" => "Abstract" "secciones" => array:1 [ 0 => array:1 [ "identificador" => "abst0005" ] ] ] 1 => array:2 [ "identificador" => "xpalclavsec616753" "titulo" => "Keywords" ] 2 => array:3 [ "identificador" => "xres602697" "titulo" => "Resumen" "secciones" => array:1 [ 0 => array:1 [ "identificador" => "abst0010" ] ] ] 3 => array:2 [ "identificador" => "xpalclavsec616754" "titulo" => "Palabras clave" ] 4 => array:2 [ "identificador" => "sec0005" "titulo" => "Introduction" ] 5 => array:3 [ "identificador" => "sec0010" "titulo" => "Materials and methods" "secciones" => array:2 [ 0 => array:2 [ "identificador" => "sec0015" "titulo" => "Microorganism and culture media" ] 1 => array:2 [ "identificador" => "sec0020" "titulo" => "Flask and bioreactor culture conditions" ] ] ] 6 => array:2 [ "identificador" => "sec0025" "titulo" => "Analytical methods" ] 7 => array:3 [ "identificador" => "sec0030" "titulo" => "Results and discussion" "secciones" => array:4 [ 0 => array:2 [ "identificador" => "sec0035" "titulo" => "Effect of the control strategy on cell specific growth rate" ] 1 => array:2 [ "identificador" => "sec0040" "titulo" => "Effect of the control strategy on f and the concentration and production of enriched surfactin in foam" ] 2 => array:2 [ "identificador" => "sec0045" "titulo" => "Influence of agitation and aeration rates on f and concentration of enriched surfactin" ] 3 => array:2 [ "identificador" => "sec0050" "titulo" => "Fed-batch culture with controlled agitation and aeration rates" ] ] ] 8 => array:2 [ "identificador" => "sec0055" "titulo" => "Conclusions" ] 9 => array:3 [ "identificador" => "sec0060" "titulo" => "Ethical disclosures" "secciones" => array:3 [ 0 => array:2 [ "identificador" => "sec0065" "titulo" => "Protection of human and animal subjects" ] 1 => array:2 [ "identificador" => "sec0070" "titulo" => "Confidentiality of data" ] 2 => array:2 [ "identificador" => "sec0075" "titulo" => "Right to privacy and informed consent" ] ] ] 10 => array:2 [ "identificador" => "sec0085" "titulo" => "Conflict of interest" ] 11 => array:2 [ "identificador" => "xack202833" "titulo" => "Acknowledgements" ] 12 => array:1 [ "titulo" => "References" ] ] ] "pdfFichero" => "main.pdf" "tienePdf" => true "fechaRecibido" => "2014-12-04" "fechaAceptado" => "2015-09-07" "PalabrasClave" => array:2 [ "en" => array:1 [ 0 => array:4 [ "clase" => "keyword" "titulo" => "Keywords" "identificador" => "xpalclavsec616753" "palabras" => array:6 [ 0 => "Surfactin" 1 => "Agitation rate" 2 => "Aeration rate" 3 => "Foam overflowing" 4 => "Fed-batch culture" 5 => "Industrial fermentation" ] ] ] "es" => array:1 [ 0 => array:4 [ "clase" => "keyword" "titulo" => "Palabras clave" "identificador" => "xpalclavsec616754" "palabras" => array:6 [ 0 => "Surfactina" 1 => "Tasa de agitación" 2 => "Tasa de aireación" 3 => "Espuma desbordante" 4 => "Cultivo fed-batch" 5 => "Fermentación industrial" ] ] ] ] "tieneResumen" => true "resumen" => array:2 [ "en" => array:2 [ "titulo" => "Abstract" "resumen" => "Bacillus amyloliquefaciens fmb50 produces a high yield of surfactin, a lipopeptide-type biosurfactant that has been widely studied and has potential applications in many fields. A foam overflowing culture has been successfully used in the combined production-enrichment fermentation of surfactin. In this study, the agitation and aeration rates were found to have relationships with foam formation and surfactin enrichment. A maximum surfactin concentration of 4.7g/l of foam was obtained after 21h of culture with an agitation rate of 150rpm and an aeration rate of 1vvm in fed-batch culture. By controlling the foam overflow rate (fout) of a fed-batch culture, surfactin concentration in the foam was continuously maintained above 4g/l." ] "es" => array:2 [ "titulo" => "Resumen" "resumen" => "Bacillus amyloliquefaciens fmb50 produce gran cantidad de surfactina, un biosurfactante de tipo lipopeptídico que ha sido objeto de estudios pormenorizados y tiene aplicaciones en muchos campos. El cultivo en espuma desbordante se ha utilizado con éxito en la fermentación combinada de producción-enriquecimiento de surfactina. En este estudio, se halló que las tasas de aireación y agitación tienen relación con la formación de espuma y el enriquecimiento de la surfactina. Se obtuvo una concentración máxima de surfactina de 4,7g/l de espuma después de 21 h de cultivo con una tasa de agitación de 150 rpm y una tasa de aireación de 1 vvm en un cultivo alimentado (fed-batch). Al controlar la tasa de espuma desbordante (fout) de un cultivo fed-batch, la concentración de surfactina en la espuma se mantuvo continua por encima de 4 g/l." ] ] "multimedia" => array:5 [ 0 => array:7 [ "identificador" => "fig0005" "etiqueta" => "Figure 1" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr1.jpeg" "Alto" => 1033 "Ancho" => 2670 "Tamanyo" => 112598 ] ] "descripcion" => array:1 [ "en" => "Time courses of CFU (A) and cell specific growth rate (B) by Bacillus amyloliquefaciens fmb50 in batch culture with three control strategies. In control 1, the agitation rate was 200rpm, and the aeration rate was 0.66vvm; in control 2, the agitation rate was 300rpm, and the aeration rate increased from 1.2 to 2.66vvm; in control 3, the agitation rate changed between 150 and 250rpm to keep the overflowing flow rate at a relatively moderate level, and the aeration rate was kept at 1.8vvm." ] ] 1 => array:7 [ "identificador" => "fig0010" "etiqueta" => "Figure 2" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr2.jpeg" "Alto" => 848 "Ancho" => 3168 "Tamanyo" => 146949 ] ] "descripcion" => array:1 [ "en" => "Time courses of foam overflowing flow rate (A), surfactin concentration in the foam (B) and specific production rate (C) in batch culture of Bacillus amyloliquefaciens fmb50 with three control strategies. In control 1, the agitation rate was 200rpm, and the aeration rate was 0.66vvm; in control 2, the agitation rate was 300rpm, and the aeration rate increased from 1.2 to 2.66vvm; in control 3, the agitation rate was changed between 150 and 250rpm to keep fout at a relatively moderate level, and the aeration rate was kept at 1.8vvm." ] ] 2 => array:7 [ "identificador" => "fig0015" "etiqueta" => "Figure 3" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr3.jpeg" "Alto" => 1884 "Ancho" => 3259 "Tamanyo" => 281994 ] ] "descripcion" => array:1 [ "en" => "Effect of agitation and aeration rate on foam overflowing flow rate (up) and surfactin concentration in the overflowing foam (down) at 15h (A and D), 21h (B and E), and 33h (C and F) by Bacillus amyloliquefaciens fmb50 in batch culture." ] ] 3 => array:7 [ "identificador" => "fig0020" "etiqueta" => "Figure 4" "tipo" => "MULTIMEDIAFIGURA" "mostrarFloat" => true "mostrarDisplay" => false "figura" => array:1 [ 0 => array:4 [ "imagen" => "gr4.jpeg" "Alto" => 1270 "Ancho" => 2166 "Tamanyo" => 126327 ] ] "descripcion" => array:1 [ "en" => "Time course profiles of CFU, surfactin concentration in foam, and foam overflowing flow rate (fout) during fed-batch fermentation of Bacillus amyloliquefaciens fmb50 under controlled agitation and aeration rate to keep fout constant." ] ] 4 => array:7 [ "identificador" => "tbl0005" "etiqueta" => "Table 1" "tipo" => "MULTIMEDIATABLA" "mostrarFloat" => true "mostrarDisplay" => false "tabla" => array:1 [ "tablatextoimagen" => array:1 [ 0 => array:2 [ "tabla" => array:1 [ 0 => """ <table border="0" frame="\n \t\t\t\t\tvoid\n \t\t\t\t" class=""><thead title="thead"><tr title="table-row"><th class="td" title="table-head " align="left" valign="top" scope="col" style="border-bottom: 2px solid black">Control \t\t\t\t\t\t\n \t\t\t\t</th><th class="td" title="table-head " align="left" valign="top" scope="col" style="border-bottom: 2px solid black">Agitation rate (rpm) \t\t\t\t\t\t\n \t\t\t\t</th><th class="td" title="table-head " align="left" valign="top" scope="col" style="border-bottom: 2px solid black">Aeration (vvm) \t\t\t\t\t\t\n \t\t\t\t</th></tr></thead><tbody title="tbody"><tr title="table-row"><td class="td-with-role" title="table-entry ; entry_with_role_rowhead " align="left" valign="top">1 \t\t\t\t\t\t\n \t\t\t\t</td><td class="td" title="table-entry " align="char" valign="top">200 \t\t\t\t\t\t\n \t\t\t\t</td><td class="td" title="table-entry " align="char" valign="top">0.66 \t\t\t\t\t\t\n \t\t\t\t</td></tr><tr title="table-row"><td class="td-with-role" title="table-entry ; entry_with_role_rowhead " align="left" valign="top">2 \t\t\t\t\t\t\n \t\t\t\t</td><td class="td" title="table-entry " align="char" valign="top">300 \t\t\t\t\t\t\n \t\t\t\t</td><td class="td" title="table-entry " align="char" valign="top">1.2–2.66 \t\t\t\t\t\t\n \t\t\t\t</td></tr><tr title="table-row"><td class="td-with-role" title="table-entry ; entry_with_role_rowhead " align="left" valign="top">3 \t\t\t\t\t\t\n \t\t\t\t</td><td class="td" title="table-entry " align="char" valign="top">150–250 \t\t\t\t\t\t\n \t\t\t\t</td><td class="td" title="table-entry " align="char" valign="top">1.8 \t\t\t\t\t\t\n \t\t\t\t</td></tr></tbody></table> """ ] "imagenFichero" => array:1 [ 0 => "xTab986780.png" ] ] ] ] "descripcion" => array:1 [ "en" => "Control strategy of agitation and aeration rates." ] ] ] "bibliografia" => array:2 [ "titulo" => "References" "seccion" => array:1 [ 0 => array:2 [ "identificador" => "bibs0005" "bibliografiaReferencia" => array:17 [ 0 => array:3 [ "identificador" => "bib0095" "etiqueta" => "1" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Surfactin, a crystalline peptidelipid surfactant produced by Bacillus subtilis: isolation, characterization and its inhibition of fibrin clot formation" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:3 [ 0 => "K. 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Chang" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:5 [ "tituloSerie" => "Process Biochem" "fecha" => "2006" "volumen" => "41" "paginaInicial" => "1799" "paginaFinal" => "1805" ] ] ] ] ] ] 16 => array:3 [ "identificador" => "bib0180" "etiqueta" => "17" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Genome shuffling of Bacillus amyloliquefaciens for improving antimicrobial lipopeptide production and an analysis of relative gene expression using FQ RT-PCR" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:8 [ 0 => "J.F. Zhao" 1 => "Y.H. Li" 2 => "C. Zhang" 3 => "Z. Yao" 4 => "L. Zhang" 5 => "X.M. Bie" 6 => "F.X. Lu" 7 => "Z.X. Lu" ] ] ] ] ] "host" => array:1 [ 0 => array:2 [ "doi" => "10.1007/s10295-012-1098-9" "Revista" => array:6 [ "tituloSerie" => "J Ind Microbiol Biotechnol" "fecha" => "2012" "volumen" => "39" "paginaInicial" => "889" "paginaFinal" => "896" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/22350068" "web" => "Medline" ] ] ] ] ] ] ] ] ] ] ] ] "agradecimientos" => array:1 [ 0 => array:4 [ "identificador" => "xack202833" "titulo" => "Acknowledgements" "texto" => "This work was supported by grants from the National Natural Science Foundation of China (grant no. 31271828) and the National Science and Technology Support program (grant no. 2011BAD23B05)." 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Control de las tasas de aireación y agitación en la producción de surfactina en un cultivo alimentado (fed-batch) en espuma desbordante con fermentación industrial

Shulin Yao, Shengming Zhao, Zhaoxin Lu, Yuqi Gao, Fengxia Lv, Xiaomei Bie

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bxm43@jiau.edu.cn

Corresponding author.

College of Food Science and Technology, Nanjing Agricultural University, Key Laboratory of Food Processing and Quality Control, Ministry of Agriculture of China, 1 Weigang, Nanjing 210095, PR China

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However&#44; after almost 50 years&#44; surfactin is not yet a viable alternative to chemical surfactants because of the low yield in bioreactors<a class="elsevierStyleCrossRef" href="#bib0160"><span class="elsevierStyleSup">13</span></a>&#44; its relatively high medium cost&#44; and severe foaming in aerated and stirred bioreactors<a class="elsevierStyleCrossRef" href="#bib0175"><span class="elsevierStyleSup">16</span></a>&#46; Therefore&#44; the production of surfactin is still limited to laboratory scale<a class="elsevierStyleCrossRef" href="#bib0120"><span class="elsevierStyleSup">5</span></a>&#46; To cope with these problems&#44; renewable substrates and foam overflow fermentation were used in recent years<a class="elsevierStyleCrossRefs" href="#bib0115"><span class="elsevierStyleSup">4&#44;9</span></a>&#46;</p><p id="par0010" class="elsevierStylePara elsevierViewall">Some studies on fed-batch culture of surfactin have been undertaken in the past few years<a class="elsevierStyleCrossRefs" href="#bib0115"><span class="elsevierStyleSup">4&#44;5&#44;16</span></a>&#46; In foam overflowing fed-batch culture &#40;FOFC&#41;&#44; the flow rates of the feed and of the overflow foam were equal&#44; and thus the broth volume in the bioreactor was kept constant&#44; and continuous enrichment of surfactin in foam overflow was accomplished<a class="elsevierStyleCrossRef" href="#bib0125"><span class="elsevierStyleSup">6</span></a>&#46; It has been shown that by controlling the agitation and aeration rates&#44; maximum surfactin productivity could be achieved when the oxygen volumetric mass transfer coefficient &#40;<span class="elsevierStyleItalic">k</span><span class="elsevierStyleInf">L</span>a&#41; value was 0&#46;0132&#47;s<a class="elsevierStyleCrossRef" href="#bib0175"><span class="elsevierStyleSup">16</span></a>&#46; However&#44; the effect of agitation and aeration rates on surfactin enrichment in the foam has not been reported&#46;</p><p id="par0015" class="elsevierStylePara elsevierViewall">In our previous studies&#44; we identified a strain of <span class="elsevierStyleItalic">Bacillus amyloliquefaciens</span> as producer of five surfactin homologues by using high performance liquid chromatography &#40;HPLC&#41; and electrospray ionization mass spectrometry methods<a class="elsevierStyleCrossRef" href="#bib0160"><span class="elsevierStyleSup">13</span></a>&#46; A high yield of surfactin from strain <span class="elsevierStyleItalic">B&#46; amyloliquefaciens</span> fmb50 was obtained by genome shuffling&#44; and a method for surfactin determination was established<a class="elsevierStyleCrossRef" href="#bib0180"><span class="elsevierStyleSup">17</span></a>&#46;</p><p id="par0020" class="elsevierStylePara elsevierViewall">In this work&#44; a semi-defined medium&#44; which is of low cost and high yield compared with the commonly used Landy medium&#44; was initially combined with foam overflow batch culture&#46; The effect of agitation and aeration rates on the foam overflowing rate &#40;<span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span>&#41; and surfactin enrichment in the batch culture process were studied&#46; By using fed-batch culture&#44; a continuous&#44; high concentration enrichment of surfactin could be achieved&#46; This novel fermentation technology has a potential for the industrial application of surfactin production&#46;</p></span><span id="sec0010" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0030">Materials and methods</span><span id="sec0015" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0035">Microorganism and culture media</span><p id="par0025" class="elsevierStylePara elsevierViewall"><span class="elsevierStyleItalic">B&#46; amyloliquefaciens</span> fmb50 &#40;CGMCC No&#46; 6249&#41;&#44; the surfactin producer used in this study&#44; is registered by the China Committee for Culture Collection of Microorganisms&#46;</p><p id="par0030" class="elsevierStylePara elsevierViewall">The seed medium &#40;BPY&#41; consisted of&#58; beef extract 5&#46;0<span class="elsevierStyleHsp" style=""></span>g&#47;l&#44; peptone 10&#46;0<span class="elsevierStyleHsp" style=""></span>g&#47;l&#44; yeast extract 5&#46;0<span class="elsevierStyleHsp" style=""></span>g&#47;l&#44; glucose 10&#46;0<span class="elsevierStyleHsp" style=""></span>g&#47;l and NaCl 5&#46;0<span class="elsevierStyleHsp" style=""></span>g&#47;l &#40;pH 7&#46;0&#41;&#46;</p><p id="par0035" class="elsevierStylePara elsevierViewall">The semi-defined medium &#40;IBM&#41; used for fermentation was optimized by the Taguchi method in our previous work&#44; and consisted of&#58; corn powder 35<span class="elsevierStyleHsp" style=""></span>g&#47;l&#44; ammonium nitrate 15<span class="elsevierStyleHsp" style=""></span>g&#47;l&#44; urea 6<span class="elsevierStyleHsp" style=""></span>g&#47;l&#44; KCl 1&#46;47<span class="elsevierStyleHsp" style=""></span>g&#47;l&#44; NaH<span class="elsevierStyleInf">2</span>PO<span class="elsevierStyleInf">4</span> 20<span class="elsevierStyleHsp" style=""></span>mmol&#47;l&#44; MnSO<span class="elsevierStyleInf">4</span> 0&#46;5<span class="elsevierStyleHsp" style=""></span>mmol&#47;l&#44; MgSO<span class="elsevierStyleInf">4</span> 0&#46;1<span class="elsevierStyleHsp" style=""></span>mmol&#47;l&#44; CuSO<span class="elsevierStyleInf">4</span> 12&#46;8<span class="elsevierStyleHsp" style=""></span>&#956;mol&#47;l&#44; FeSO<span class="elsevierStyleInf">4</span> 1<span class="elsevierStyleHsp" style=""></span>&#956;mol&#47;l and CaCl<span class="elsevierStyleInf">2</span> 0&#46;5<span class="elsevierStyleHsp" style=""></span>&#956;mol&#47;l &#40;pH 7&#46;0&#41;&#46;</p></span><span id="sec0020" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0040">Flask and bioreactor culture conditions</span><p id="par0040" class="elsevierStylePara elsevierViewall">In the primary inoculum&#44; a loop of colonies from a fresh potato dextrose agar-slant was transferred into 50<span class="elsevierStyleHsp" style=""></span>ml BPY medium in shaken flasks &#40;250<span class="elsevierStyleHsp" style=""></span>ml&#41; and cultured at 37<span class="elsevierStyleHsp" style=""></span>&#176;C and 180<span class="elsevierStyleHsp" style=""></span>rpm for 12<span class="elsevierStyleHsp" style=""></span>h&#46; For the secondary inoculation&#44; 10<span class="elsevierStyleHsp" style=""></span>ml of the primary culture was inoculated into 200<span class="elsevierStyleHsp" style=""></span>ml BPY medium in shaken flasks<a class="elsevierStyleCrossRef" href="#bib0150"><span class="elsevierStyleSup">11</span></a>&#44; and cultured in the same conditions as those in the primary inoculum&#46;</p><p id="par0045" class="elsevierStylePara elsevierViewall">Bioreactor cultivation was performed in a 19 l bioreactor containing 12<span class="elsevierStyleHsp" style=""></span>l of medium &#40;L1523&#44; Bioengineering AG&#44; Switzerland&#41;&#59; the temperature was controlled at 32<span class="elsevierStyleHsp" style=""></span>&#176;C and the pH was maintained at 7&#46;0 with the automatic addition of 4&#46;0<span class="elsevierStyleHsp" style=""></span>mol&#47;l NaOH&#46; Three control strategies were adopted in batch cultivation to investigate the effects of agitation and aeration rates on <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> and surfactin enrichment in foam from overflowing cultures&#46; The agitation and aeration rates were controlled as shown in <a class="elsevierStyleCrossRef" href="#tbl0005">Table 1</a>&#46; In control 1&#44; the agitation and aeration rates were controlled at 200<span class="elsevierStyleHsp" style=""></span>rpm and at 0&#46;66<span class="elsevierStyleHsp" style=""></span>vvm respectively&#46; In control 2&#44; the agitation and aeration rates were controlled at a relatively high level&#59; the agitation rate was 300<span class="elsevierStyleHsp" style=""></span>rpm and the aeration was increased from 1&#46;2 to 2&#46;66<span class="elsevierStyleHsp" style=""></span>vvm as the foam overflowed and the medium volume in the bioreactor was reduced&#46;</p><elsevierMultimedia ident="tbl0005"></elsevierMultimedia></span></span><span id="sec0025" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0045">Analytical methods</span><p id="par0050" class="elsevierStylePara elsevierViewall">The colony forming units &#40;CFU&#41; were calculated using plate colony-counting methods&#46; Dry cell weight &#40;DCW&#41; was obtained by collecting the fermentation broth with different incubation times in the IBM medium&#46; After centrifugation at 1000<span class="elsevierStyleHsp" style=""></span>&#215;<span class="elsevierStyleHsp" style=""></span><span class="elsevierStyleItalic">g</span> for 15<span class="elsevierStyleHsp" style=""></span>min&#44; the collected pellets were dried for 8<span class="elsevierStyleHsp" style=""></span>h at 90<span class="elsevierStyleHsp" style=""></span>&#176;C and the dry cells weighed<a class="elsevierStyleCrossRef" href="#bib0135"><span class="elsevierStyleSup">8</span></a>&#46; Then&#44; the dry cell weight &#40;DCW&#41; was determined using a pre-determined standard curve relating the number of colony forming units &#40;CFU&#41; to the dry weight&#58; DCW<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>2&#46;43<span class="elsevierStyleHsp" style=""></span>&#215;<span class="elsevierStyleHsp" style=""></span>log CFU&#47;ml<span class="elsevierStyleHsp" style=""></span>&#8722;<span class="elsevierStyleHsp" style=""></span>18&#46;63&#44; with an <span class="elsevierStyleItalic">R</span><span class="elsevierStyleSup">2</span> value of 0&#46;92&#46; Results were represented as CFU per milliliter&#46;</p><p id="par0055" class="elsevierStylePara elsevierViewall">Culture samples were precipitated and then extracted with methanol as described by Cooper <span class="elsevierStyleItalic">et al</span>&#46;<a class="elsevierStyleCrossRef" href="#bib0110"><span class="elsevierStyleSup">3</span></a> in the extraction of surfactin&#46; Surfactin concentration in crude samples was determined by reverse phase HPLC &#40;U-3000&#44; Dionex&#44; United States&#41; equipped with an Agilent C18 column &#40;4&#46;5<span class="elsevierStyleHsp" style=""></span>mm<span class="elsevierStyleHsp" style=""></span>&#215;<span class="elsevierStyleHsp" style=""></span>250<span class="elsevierStyleHsp" style=""></span>mm&#44; Agilent&#44; United States&#41; and a UV detector&#46; About 20<span class="elsevierStyleHsp" style=""></span>&#956;l of the surfactin sample was injected into the column and then eluted with acetonitrile with 0&#46;1<span class="elsevierStyleHsp" style=""></span>&#37; TFA at a flow rate of 0&#46;84<span class="elsevierStyleHsp" style=""></span>ml&#47;min&#46; Eluent absorbance was monitored at 210<span class="elsevierStyleHsp" style=""></span>nm<a class="elsevierStyleCrossRef" href="#bib0180"><span class="elsevierStyleSup">17</span></a>&#46; Quantization was performed based on a standard curve using a standard sample of surfactin &#40;Sigma&#8211;Aldrich Co&#46;&#44; United Sates&#41;&#46; The standard curve equation is&#58; <span class="elsevierStyleItalic">y</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>17&#46;771<span class="elsevierStyleItalic">x</span><span class="elsevierStyleHsp" style=""></span>&#8722;<span class="elsevierStyleHsp" style=""></span>38&#46;742 &#40;<span class="elsevierStyleItalic">R</span><span class="elsevierStyleSup">2</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;999&#41;&#46; The equation for the calculation of the specific growth rate &#40;<span class="elsevierStyleItalic">&#956;</span>&#47;<span class="elsevierStyleItalic">h</span>&#41; was&#58; <span class="elsevierStyleItalic">&#956;</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>d<span class="elsevierStyleItalic">X</span>&#47;&#40;<span class="elsevierStyleItalic">X</span>&#183;d<span class="elsevierStyleItalic">t</span>&#41;&#59; the equation for the calculation of the specific growth rate &#40;<span class="elsevierStyleItalic">qp</span>&#47;<span class="elsevierStyleItalic">g</span>&#47;&#40;<span class="elsevierStyleItalic">g</span>&#42;<span class="elsevierStyleItalic">h</span>&#41;&#41; was&#58; qp<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>dP&#47;&#40;<span class="elsevierStyleItalic">X</span>&#183;d<span class="elsevierStyleItalic">t</span>&#41;&#44; <span class="elsevierStyleItalic">X</span>&#58; cell yield &#40;g&#47;l&#41;&#59; <span class="elsevierStyleItalic">P</span>&#58; product yield &#40;U&#47;ml&#41;&#59; <span class="elsevierStyleItalic">t</span>&#58; fermentation time &#40;h&#41;&#46;</p></span><span id="sec0030" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0050">Results and discussion</span><span id="sec0035" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0055">Effect of the control strategy on cell specific growth rate</span><p id="par0060" class="elsevierStylePara elsevierViewall">The effect of agitation and aeration rates on cell growth in the three control strategies adopted in this study in batch culture is shown in <a class="elsevierStyleCrossRef" href="#fig0005">Figure 1</a>&#46; In controls 1 and 2&#44; <span class="elsevierStyleItalic">B&#46; amyloliquefaciens</span> fmb50 in the bioreactor reached up to 10<span class="elsevierStyleSup">9</span><span class="elsevierStyleHsp" style=""></span>CFU&#47;ml after 15<span class="elsevierStyleHsp" style=""></span>&#177;<span class="elsevierStyleHsp" style=""></span>0&#46;2<span class="elsevierStyleHsp" style=""></span>h culture&#44; and then the cell growth went into stationary phase &#40;<a class="elsevierStyleCrossRef" href="#fig0005">Fig&#46; 1</a>A&#41;&#46; However&#44; in control 3&#44; cell growth declined from 12<span class="elsevierStyleHsp" style=""></span>h of cultivation&#46; That may be due to the agitation rate changing between 150 and 250<span class="elsevierStyleHsp" style=""></span>rpm&#59; normal cell growth seemed to be inhibited and the biomass decreased due to the simultaneous increase of the overflow&#46; <span class="elsevierStyleItalic">B&#46; amyloliquefaciens</span> fmb50 reached its maximum specific cell growth rate at 6<span class="elsevierStyleHsp" style=""></span>h of cultivation in control 1&#44; which was 4<span class="elsevierStyleHsp" style=""></span>h earlier than in control 2 &#40;<a class="elsevierStyleCrossRef" href="#fig0005">Fig&#46; 1</a>B&#41;&#46; In control 3&#44; where cell growth was disturbed&#44; a lower specific cell growth rate was observed&#46; The maximum specific cell growth rate was lowered nearly by half with respect to controls 1 and 2&#46; Although the increase in the agitation rate promoted quadratic cell growth&#44; this one was not delayed&#46;</p><elsevierMultimedia ident="fig0005"></elsevierMultimedia><p id="par0065" class="elsevierStylePara elsevierViewall">Surfactin production is essentially associated with cell growth<a class="elsevierStyleCrossRefs" href="#bib0130"><span class="elsevierStyleSup">7&#44;16</span></a>&#44; although high biomass does not necessarily mean high surfactin production&#46; It seems that a lower specific growth rate is more conducive to the production of biosurfactants such as mycosubtilin<a class="elsevierStyleCrossRef" href="#bib0115"><span class="elsevierStyleSup">4</span></a>&#46; The effect of agitation and aeration rates on cell growth and cell specific growth rate was studied in this paper and the best control strategy was chosen to enhance surfactin production&#46;</p></span><span id="sec0040" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0060">Effect of the control strategy on <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf"><span class="elsevierStyleItalic">out</span></span> and the concentration and production of enriched surfactin in foam</span><p id="par0070" class="elsevierStylePara elsevierViewall">Foam began to overflow outside the bioreactor after 6<span class="elsevierStyleHsp" style=""></span>h of cultivation &#40;<a class="elsevierStyleCrossRef" href="#fig0010">Fig&#46; 2</a>A&#41;&#46; The maximum <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> in control 1 was 0&#46;3<span class="elsevierStyleHsp" style=""></span>l&#47;h while in control 2 it was 1&#46;3<span class="elsevierStyleHsp" style=""></span>l&#47;h&#44; which indicated that the foam overflow was significantly affected by the agitation and aeration rates used in different strategies&#46; The surfactin concentration in the overflow foam also changed with time&#46; As shown in <a class="elsevierStyleCrossRef" href="#fig0010">Figure 2</a>B&#44; different trends were observed in the three control strategies&#46; In control 3&#44; surfactin concentration increased linearly and showed the highest specific production rate&#44; reaching a maximum of 0&#46;426<span class="elsevierStyleHsp" style=""></span>l&#47;h &#40;<a class="elsevierStyleCrossRef" href="#fig0010">Fig&#46; 2</a>C&#41;&#46;</p><elsevierMultimedia ident="fig0010"></elsevierMultimedia><p id="par0075" class="elsevierStylePara elsevierViewall">The linear increase of surfactin concentration and the relatively high specific production rate in control 3 &#40;<a class="elsevierStyleCrossRef" href="#fig0010">Fig&#46; 2</a>B and C&#41; indicated that high surfactin enrichment in foam may be achieved by a combined control of the agitation and aeration rates at a reasonable level&#46; As shown in <a class="elsevierStyleCrossRef" href="#fig0010">Figure 2</a>B&#44; the maximum surfactin concentration in foam reached 3&#46;342<span class="elsevierStyleHsp" style=""></span>g&#47;ml and the maximum surfactin production in foam reached 2&#46;525<span class="elsevierStyleHsp" style=""></span>g &#40;approximately overflowing 7&#46;6<span class="elsevierStyleHsp" style=""></span>l foam&#41; in control 3&#46; However&#44; maximum surfactin concentration and maximum surfactin production were 1&#46;8<span class="elsevierStyleHsp" style=""></span>g&#47;ml and 8&#46;46<span class="elsevierStyleHsp" style=""></span>g &#40;approximately overflowing 4&#46;7<span class="elsevierStyleHsp" style=""></span>l of foam&#41; in control 1&#44; respectively&#46; These results in control 3&#44; which show the linear increase of surfactin concentration and the relatively high specific production rate&#44; are similar to those in the study by Davis <span class="elsevierStyleItalic">et al</span>&#46;<a class="elsevierStyleCrossRef" href="#bib0125"><span class="elsevierStyleSup">6</span></a>&#44; the enrichment of surfactin in the foam was enhanced to 0&#46;44<span class="elsevierStyleHsp" style=""></span>g&#47;l with the time going by supplementing culture broth in the stationary phase<a class="elsevierStyleCrossRef" href="#bib0125"><span class="elsevierStyleSup">6</span></a>&#46; Foam stopped overflowing at 22<span class="elsevierStyleHsp" style=""></span>h in control 1 and at 26<span class="elsevierStyleHsp" style=""></span>h in controls 2 and 3&#46; This was partly due to the medium level in the bioreactor dropping as overflowing foam&#58; however&#59; the surfactin in the foam was thus separated from the broth almost completely<a class="elsevierStyleCrossRef" href="#bib0125"><span class="elsevierStyleSup">6</span></a>&#46; Apparently&#44; continuous surfactin production in the bioreactor ensured an increasing trend of surfactin concentration in the foam&#46; With the agitation and aeration rate controlled under different strategies&#44; the surfactin production presented a different kinetic trend&#46; Control 3&#44; under which <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> was below 0&#46;7<span class="elsevierStyleHsp" style=""></span>l&#47;h&#44; indicated that surfactin enrichment may be improved through stir speed and aeration control&#46;</p></span><span id="sec0045" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0065">Influence of agitation and aeration rates on <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf"><span class="elsevierStyleItalic">out</span></span> and concentration of enriched surfactin</span><p id="par0080" class="elsevierStylePara elsevierViewall">The influence of agitation and aeration rates on <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> and surfactin enrichment in the overflowing foam were investigated in batch culture experiments conducted using a series of combinations of aeration and agitation rates&#46; Foam overflow began after 6<span class="elsevierStyleHsp" style=""></span>h of cultivation &#40;<a class="elsevierStyleCrossRef" href="#fig0010">Fig&#46; 2</a>A&#41;&#46; As indicated in <a class="elsevierStyleCrossRef" href="#fig0015">Fig&#46; 3</a>&#40;A&#8211;C&#41;&#44; <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> tended to increase with an increased aeration rate from 1 to 1&#46;5<span class="elsevierStyleHsp" style=""></span>vvm and agitation rate from 150 to 250<span class="elsevierStyleHsp" style=""></span>rpm&#46; At 33<span class="elsevierStyleHsp" style=""></span>h&#44; <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> was extremely low&#44; only a maximum 0&#46;14<span class="elsevierStyleHsp" style=""></span>l per hour &#40;<a class="elsevierStyleCrossRef" href="#fig0015">Fig&#46; 3</a>C&#41;&#59; however&#44; raising the aeration rate to 250<span class="elsevierStyleHsp" style=""></span>rpm&#44; the <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> could increase to a maximum 1&#46;76 per hour&#46;</p><elsevierMultimedia ident="fig0015"></elsevierMultimedia><p id="par0085" class="elsevierStylePara elsevierViewall">Agitation and aeration rates significantly affected the concentration of surfactin in the foam&#46; In the early stage of fermentation &#40;i&#46;e&#46; at 15<span class="elsevierStyleHsp" style=""></span>h&#59; <a class="elsevierStyleCrossRef" href="#fig0015">Fig&#46; 3</a>A&#41;&#44; an increase in agitation rate increased <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span>&#59; however&#44; the surfactin concentration was at a relatively low level and decreased from 1&#46;6<span class="elsevierStyleHsp" style=""></span>g&#47;l to 1<span class="elsevierStyleHsp" style=""></span>g&#47;l with the agitation increase&#46; At 21<span class="elsevierStyleHsp" style=""></span>h&#44; the maximum surfactin concentration of 4&#46;7<span class="elsevierStyleHsp" style=""></span>g&#47;l occurred at 150<span class="elsevierStyleHsp" style=""></span>rpm agitation and 1<span class="elsevierStyleHsp" style=""></span>vvm aeration&#46; With the increase of agitation to 250<span class="elsevierStyleHsp" style=""></span>rpm&#44; the surfactin concentration was reduced to 1&#46;5<span class="elsevierStyleHsp" style=""></span>g&#47;l&#46; Interestingly&#44; the change in the agitation rate had no effect on the surfactin concentration at 1&#46;5<span class="elsevierStyleHsp" style=""></span>vvm aeration &#40;<a class="elsevierStyleCrossRef" href="#fig0015">Fig&#46; 3</a>E&#41;&#46; At 33<span class="elsevierStyleHsp" style=""></span>h&#44; the surfactin concentration in the foam was maintained at &#62;2&#46;5<span class="elsevierStyleHsp" style=""></span>g&#47;l with an aeration rate of 1&#46;5<span class="elsevierStyleHsp" style=""></span>vvm and it did not drop with an increasing agitation rate&#46; However&#44; under the condition of aeration rate at 1<span class="elsevierStyleHsp" style=""></span>vvm&#44; the surfactin concentration in the foam was 2&#46;5<span class="elsevierStyleHsp" style=""></span>g&#47;l only with the agitation rate of 250<span class="elsevierStyleHsp" style=""></span>rpm&#46; Apparently&#44; the agitation and aeration rates significantly affected the concentration of surfactin enriched in the foam&#46;</p><p id="par0090" class="elsevierStylePara elsevierViewall">The agitation and aeration rates in the aerobic fermentation greatly affected the foam formation rate and the surfactin concentration in the foam&#46; Similar to our case&#44; a previous study observed that a higher stirring speed or aeration rate led to a higher <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span><a class="elsevierStyleCrossRef" href="#bib0125"><span class="elsevierStyleSup">6</span></a>&#46; However&#44; surfactin enrichment in this study presented a different kinetic relationship with <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> compared to that in the basic study by Davis <span class="elsevierStyleItalic">et al</span>&#46;<a class="elsevierStyleCrossRef" href="#bib0125"><span class="elsevierStyleSup">6</span></a>&#44; in which the foam continuously overflowed under a constant agitation rate&#46; In the late stages of batch FOFC&#44; such as at 33<span class="elsevierStyleHsp" style=""></span>h &#40;<a class="elsevierStyleCrossRef" href="#fig0015">Fig&#46; 3</a>F&#41;&#44; the surfactin concentration in the foam was lower than at an earlier stage &#40;21<span class="elsevierStyleHsp" style=""></span>h&#41;&#44; because &#62; 50&#37; of the culture broth in the bioreactor was gone&#46; Thus&#44; the broth level in the bioreactor was too low to maintain the overflowing foam and further foam formation was also limited because almost all the surfactin had already been drained away in the foam&#46; Both the surfactin concentration in the foam and <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> were determined by the surfactin produced in the broth&#46; Furthermore&#44; the <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> level affected the surfactin enrichment&#44; and at the same time&#44; <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> was influenced by the agitation and aeration rates&#46;</p></span><span id="sec0050" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0070">Fed-batch culture with controlled agitation and aeration rates</span><p id="par0095" class="elsevierStylePara elsevierViewall">A fed-batch culture process was also adopted in this study&#46; <a class="elsevierStyleCrossRef" href="#fig0020">Figure 4</a> shows that foam began to overflow after 6<span class="elsevierStyleHsp" style=""></span>h of cultivation&#46; Afterwards&#44; <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> gradually dropped from 0&#46;6<span class="elsevierStyleHsp" style=""></span>l&#47;h to 0&#46;2<span class="elsevierStyleHsp" style=""></span>l&#47;h by real-time control of the agitation and aeration rates and was maintained at this rate&#46; From 15<span class="elsevierStyleHsp" style=""></span>h&#44; the IBM medium was fed into the bioreactor at a 0&#46;2<span class="elsevierStyleHsp" style=""></span>l&#47;h feed rate &#40;matching <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span>&#41;&#46; In this culture&#44; the biomass was kept above 10<span class="elsevierStyleSup">9</span><span class="elsevierStyleHsp" style=""></span>CFU&#47;ml&#46; Foam continued to overflow until the cultivation terminated and surfactin was enriched in the foam with a concentration &#62;4<span class="elsevierStyleHsp" style=""></span>g&#47;l&#59; the maximum concentration reached 4&#46;7<span class="elsevierStyleHsp" style=""></span>g&#47;l&#46; At the end of the fermentation&#44; the surfactin concentration in the culture broth in the bioreactor was very low &#40;data not shown&#41;&#44; which means that almost all the surfactin was transferred into the foam&#46;</p><elsevierMultimedia ident="fig0020"></elsevierMultimedia><p id="par0100" class="elsevierStylePara elsevierViewall">In other studies of foam overflowing cultures&#44; although the product was enriched and separated in foam&#44; the foam overflowed spontaneously&#44; and thus the surfactin concentration in it fluctuated<a class="elsevierStyleCrossRefs" href="#bib0125"><span class="elsevierStyleSup">6&#44;10&#44;16</span></a>&#46; For this reason&#44; surfactin in the foam was not enriched to a very high level&#59; in previous studies&#44; in which surfactin was produced by potato process effluent&#44; it was 1&#46;67<span class="elsevierStyleHsp" style=""></span>g&#47;l and 0&#46;9<span class="elsevierStyleHsp" style=""></span>g&#47;l<a class="elsevierStyleCrossRefs" href="#bib0125"><span class="elsevierStyleSup">6&#44;10</span></a>&#46; Thus&#44; controlled foam overflow in this work achieved almost 3 and 4 times the surfactin concentration compared with the previous studies&#44; respectively<a class="elsevierStyleCrossRefs" href="#bib0125"><span class="elsevierStyleSup">6&#44;10</span></a>&#46; Furthermore&#44; the medium feeding in our study overcame disruption of cell growth&#44; which ensured that surfactin was continuously produced&#46; Combined with control of <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> at a low level&#44; the surfactin concentration in the foam was maintained above 4<span class="elsevierStyleHsp" style=""></span>g&#47;l&#44; which will significantly reduce production costs and improve industrial production capacity&#46;</p></span></span><span id="sec0055" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0075">Conclusions</span><p id="par0105" class="elsevierStylePara elsevierViewall">Our study focused on the control of agitation and aeration rates in foam overflowing fermentation to improve surfactin enrichment and continuous high-level production&#46; In batch mode&#44; the agitation and aeration rates were found to have a close relationship with cell growth and surfactin production&#46; In further research&#44; our study revealed that <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> does not always negatively impact on the surfactin concentration in the foam&#46; The broth level in the bioreactor and the surfactin residue in the broth also affect both <span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span> and the surfactin enrichment of the foam&#46;</p><p id="par0110" class="elsevierStylePara elsevierViewall">Through feeding the medium into the bioreactor&#44; a fed-batch fermentation process was successfully established in which foam overflowed at a controlled flow rate of 0&#46;2<span class="elsevierStyleHsp" style=""></span>l&#47;h&#44; and surfactin in the enriched foam was kept at a level above 4<span class="elsevierStyleHsp" style=""></span>g&#47;l&#46;</p></span><span id="sec0060" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0080">Ethical disclosures</span><span id="sec0065" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0085">Protection of human and animal subjects</span><p id="par0140" class="elsevierStylePara elsevierViewall">The authors declare that no experiments were performed on humans or animals for this study&#46;</p></span><span id="sec0070" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0090">Confidentiality of data</span><p id="par0145" class="elsevierStylePara elsevierViewall">The authors declare that no patient data appear in this article&#46;</p></span><span id="sec0075" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0095">Right to privacy and informed consent</span><p id="par0150" class="elsevierStylePara elsevierViewall">The authors declare that no patient data appear in this article&#46;</p></span></span><span id="sec0085" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0100">Conflict of interest</span><p id="par0135" class="elsevierStylePara elsevierViewall">The authors declare that they have no conflicts of interest&#46;</p></span></span>"
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              "titulo" => "Effect of the control strategy on f and the concentration and production of enriched surfactin in foam"
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              "titulo" => "Influence of agitation and aeration rates on f and concentration of enriched surfactin"
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              "titulo" => "Fed-batch culture with controlled agitation and aeration rates"
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            1 => "Agitation rate"
            2 => "Aeration rate"
            3 => "Foam overflowing"
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            2 => "Tasa de aireaci&#243;n"
            3 => "Espuma desbordante"
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        "titulo" => "Abstract"
        "resumen" => "<span id="abst0005" class="elsevierStyleSection elsevierViewall"><p id="spar0005" class="elsevierStyleSimplePara elsevierViewall"><span class="elsevierStyleItalic">Bacillus amyloliquefaciens</span> fmb50 produces a high yield of surfactin&#44; a lipopeptide-type biosurfactant that has been widely studied and has potential applications in many fields&#46; A foam overflowing culture has been successfully used in the combined production-enrichment fermentation of surfactin&#46; In this study&#44; the agitation and aeration rates were found to have relationships with foam formation and surfactin enrichment&#46; A maximum surfactin concentration of 4&#46;7<span class="elsevierStyleHsp" style=""></span>g&#47;l of foam was obtained after 21<span class="elsevierStyleHsp" style=""></span>h of culture with an agitation rate of 150<span class="elsevierStyleHsp" style=""></span>rpm and an aeration rate of 1<span class="elsevierStyleHsp" style=""></span>vvm in fed-batch culture&#46; By controlling the foam overflow rate &#40;<span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span>&#41; of a fed-batch culture&#44; surfactin concentration in the foam was continuously maintained above 4<span class="elsevierStyleHsp" style=""></span>g&#47;l&#46;</p></span>"
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        "resumen" => "<span id="abst0010" class="elsevierStyleSection elsevierViewall"><p id="spar0010" class="elsevierStyleSimplePara elsevierViewall"><span class="elsevierStyleItalic">Bacillus amyloliquefaciens</span> fmb50 produce gran cantidad de surfactina&#44; un biosurfactante de tipo lipopept&#237;dico que ha sido objeto de estudios pormenorizados y tiene aplicaciones en muchos campos&#46; El cultivo en espuma desbordante se ha utilizado con &#233;xito en la fermentaci&#243;n combinada de producci&#243;n-enriquecimiento de surfactina&#46; En este estudio&#44; se hall&#243; que las tasas de aireaci&#243;n y agitaci&#243;n tienen relaci&#243;n con la formaci&#243;n de espuma y el enriquecimiento de la surfactina&#46; Se obtuvo una concentraci&#243;n m&#225;xima de surfactina de 4&#44;7<span class="elsevierStyleHsp" style=""></span>g&#47;l de espuma despu&#233;s de 21 h de cultivo con una tasa de agitaci&#243;n de 150 rpm y una tasa de aireaci&#243;n de 1 vvm en un cultivo alimentado <span class="elsevierStyleItalic">&#40;fed-batch&#41;</span>&#46; Al controlar la tasa de espuma desbordante &#40;<span class="elsevierStyleItalic">f</span><span class="elsevierStyleInf">out</span>&#41; de un cultivo <span class="elsevierStyleItalic">fed-batch</span>&#44; la concentraci&#243;n de surfactina en la espuma se mantuvo continua por encima de 4 g&#47;l&#46;</p></span>"
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                        0 => array:2 [
                          "etal" => false
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                            0 => "K&#46; Arima"
                            1 => "A&#46; Kakinuma"
                            2 => "G&#46; Tamura"
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                        ]
                      ]
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                      "Revista" => array:6 [
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              ]
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                      "titulo" => "Biosurfactants production and possible uses in microbial enhanced oil recovery and oil pollution remediation"
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                        0 => array:2 [
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                            0 => "I&#46;M&#46; Banat"
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                            1 => "C&#46;R&#46; MacDonald"
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                  "contribucion" => array:1 [
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                      "titulo" => "Control of the specific growth rate of <span class="elsevierStyleItalic">Bacillus subtilis</span> for the production of biosurfactant lipopeptides in bioreactors with foam overflow"
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                            0 => "S&#46; Chenikher"
                            1 => "J&#46;S&#46; Guez"
                            2 => "F&#46; Coutte"
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              "referencia" => array:1 [
                0 => array:2 [
                  "contribucion" => array:1 [
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                      "titulo" => "Production of surfactin and fengycin by <span class="elsevierStyleItalic">Bacillus subtilis</span> in a bubbleless membrane bioreactor"
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Article information

ISSN: 03257541
Original language: English

DOI:10.1016/j.ram.2015.09.003

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Year/Month	Html	Pdf	Total

2024 October	18	4	22
2024 September	32	11	43
2024 August	21	4	25
2024 July	25	5	30
2024 June	26	8	34
2024 May	27	10	37
2024 April	32	12	44
2024 March	55	14	69
2024 February	53	3	56
2024 January	54	6	60
2023 December	40	8	48
2023 November	46	10	56
2023 October	59	6	65
2023 September	33	5	38
2023 August	19	2	21
2023 July	38	10	48
2023 June	30	4	34
2023 May	31	8	39
2023 April	28	5	33
2023 March	28	6	34
2023 February	55	13	68
2023 January	31	5	36
2022 December	51	11	62
2022 November	59	12	71
2022 October	55	10	65
2022 September	35	12	47
2022 August	48	17	65
2022 July	46	14	60
2022 June	39	7	46
2022 May	50	9	59
2022 April	69	16	85
2022 March	112	19	131
2022 February	137	13	150
2022 January	92	9	101
2021 December	78	18	96
2021 November	79	16	95
2021 October	90	17	107
2021 September	65	14	79
2021 August	86	29	115
2021 July	56	12	68
2021 June	45	13	58
2021 May	68	19	87
2021 April	137	66	203
2021 March	61	22	83
2021 February	62	12	74
2021 January	49	9	58
2020 December	51	6	57
2020 November	57	22	79
2020 October	29	3	32
2020 September	30	9	39
2020 August	38	4	42
2020 July	41	6	47
2020 June	41	13	54
2020 May	51	15	66
2020 April	53	13	66
2020 March	54	8	62
2020 February	57	14	71
2020 January	36	3	39
2019 December	31	6	37
2019 November	50	11	61
2019 October	40	13	53
2019 September	53	7	60
2019 August	28	11	39
2019 July	35	6	41
2019 June	81	8	89
2019 May	190	1	191
2019 April	89	14	103
2019 March	26	4	30
2019 February	25	6	31
2019 January	19	4	23
2018 December	20	6	26
2018 November	35	6	41
2018 October	45	12	57
2018 September	40	5	45
2018 August	15	15	30
2018 July	17	2	19
2018 June	15	2	17
2018 May	22	9	31
2018 April	27	13	40
2018 March	19	2	21
2018 February	8	4	12
2018 January	11	4	15
2017 December	16	2	18
2017 November	14	5	19
2017 October	9	10	19
2017 September	13	4	17
2017 August	9	6	15
2017 July	8	5	13
2017 June	10	8	18
2017 May	20	9	29
2017 April	14	15	29
2017 March	11	52	63
2017 February	30	1	31
2017 January	10	7	17
2016 December	23	9	32
2016 November	35	9	44
2016 October	42	21	63
2016 September	53	11	64
2016 August	45	11	56
2016 July	20	2	22
2016 June	41	22	63
2016 May	29	19	48
2016 April	30	13	43
2016 March	58	28	86
2016 February	31	26	57
2016 January	43	28	71
2015 December	9	6	15

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