Production of citric acid using its extraction wastewater treated by anaerobic digestion and ion exchange in an integrated citric acid-methane fermentation process

In order to solve the problem of extraction wastewater pollution in citric acid industry, an integrated citric acid-methane fermentation process is proposed in this study. Extraction wastewater was treated by mesophilic anaerobic digestion and then used to make mash for the next batch of citric acid fermentation. The recycling process was done for seven batches. Citric acid production (82.4 g/L on average) decreased by 34.1?% in the recycling batches (2nd–7th) compared with the first batch. And the residual reducing sugar exceeded 40 g/L on average in the recycling batches. Pigment substances, acetic acid, ammonium, and metal ions in anaerobic digestion effluent (ADE) were considered to be the inhibitors, and their effects on the fermentation were studied. Results indicated that ammonium, Na⁺ and K⁺ in the ADE significantly inhibited citric acid fermentation. Therefore, the ADE was treated by acidic cation exchange resin prior to reuse to make mash for citric acid fermentation. The recycling process was performed for ten batches, and citric acid productions in the recycling batches were 126.6 g/L on average, increasing by 1.7 % compared with the first batch. This process could eliminate extraction wastewater discharge and reduce water resource consumption.     

Effect of propionic acid on citric acid fermentation in an integrated citric acid–methane fermentation process

In this study, an integrated citric acid-methane fermentation process was established to solve the problem of wastewater treatment in citric acid production. Citric acid wastewater was treated through anaerobic digestion and then the anaerobic digestion effluent (ADE) was further treated and recycled for the next batch citric acid fermentation. This process could eliminate wastewater discharge and reduce water resource consumption. Propionic acid was found in the ADE and its concentration continually increased in recycling. Effect of propionic acid on citric acid fermentation was investigated, and results indicated that influence of propionic acid on citric acid fermentation was contributed to the undissociated form. Citric acid fermentation was inhibited when the concentration of propionic acid was above 2, 4, and 6 mM in initial pH 4.0, 4.5 and, 5.0, respectively. However, low concentration of propionic acid could promote isomaltase activity which converted more isomaltose to available sugar, thereby increasing citric acid production. High concentration of propionic acid could influence the vitality of cell and prolong the lag phase, causing large amount of glucose still remaining in medium at the end of fermentation and decreasing citric acid production.     

Identification and characterization of a novel endoglucanase (CMCase) isolated from the larval gut of Bombyx mori

While screening for cellulase-producing fungi from insect gut, a fungus with high endoglucanase (carboxymethyl cellulase; CMCase) activity was isolated from the larval gut of Bombyx mori. Based on morphological characteristics and using an 18S rRNA-based molecular phylogenetic approach, the fungus, strain BMC-2, was identified as a Mucor sp. expressing a novel alkalotolerant cellulase. The maximum production of cellulase by the BMC-2 strain was observed at 55 °C and pH 8.0. The CMCase activity was inhibited by Cu^2 + > Na⁺ > Zn^2 + > Mg^2 + > Ba^2 +, and induced by Ca^2 +, Mn^2 +, Fe^2 +, and K⁺.     

Synthesis and ionophoric activities of functionalized bis(choloyl) conjugates with a rigid core

Three bis(choloyl) conjugates bearing a rigid p-phenylenediamine/p-bis(aminomethyl)benzene linker and amino/acetamido groups were synthesized, and fully characterized on the basis of ¹H NMR, ESI-MS and HRMS. Their ionophoric activities were investigated by means of pH discharge assay. The results indicate that these conjugates exhibit potent ionophoric activities across egg-yolk l-α-phosphatidylcholine (EYPC)-based liposomal membranes, via a cation/proton antiport mechanism. They show moderate ion selectivity among alkali metal ions. Of the three conjugates, the ones having amino groups transport alkali metal ions in the order of Na⁺ > Li⁺ > K⁺ ≈ Rb⁺ ≈ Cs⁺, whereas the one having acetamido groups functions in the order of Li⁺ > Na⁺ > K⁺ ≈ Rb⁺ ≈ Cs⁺.     

Applying geographic information systems and remote sensing for water quality assessment of mangrove forest

The study of environmental conditions is one of the most important measures in the field of reforestation. The present study was undertaken to assess the environmental status of the mangrove forest of Alibaug, Maharashtra, India with respect to different sixteen physicochemical parameters of water using Geographical information system (GIS) for rehabilitation, conservation and development of the destructed area of the mangrove forest. The Base map of study area was prepared using topographic map and the remote sensing data of Landsat 7 ETM + for spatial analysis. The distributions of water pollutants were assigned using a GIS approach of Inverse Distance Weighted (IDW). The results showed that the amounts of EC, COD, hardness, O&G, Cl^?, Na⁺, Ca^2 +, Mg^2 +, NO₃^? and PO₄^3? are higher than the normal ranges in mangrove forest due to natural processes and human activity, industrial and domestic wastewater disposal, oil spillage and agricultural runoff which all eventually affect the water quality of mangrove forest of Alibaug. To identify the areas within the normal ranges of 16 studied parameter, suitability map of water was prepared through an integration of 16 suitability maps of the studied parameters. The suitability map of water classified the water to six classes of suitability in order of moderate > moderate to high > low to moderate > high > low suitable. The areas with classes of 1 and 2 were suitable for the protective measures. Classes 3 and 4 were suitable for replantation and restoration of native mangrove species as well as local communities' cooperation in the participatory protection measures. The areas of classes 5 and 0 need to be designed an urgent management and mitigation plan to reduce impact of human activities. The result of the study also proves the use of GIS as a powerful tool in addressing assessment and monitoring programs of the water quality in the mangrove ecosystems.     

Long chain fatty acids degradation in anaerobic digester: Thermodynamic equilibrium consideration

The biological oxidations and reductions occurring in batch anaerobic digesters may approach equilibrium. The approach depends strongly on the activities of the micro-organisms present. The thermodynamics of linear long chain fatty acid degradation in a batch reactor was modeled. The substrates considered are the saturated fatty acids from acetic acid to stearic acid. From the thermodynamic perspective, the fermentation (acidogenesis and acetogenesis), decomposing the long chain saturated fatty acids to acetic acid though shorter chain acids, could not proceed spontaneously (ΔH ≫ 0 and ΔG ≫ 0). However the model suggests that the major driving force for the fermentation may be found in the methanogenesis. The model results show two distinct cases: (ΔS > 0 and ΔH > 0) and (ΔS < 0 and ΔH < 0), relating to spontaneous but endothermic and non-spontaneous but exothermic processes respectively. Where, spontaneous digestion is associated with high initial concentrations of LCFA and endotherm. This implies that the digestion of this type of substrate might be better facilitated by the supply of supplemental heat. The digestion of very low concentrations of LCFA is found to be non-spontaneous due in large part to the solubility of carbon dioxide. This implies that the digestion of this type of substrate might be enabled by selectively removing carbon dioxide.     

Purification and biochemical properties of an extracellular acid phytase produced by the Saccharomyces cerevisiae CY strain

An extracellular acid phytase was purified to homogeneity from the culture supernatant of the Saccharomyces cerevisiae CY strain by ultrafiltration, DEAE-Sepharose column chromatography, and Sephacryl S-300 gel filtration. The molecular weight of the purified enzyme was estimated to be 630 kDa by gel filtration. Removing the sugar chain by endoglycosidase H digestion revealed that the molecular mass of the protein decreased to 446 kDa by gel filtration and gave a band of 55 kDa by SDS-PAGE. The purified enzyme was most active at pH 3.6 and 40 °C and was fairly stable from pH 2.5 to 5.0. The phytase displayed broad substrate specificity and had a K_m value of 0.66 mM (sodium phytate, pH 3.6, 40 °C). The phytase activity was completely inhibited by Fe³⁺ and Hg²⁺, and strongly inhibited (maximum of 91%) by Ba²⁺, Co²⁺, Cu⁺, Cu²⁺, Fe²⁺, Mg²⁺, and Sn²⁺ at 5 mM concentrations.     

Salt effects on β-glucosidase: pH-profile narrowing

Salts inhibit the activity of sweet almond β-glucosidase. For cations (Cl⁻ salts) the effectiveness follows the series: Cu⁺², Fe⁺² > Zn⁺² > Li⁺ > Ca⁺² > Mg⁺² > Cs⁺ > NH₄⁺ > Rb⁺ > K⁺ > Na⁺ and for anions (Na⁺ salts) the series is: I⁻ > ClO₄⁻ > ⁻SCN > Br⁻ ≈ NO₃⁻ > Cl⁻ ≈ ⁻OAc > F⁻ ≈ SO₄^− 2. The activity of the enzyme, like that of most glycohydrolases, depends on a deprotonated carboxylate (nucleophile) and a protonated carboxylic acid for optimal activity. The resulting pH-profile of k_cat/K_m for the β-glucosidase-catalyzed hydrolysis of p-nitrophenyl glucoside is characterized by a width at half height that is strongly sensitive to the nature and concentration of the salt. Most of the inhibition is due to a shift in the enzymic pK_as and not to an effect on the pH-independent second-order rate constant, (k_cat/K_m)^lim. For example, as the NaCl concentration is increased from 0.01 M to 1.0 M the apparent pK_a1 increases (from 3.7 to 4.9) and the apparent pK_a2 decreases (from 7.2 to 5.9). With p-nitrophenyl glucoside, the value of the pH-independent (k_cat/K_m)^lim (= 9 × 10⁴ M^− 1 s^− 1) is reduced by less than 4% as the NaCl concentration is increased. There is a similar shift in the pK_as when the LiCl concentration is increased to 1.0 M. The results of these salt-induced pK_a shifts rule out a significant contribution of reverse protonation to the catalytic efficiency of the enzyme. At low salt concentration, the fraction of the catalytically active monoprotonated enzyme in the reverse protonated form (i.e., proton on the group with a pK_a of 3.7 and dissociated from the group with a pK_a of 7.2) is very small (≈ 0.03%). At higher salt concentrations, where the two pK_as become closer, the fraction of the monoprotonated enzyme in the reverse protonated form increases over 300-fold. However, there is no increase in the intrinsic reactivity, (k_cat/K_m)^lim, of the monoprotonated species. For other enzymes which may show such salt-induced pK_a shifts, this provides a convenient test for the role of reverse protonation.     

Fluoroquinolones stimulate the DNA cleavage activity of topoisomerase IV by promoting the binding of Mg2 + to the second metal binding site

BackgroundFluoroquinolones target bacterial type IIA topoisomerases, DNA gyrase and topoisomerase IV (Topo IV). Fluoroquinolones trap a topoisomerase–DNA covalent complex as a topoisomerase–fluoroquinolone–DNA ternary complex and ternary complex formation is critical for their cytotoxicity. A divalent metal ion is required for type IIA topoisomerase-catalyzed strand breakage and religation reactions. Recent studies have suggested that type IIA topoisomerases use two metal ions, one structural and one catalytic, to carry out the strand breakage reaction.MethodsWe conducted a series of DNA cleavage assays to examine the effects of fluoroquinolones and quinazolinediones on Mg^2 +-, Mn^2 +-, or Ca^2 +-supported DNA cleavage activity of Escherichia coli Topo IV.ResultsIn the absence of any drug, 20–30 mM Mg^2 + was required for the maximum levels of the DNA cleavage activity of Topo IV, whereas approximately 1 mM of either Mn^2 + or Ca^2 + was sufficient to support the maximum levels of the DNA cleavage activity of Topo IV. Fluoroquinolones promoted the Topo IV-catalyzed strand breakage reaction at low Mg^2 + concentrations where Topo IV alone could not efficiently cleave DNA.Conclusions and general significanceAt low Mg^2 + concentrations, fluoroquinolones may stimulate the Topo IV-catalyzed strand breakage reaction by promoting Mg^2 + binding to metal binding site B through the structural distortion in DNA. As Mg^2 + concentration increases, fluoroquinolones may inhibit the religation reaction by either stabilizing Mg^2 + at site B or inhibition the binding of Mg^2 + to site A. This study provides a molecular basis of how fluoroquinolones stimulate the Topo IV-catalyzed strand breakage reaction by modulating Mg^2 + binding.     

Supplementation of ammonium bicarbonates for the treatment of fruit cordial wastewater by anaerobic digestion process

Lack of nitrogenous substrate and buffering capacity have been identified as causing failure in previous work on the treatment of fruit cordial wastewater using anaerobic continuous stirred tank reactors. In this study, ammonium bicarbonate was proposed to be used as the substrate for nitrogenous and buffering resources. In order to determine the toxicity effect of the ammonium salts on the anaerobic system, a series of concentration from 0 to 40 mg L^?1 was tested. Biogas production was used as the indicator for NH₄⁺ toxicity. The results showed no indication that methanogen was affected by the additional ammonium salt within the dosing regime. Application of the specific mathematical function (G = G_m^k/t) to describe the kinetic of biogas production, suggested that the optimal concentration of ammonium bicarbonate that can be used is 10 mg L^?1. This study also shows that the dosage regime up to 40 mg L^?1 can be used to supplement the lack of nitrogenous and buffering capacity for the anaerobic digestion process of the fruit cordial wastewater using CSTR.     

A novel three-TMH Na+/H+ antiporter and the functional role of its oligomerization

Na⁺/H⁺ antiporters are a category of ubiquitous transmembrane proteins with various important physiological roles in almost all living organisms ranging from bacteria to humans. However, the knowledge of novel Na⁺/H⁺ antiporters remains to be broadened, and the functional roles of oligomerization in these antiporters have not yet been thoroughly understood. Here, we reported functional analysis of an unknown transmembrane protein composed of 103 amino acid residues. This protein was found to function as a Na⁺(Li⁺, K⁺)/H⁺ antiporter. To the best of our knowledge, this antiporter is the minimal one of known Na⁺/H⁺ antiporters and thus designated as NhaM to represent the minimal Na⁺/H⁺ antiporter. NhaM and its homologs have not yet been classified into any protein family. Based on phylogenetic analysis and protein alignment, we propose NhaM and its homologs to constitute a novel transporter family designated as NhaM family. More importantly, we found that NhaM is assembled with parallel protomers into a homo-oligomer and oligomerization is vital for the function of this antiporter. This implies that NhaM may adopt and require an oligomer structure for its normal function to create a similar X-shaped structure to that of the NhaA fold. Taken together, current findings not only present the proposal of a novel transporter family but also positively contribute to the functional roles of oligomerization in Na⁺/H⁺ antiporters.     

Magnesium uptake of Arabidopsis transporters,AtMRS2-10 and AtMRS2-11, expressed in Escherichia coli mutants: Complementation and growth inhibition by aluminum

Magnesium (Mg^2 +) plays a critical role in many physiological processes. Mg^2 + transport systems in Salmonella have been well documented, but those in Escherichia coli have not been fully elucidated. We examined the effects of corA, mgtA, yhiD and corC gene deletion on Mg^2 + transport in E. coli. We obtained every combination of double, triple and quadruple mutants. The corA and mgtA double mutant required addition of 10 mM Mg^2 + to Luria-Bertani (LB) medium for growth, and the corA, mgtA and yhiD triple mutant TM2 required a higher Mg^2 + concentration. The Mg^2 + requirement of the quadruple mutant was similar to that of TM2. The results demonstrated that either CorA or MgtA is necessary for normal E. coli growth in LB medium and that YhiD plays a role in Mg^2 + transport under high Mg^2 + growth conditions in E. coli. The Arabidopsis Mg^2 + transporters, AtMRS2-10 and AtMRS2-11, were heterologously expressed in TM2 cells. TM2 cells expressing AtMRS2-10 and AtMRS2-11 could grow in LB medium that had been supplemented with 1 mM Mg^2 + and without Mg^2 + supplementation, respectively, and cell growth was inhibited by 2 mM AlCl₃. The results indicated that the growth of TM2 expressing AtMRS2-10 and AtMRS2-11 reflected these AtMRS2 function for Mg^2 + and aluminum. The E. coli TM2 cells are useful for functional analysis of Arabidopsis MRS2 proteins.     

Identification of microorganisms in the granules generated during methane fermentation of the syrup wastewater produced while canning fruit

The wastewater produced in the process of canning fruit contains a syrup that consists mainly of sucrose. This syrup wastewater was treated by methane fermentation in an upflow anaerobic sludge blanket reactor. The organic loading rate of syrup wastewater was increased gradually as fermentation progressed. The higher the organic loading rate, the more methane gas evolved until the organic loading rate reached 30.3 kg COD m^?3 d^?1, at which point methane generation abruptly diminished because the loading rate was too high to stably operate the reactor. The changes in the microbial community, that of both bacteria and archaea in the granules, were analyzed simultaneously using PCR-DGGE during the fermentation process. Methanosaeta spp., which are methanogenic archaea that produce extracellular polymers indispensable for the formation of granules, were dominant when the methane gas vigorously evolved, and the iron-reducing bacterium belonging to genus Geobacter, which outcompetes methanogens, grew proportionally with the deterioration of methane fermentation.     

Modelling and simulation of continuous L (+) lactic acid production from sugarcane juice in membrane integrated hybrid-reactor system

Modelling and simulation was done for a two-stage membrane-integrated hybrid reactor system for continuous production of L (+) lactic acid under non-neutralizing conditions. The model captures microbial conversion of sugar cane juice to lactic acid under substrate–product inhibitions with downstream purification by nanofiltration. All the major phenomena and the governing parameters like fluid flow, feed dilution, substrate–product inhibitions, Donnan and steric effects during micro and nanofiltration for cell recycle, product separation and purification have been reflected in the modelling. The model describes a green, integrated continuous process of direct lactic acid production starting with a cheap, renewable carbon source. The highest lactic acid concentration achieved after the final stage of nanofiltration was 66.97 g/L at 13 kg/cm² operating pressure when the overall productivity reached 12.40 g/(L h). The developed model could successfully predict production, purification and transport of lactic acid through two stage membrane modules. Performance of the model was very good as indicated in the high overall correlation coefficient (R² > 0.980) and the low relative error (RE < 0.1).     

Improving biogas production from protein-rich distillery wastewater by decreasing ammonia inhibition

A large quantity of protein-rich distillery wastewater is produced during the process of bio-ethanol production from kitchen waste. It is difficult, however, to treat protein-rich distillery wastewater by anaerobic digestion due to ammonia inhibition. In this study, a novel method was investigated to reduce ammonia inhibition during thermophilic anaerobic digestion through the recirculation of water-washed biogas into the headspace (R1 system) or liquid phase (R2 system) of the reactors. The results show that the method greatly improved biogas production from distillery wastewater. R2 system achieved stable biogas production at a higher organic loading rate (OLR) of 4.0 g VTS/L/d than R1 system at 3.0 g VTS/L/d. At the same OLR, we observed a higher biogas production rate but lower accumulation of NH₄⁺ and volatile fatty acids in the reactor, and higher ammonia absorption rate in the water tank of R2 system than R1 system. The better performance of R2 system could be attributed to the more efficient removal of ammonia from liquid phase. In addition, adjusting the C/N ratio of distillery wastewater from 9.0 to 11.4 significantly enhanced the maximum OLR from 3.0 to 7.0 g VTS/L/d in R1 system.     

Thapsigargin decreases the Na+- Ca2 + exchanger mediated Ca2 + entry in pig coronary artery smooth muscle

Na⁺- Ca^2 + exchanger (NCX) has been proposed to play a role in refilling the sarco/endoplasmic reticulum (SER) Ca^2 + pool along with the SER Ca^2 + pump (SERCA). Here, SERCA inhibitor thapsigargin was used to determine the effects of SER Ca^2 + depletion on NCX–SERCA interactions in smooth muscle cells cultured from pig coronary artery. The cells were Na⁺-loaded and then placed in either a Na⁺-containing or in a Na⁺-substituted solution. Subsequently, the difference in Ca^2 + entry between the two groups was examined and defined as the NCX mediated Ca^2 + entry. The NCX mediated Ca^2 + entry in the smooth muscle cells was monitored using two methods: Ca^2 +sensitive fluorescence dye Fluo-4 and radioactive Ca^2 +. Ca^2 +-entry was greater in the Na⁺-substituted cells than in the Na⁺-containing cells when measured by either method. This difference was established to be NCX-mediated as it was sensitive to the NCX inhibitors. Thapsigargin diminished the NCX mediated Ca^2 + entry as determined by either method. Immunofluorescence confocal microscopy was used to determine the co-localization of NCX1 and subsarcolemmal SERCA2 in the cells incubated in the Na⁺-substituted solution with or without thapsigargin. SER Ca^2 + depletion with thapsigargin increased the co-localization between NCX1 and the subsarcolemmal SERCA2. Thus, inhibition of SERCA2 leads to blockade of constant Ca^2 + entry through NCX1 and also increases proximity between NCX1 and SERCA2. This blockade of Ca^2 + entry may protect the cells against Ca^2 +-overload during ischemia–reperfusion when SERCA2 is known to be damaged.     

Significance of metal ion supplementation in the fermentation medium on the structure and anti-tumor activity of Tuber polysaccharides produced by submerged culture of Tuber melanosporum

The significance of metal ion supplementation in the fermentation medium on the structure and anti-tumor activity of Tuber polysaccharides was systematically studied in the submerged fermentation of Tuber melanosporum. The lowest weight-average molecular weight (M_w) (i.e., 115.3 × 10⁴ g/mol) of intracellular polysaccharides (IPS) was obtained when Mg²⁺ and K⁺ was added in the fermentation medium. The IPS with the lower M_w exhibited a higher inhibition ratio against S-180 tumor cells. The compact conformation of extracellular polysaccharides (EPS) was formed when only K⁺ was supplied in the fermentation medium. Interestingly, EPS with compact conformation exhibited a higher inhibition ratio (i.e., 59.2%) than EPS with branched polymer chain (i.e., 9.2%) against A549 tumor cells. The highest inhibition ratio for EPS with α-glycosidic linkages against the tumor cell line HepG2 reached 32.2% when Mg²⁺ or K⁺ was supplied in the fermentation medium. The addition of metal ion Mg²⁺, K⁺, and their combination to the fermentation medium is a vital factor affecting the structures of Tuber polysaccharides, which further determine their anti-tumor activities. The information obtained in this work will be useful for the efficient and directed production of polysaccharides with anti-tumor activities by the submerged fermentation of edible fungi mycelium.     

Effects of redox potential control on succinic acid production by engineered Escherichia coli under anaerobic conditions

The effects of oxido-reduction potential (ORP) control on succinic acid production have been investigated in Escherichia coli LL016. In LL016, two CO₂ fixation pathways were achieved and NAD⁺ supply was enhanced by co-expression of heterologous pyruvate carboxylase (PYC) and nicotinic acid phosphoribosyltransferase (NAPRTase). During anaerobic fermentation, cell growth and metabolite distribution were changed with redox potential levels in the range of −200 to −400 mV. From the results, the ORP level of −400 mV was preferable, which resulted in the high succinic acid concentration (28.6 g/L) and high succinic acid productivity (0.33 g/L/h). Meanwhile, the yield of succinic acid at the ORP level of −400 mV was 39% higher than that at the ORP level of −200 mV. In addition, a higher NADH/NAD⁺ ratio and increased enzyme activities were also achieved by regulating the culture to a more reductive environment, which further enhanced the succinic acid production.     

Removal of nutrients from wastewater with Canna indica L. under different vertical-flow constructed wetland conditions

Constructed wetlands are becoming increasingly popular worldwide for removing contaminants from domestic wastewater. This study investigated the removal efficiency of nitrogen (N) and phosphorus (P) from wastewater with the simulated vertical-flow constructed wetlands (VFCWs) under three different substrates (i.e., BFAS or blast furnace artificial slag, CBAS or coal burn artificial slag, and MSAS or midsized sand artificial slag), hydraulic loading rates (i.e., 7, 14, and 21 cm d^?1), and wetland operational periods (0.5, 1, and 2 years) as well as with and without planting Canna indica L. The wastewater was collected from the campus of South China Agricultural University, Guangzhou, China. Results show that the percent removal of total P (TP) and ammonium N (NH₄⁺-N) by the substrates was BFAS > CBAS > MSAS due to the high contents of Ca and Al in substrate BFAS. In contrast, the percent removal of total N (TN) by the substrates was CBAS > MSAS > BFAS due to the complicated nitrification/denitrification processes. The percent removal of nutrients by all of the substrates was TP > NH₄⁺-N > TN. About 10% more TN was removed from the wastewater after planting Canna indica L. A lower hydraulic loading rate or longer hydraulic retention time (HRT) resulted in a higher removal of TP, NH₄⁺-N, and TN because of more contacts and interactions among nutrients, substrates, and roots under the longer HRT. Removal of NO₃^?N from the simulated VFCWs is a complex process. A high concentration of NO₃^?N in the effluent was observed under the high hydraulic loading rate because more NH₄⁺-N and oxygen were available for nitrification and a shorter HRT was unfavorable for denitrification. In general, a longer operational period had a highest removal rate for nutrients in the VFCWs.     

Secretion of Na+, K+ and fluid by the Malpighian (renal) tubule of the larval cabbage looper Trichoplusia ni (Lepidoptera: Noctuidae)

The Malpighian (renal) tubules play important roles in ionic and osmotic homeostasis in insects. In Lepidoptera, the Malpighian tubules are structurally regionalized and the concentration of Na⁺ and K⁺ in the secreted fluid varies depending on the segment of tubule analyzed. In this work, we have characterized fluid and ion (Na⁺, K⁺, H⁺) transport by tubules of the larval stage of the cabbage looper Trichoplusia ni; we have also evaluated the effects of fluid secretion inhibitors and stimulants on fluid and ion transport. Ramsay assays showed that fluid was secreted by the iliac plexus but not by the yellow and white regions of the tubule. K⁺ and Na⁺ were secreted by the distal iliac plexus (DIP) and K⁺ was reabsorbed in downstream regions. The fluid secretion rate decreased > 50% after 25 μM bafilomycin A1, 500 μM amiloride or 50 μM bumetanide was added to the bath. The concentration of K⁺ in the secreted fluid did not change, whereas the concentration of Na⁺ in the secreted fluid decreased significantly when tubules were exposed to bafilomycin A1 or amiloride. Addition of 500 μM cAMP or 1 μM 5-HT to the bath stimulated fluid secretion and resulted in a decrease in K⁺ concentration in the secreted fluid. An increase in Na⁺ concentration in the secreted fluid was observed only in cAMP-stimulated tubules. Secreted fluid pH and the transepithelial electrical potential (TEP) did not change when tubules were stimulated. Taken together, our results show that the secretion of fluid is carried out by the upper regions (DIP) in T. ni Malpighian tubules. Upper regions of the tubules secrete K⁺, whereas lower regions reabsorb it. Stimulation of fluid secretion is correlated with a decrease in the K⁺/Na⁺ ratio.