Extracellular Phenol Oxidase Research Articles

Исследование влияния солей металлов на активность ферментов фенолоксидазного комплекса бактерий рода Azospirillum

Recently, much attention has been paid to the development of technologies for biodegradation of organopollutants and the search for promising biodestructors. The environmental accumulation of lignin-like compounds and synthetic dyes poses a huge threat not only to ecosystems and biodiversity, but also to human health. Phenol oxidases are enzymes with broad substrate specificity, with oxidizing ability towards various polyphenols and aromatic amines. Therefore the use of phenol oxydases as bioremediation agents is promising due to their unique catalytic properties. In this work we present the results of a study of the effect of metal ions on the activity of the azospirilla phenol oxidase complex. It was demonstrated that extracellular laccases of lignin- and Mn-peroxidases of strains Azospirillum baldaniorum Sp245 and Azospirillum brasilense SR80 are quite stable in the presence of the studied metal salts. The enzymatic activity decreased and the effectiveness of the organopollutants’ biodegradation efficacy was inhibited in the presence of Zn2+ ions. The laccase and lignin-peroxidase activity induced by copper ions positively correlated with the ability of lignin degradation by azospirillum. Analysis of the obtained data showed that inhibitors and inducers of authentic extracellular phenol oxidases of both fungi and bacteria are typical for azospirillum enzymes.

Enzymatic and Antimicrobial Activities in Polar Strains of Microscopic Soil Fungi.

Potential to produce inducible enzymes (several hydrolases and oxidases) and antibiotics as secondary metabolites was studied in soil micromycete strains from the Arctic (Franz Josef Land and Novaya Zemlya) and Antarctica (the oases Thala Hills, Larsemann Hills, Schirmacher, and Marie Byrd Land). Maximal esterase activity was observed in strains of two typical Antarctic species, Hyphozyma variabilis 218 and Thelebolus ellipsoideus 210 (51 and 29 nmol FDA/((g myceliumh), respectively). Cellulolytic activity was maximal (89 µmol glucose/mg biomass) in Ascochyta pisi 192. Extracellular phenol oxidase (laccase) and peroxidase activities were not detected in the strains examined. Antibacterial activity toward Bacillus subtilis ATCC 6633 was observed in 75% of the Antarctic micromycete strains. Higher-activity strains were isolated from organic-rich moist habitats with a moss or lichen cover. Maximal activities were displayed by Paecilomyces marquandii 166, Penicillium janczewskii 165, Penicillium roseopurpureum 169, and Thelebolus ellipsoideus 210. Antagonistic activity toward Antarctic bacterial strains was shown by 77% of the microfungal strains examined. Maximal inhibition was observed with strains of the typical Antarctic species Antarctomyces psychrotrophicus MT303855 and the eurytopic species Sarocladium kiliense MT303856. Antimycotic activity was observed in 42% of the strains. Both activities were detected in 38% of the Antarctic strains.

Differential Activity of the Extracellular Phenoloxidases in Different Strains of the Phytopathogenic Fungus, Microdochium nivale.

To cause plant diseases, phytopathogenic fungi use numerous extracellular enzymes, among which, the phenoloxidases (POs) seem underestimated for the pathogens of non-woody plants. Our study aimed to (1) compare extracellular PO activities (lignin peroxidase, Mn peroxidase, laccase, and tyrosinase) in differentially virulent strains (inhabiting winter rye in a single field) of the phytopathogenic species, Microdochium nivale; (2) check whether these activities are responsive to host plant metabolites; and (3) search for correlations between the activities, lignin-decomposing capacity, and virulence. All strains displayed all four enzymatic activities, but their levels and dynamics depended on the particular strain. The activities displayed the hallmarks of co-regulation and responsiveness to the host plant extract. No relationships between the virulence of strains and levels of their extracellular PO activities or lignin-degrading capacity were revealed. We consider that different strains may rely on different POs for plant colonization, and that different POs contribute to the “uniqueness” of the enzymatic cocktails that are delivered into host plant tissues by different virulent strains of M. nivale. Our study supports the hypothesis of the differential behavior of closely related M. nivale strains, and discusses an important role of POs in the interactions of phytopathogens with herbaceous plants.

Vitamin C elicits the activation of immunological responses in swimming crab (Portunus trituberculatus) hemocytes against Mesanophrys sp.

Under suitable conditions, Vitamin C (vit. C) is essential for immunological responses and positively influences the function of stimulated immune cells. Therefore, this study evaluated the specific mechanisms of innate immunity in crab after supplementing the cell culture media with vit. C. We used the primary culture media of swimming crab hemocytes to detect some immune factors and immune-related signaling pathways via 24 h stimulation in vitro assays. Besides, we evaluated the anti-parasitic action of these hemocytes. Herein, the cellular and humoral factors, including intra- and extracellular phenol oxidase, nitrite oxidase synthase, lysozyme, and intracellular O2− were significantly (P < 0.05) enhanced after the culture media was supplemented with vit. C as compared to control. A two-slope broken-line analysis indicated that the optimum vit. C level for intracellular O2− was 118.91 μg mL−1 of the cell culture media (R2 = 0.73). However, for the extracellular O2−, the vit. C supplementation showed a downward quadratic relationship (R2 = 0.59). qRT-PCR demonstrated that relative mRNA expression of immune-related genes, including proPO, serine proteases, a2M, and alf-1 was significantly (P < 0.05) increased with vit. C media supplementation. Further, the results indicated that the anti-parasitic response and survival rate (%) of hemocytes after Mesanophrys sp. challenge was also significantly (P < 0.05) augmented in media supplemented with vit. C, relative to the control. Thus, the different vit. C in vitro levels were necessary for optimal immunological responses of crab hemocytes. In summary, vit. C supplementation enhanced and modulated the innate immune response of hemocytes against Mesanophrys sp., highlighting its potential application as an immunostimulant.

Low Biodegradability of Dissolved Organic Matter From Southeast Asian Peat‐Draining Rivers

AbstractSoutheast Asia's extensive tropical peatlands account for a significant proportion of the global riverine dissolved organic carbon (DOC) flux to the ocean. Peat‐derived DOC is rich in polyphenolic compounds, the microbial degradation of which is thought to rely on extracellular phenol oxidases. Despite substantial interest in the biogeochemical fate of terrigenous DOC (tDOC), few studies have quantified phenol oxidase activity in aquatic environments, and microbial remineralization rates of tDOC have never been measured in Southeast Asia. Here, we assess the potential for using phenol oxidase assays as a proxy for tDOC biodegradation across peat‐draining rivers and the coastal waters of Sarawak, Borneo, and report experimental measurements of microbial tDOC remineralization rates from this region. We first show that phenol oxidase assays in aquatic samples are problematic because of the rapid, pH‐dependent autoxidation of the assay substrate. Our field measurements of phenol oxidase activity detected only substrate autoxidation, suggesting that real phenol oxidase activity was low or absent. Second, we report that peatland tDOC, collected from one of the few remaining intact peatlands on Borneo, showed at most very limited biodegradation (0%–6% loss of DOC, and 0%–12% loss of colored dissolved organic matter) during several 56‐day incubation experiments at an in situ temperature of ∼30°C, even when diluted with seawater or amended with nutrients. Our results suggest that direct microbial respiration is perhaps not a major pathway for peatland tDOC remineralization in Southeast Asia and that photo‐oxidation is more likely to control the fate of this carbon.

Scenedesmus vacuolatus cultures for possible combined laccase-like phenoloxidase activity and biodiesel production

A key aspect of the industrial development of microalgal production processes is the excessive cost of biomass production. A solution is a combination of biodiesel production and wastewater treatment. The microalga Scenedesmus has a high lipid content and a potential extracellular phenoloxidase activity, which could improve the phycoremediation of phenolic pollutants. In this work, the most suitable growth conditions to obtain this twofold aim were analyzed. First, different strains of Scenedesmus vacuolatus microalga were tested at different pH, salinity and CO2 concentration in the gas phase. The two most promising strains were then cultivated in autotrophic and heterotrophic conditions, and were investigated in terms of efficient nitrogen removal, fatty acid profile and maximized extracellular phenoloxidase activity in the medium. The results showed two extreme conditions: (1) biomass productivity doubled when photobioreactors were sparged with 5% CO2 supplemented air with respect to cultures sparged with air (the steady state values of strain 53 were 0.138 g L−1 day−1 in the presence of air, and 0.243 in the presence of CO2 addition), and N-starvation under 5% CO2enhanced the transesterified fraction of lipids (strain 53 FAME fraction in the presence of N-starvation was 33%, in the presence of nitrogen FAME fraction was 22%); (2) phenoloxidase activity was completely suppressed by presence of 5% CO2 in the gas phase (strain 53 0.21 U mL−1), indicating clear catabolite repression for the induction of this enzyme in the algal metabolism.

Ligninolytic Activity at 0°C of Fungi on Oak Leaves Under Snow Cover in a Mixed Forest in Japan.

Despite the importance of litter decomposition under snow cover in boreal forests and tundra, very little is known regarding the characteristics and functions of litter-decomposing fungi adapted to the cold climate. We investigated the decomposition of oak leaves in a heavy snowfall forest region of Japan. The rate of litter weight loss reached 26.5% during the snow cover period for 7months and accounted for 64.6% of the annual loss (41.1%). Although no statistically significant lignin loss was detected, decolourization portions of oak leaf litter, which was attributable to the activities of ligninolytic fungi, were observed during snow cover period. This suggests that fungi involved in litter decomposition can produce extracellular enzymes to degrade lignin that remain active at 0°C. Fungi were isolated from oak leaves collected from the forest floor under the snow layer. One hundred and sixty-six strains were isolated and classified into 33 operational taxonomic units (OTUs) based on culture characteristics and nuclear rDNA internal transcribed spacer (ITS) region sequences. To test the ability to degrade lignin, the production of extracellular phenoloxidases by isolates was quantified at 0°C. Ten OTUs (9 Ascomycota and 1 Basidiomycota) of fungi exhibited mycelial growth and ligninolytic activity. These results suggested that some litter-decomposing fungi that had the potential to degrade lignin at 0°C significantly contribute to litter decomposition under snow cover.

Are colorimetric assays appropriate for measuring phenol oxidase activity in peat soils?

The activity of extracellular phenol oxidases is believed to play a critical role in decomposition processes in peatlands. The water logged, acidic conditions, and recalcitrant litter from the peatland vegetation, lead to exceptionally high phenolics in the peat. In order to quantify the activity of oxidative enzymes involved in the modification and break down of phenolic compounds two types of assays are primarily utilized: L-DOPA and ABTS. This note focuses on the strengths and weaknesses of both approaches. Both assays involve a redox reaction and the resulting oxidized chromophore is measured spectrophotometrically. However, in the presence of reducing agents such as the phenolics commonly found in peat the colorimetric reaction is reversed and cannot be used to quantify phenol oxidase activity.

Diversity and Distribution of Phenol Oxidase Producing Fungi from Soda Lake and Description of Curvularia lonarensis sp. nov.

Soda lake is hyper alkaline and saline habitat located in closed craters with high evaporation rate. In current study fungal diversity from water and sediment samples of a soda lake (Lonar lake) located in Buldhana district of Maharashtra, India was investigated using extensive culturomics approach and mimicking the natural conditions of Lonar lake in culture media. A total of 104 diverse isolates of extremophilic fungi were recovered from this study and phylogenetically characterized by internal transcribed spacer (ITS) region sequencing. In addition, due to important role of phenol oxidase, and peroxidase in degradation of toxic phenol, lignin, etc., all isolated pure cultures were also screened for extracellular phenol oxidase and peroxidase production potential. Diversity analysis indicated that different groups of extremophilic fungi are present in the water and sediment samples of Lonar lake. A total of 38 species of fungi belonging to 18-different genera were recovered. Out of 104 isolates 32 showed ≤97% sequences similarity, which were morphologically different and could be potential novel isolates of extremophilic fungi. However, out of 104 isolates only 14 showed the extracellular phenol oxidase production potentials at alkaline pH. Curvularia sp. strain MEF018 showed highest phenol oxidase production at alkaline condition and had low sequence similarity with previously characterized species (96% with Curvularia pseudorobusta). Taxonomic characterization (morphological and physiological) and multi locus sequence analysis (MLSA) using combined alignment of ITS-LSU-gpd of strain MEF018 showed that it is a novel species of the genus Curvularia and hence proposed as Curvularia lonarensis sp. nov.

Transformation of triclosan by laccase catalyzed oxidation: The influence of humic acid-metal binding process

Laccase is a widely present extracellular phenoloxidase excreted by fungi, bacteria, and high plants. It is able to catalyze one-electron oxidation of phenolic compounds into radical intermediates that can subsequently couple to each other via covalent bonds. These reactions are believed to play an important role in humification process and the transformation of contaminants containing phenolic functionalities in the environment. In this study, we investigated the kinetics of triclosan transformation catalyzed by laccase. It was found that the rate of triclosan oxidation was first order to the concentrations of both substrate and enzyme. Humic acid (HA) could inhibit the reaction by quenching the radical intermediate of triclosan generated by laccase oxidation. Such inhibition was more significant in the presence of divalent metal cations. This is because that binding to metal ions neutralized the negative charge of HA molecules, thus making them more accessible to laccase molecule that is also negatively charged. Therefore, it has greater chance to quench the radical intermediate that is very unstable and can only diffuse a limited distance after being released from the enzyme catalytic center. Based on these understandings, a reaction model was developed by integration of metal-HA binding equilibriums and kinetic equations. This model precisely predicted the transformation rate of triclosan in the presence of HA and divalent metal ions including Ca2+, Mg2+, Cd2+, Co2+, Mn2+, Ba2+, and Zn2+. Overall, this work reveals important insights into laccase catalyzed oxidative coupling process.

Correlation between white-rot fungi potential to production of extracellular phenol oxidases and bioremediation of aquatic environments polluted with pharmaceuticals

Correlation between white-rot fungi potential to production of extracellular phenol oxidases and bioremediation of aquatic environments polluted with pharmaceuticals

First laccase in green algae: purification and characterization of an extracellular phenol oxidase from Tetracystis aeria.

A green algal phenol oxidase was firstly purified, confirmed to be a laccase, and a hetero-oligomeric quaternary structure is suggested. The operation of a laccase-mediator system is firstly described in algae. Laccases (EC 1.10.3.2) catalyze the oxidation of a multitude of aromatic substrates. They are well known in higher plants and fungi, while their presence in green algae appears uncertain. Extracellular laccase-like enzyme activity has previously been described in culture supernatants of the green soil alga Tetracystis aeria [Otto et al. in Arch Microbiol 192:759-768, (2010)]. As reported herein, the T. aeria enzyme was purified 120-fold by employing a combination of anion exchange and size exclusion chromatography. The purified enzyme was confirmed to be a laccase according to its substrate specificity. It oxidizes 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), syringaldazine, and 2,6-dimethoxyphenol (pH optima of pH ≤ 2.5, 7.0, and 6.5; K m values of 28.8, 40.5, and 1,830 µM; respectively), but not L-tyrosine or Fe(2+). ABTS is by far the most efficient substrate. Two polypeptides, A (~110 kDa) and B (71 kDa), were co-purified by the applied procedure, both being highly N-glycosylated (≥~53 and ≥ 27 %, respectively). As suggested by various gel electrophoretic analyses, the native enzyme (apparent molecular mass of ~220 kDa) most probably is a hetero-oligomer with the composition AB 2 , wherein A is the catalytic subunit and B forms a disulfide-linked homo-dimer B2. The decolorization of anthraquinone (Acid Blue 62 and Remazol Brilliant Blue R) and diazo dyes (Reactive Black 5) was studied in the presence of redox-mediating compounds (ABTS and syringaldehyde), demonstrating the operation of the laccase-mediator system in algae for the first time. Thus, laccases from green algae may participate in the biotransformation of a wide spectrum of natural and xenobiotic compounds.

Diversity of Two-Domain Laccase-Like Multicopper Oxidase Genes in Streptomyces spp.: Identification of Genes Potentially Involved in Extracellular Activities and Lignocellulose Degradation during Composting of Agricultural Waste

Traditional three-domain fungal and bacterial laccases have been extensively studied for their significance in various biotechnological applications. Growing molecular evidence points to a wide occurrence of more recently recognized two-domain laccase-like multicopper oxidase (LMCO) genes in Streptomyces spp. However, the current knowledge about their ecological role and distribution in natural or artificial ecosystems is insufficient. The aim of this study was to investigate the diversity and composition of Streptomyces two-domain LMCO genes in agricultural waste composting, which will contribute to the understanding of the ecological function of Streptomyces two-domain LMCOs with potential extracellular activity and ligninolytic capacity. A new specific PCR primer pair was designed to target the two conserved copper binding regions of Streptomyces two-domain LMCO genes. The obtained sequences mainly clustered with Streptomyces coelicolor, Streptomyces violaceusniger, and Streptomyces griseus. Gene libraries retrieved from six composting samples revealed high diversity and a rapid succession of Streptomyces two-domain LMCO genes during composting. The obtained sequence types cluster in 8 distinct clades, most of which are homologous with Streptomyces two-domain LMCO genes, but the sequences of clades III and VIII do not match with any reference sequence of known streptomycetes. Both lignocellulose degradation rates and phenol oxidase activity at pH 8.0 in the composting process were found to be positively associated with the abundance of Streptomyces two-domain LMCO genes. These observations provide important clues that Streptomyces two-domain LMCOs are potentially involved in bacterial extracellular phenol oxidase activities and lignocellulose breakdown during agricultural waste composting.

A correlation between the fate and non-extractable residue formation of 14C-metalaxyl and enzymatic activities in soil

Extracellular, oxidative soil enzymes like monophenol oxidases and peroxidases play an important role in transformation of xenobiotics and the formation of organic matter in soil. Additionally, these enzymes may be involved in the formation of non-extractable residues (NERs) of xenobiotics during humification processes. To examine this correlation, the fate of the fungicide 14C metalaxyl in soil samples from Ultuna (Sweden) was studied. Using different soil sterilization techniques, it was possible to differentiate between free, immobilized, and abiotic (“pseudoenzyme”-like) oxidative activities. A correlation between the formation of metalaxyl NER and soil organic matter content, biotic activities, as well as extracellular phenoloxidase and peroxidase activities in the bulk soil and its particle size fractions was determined. Extracellular soil-bound enzymes were involved in NER formation (up to 8% of applied radioactivity after 92 days) of the fungicide independently from the presence of living microbes and different distributions of the NER in the soil humic subfractions.

Erratum to: The roles of serine protease, intracellular and extracellular phenoloxidase in activation of prophenoloxidase system, and characterization of phenoloxidase from shrimp haemocytes induced by lipopolysaccharide or dopamine

Erratum to: The roles of serine protease, intracellular and extracellular phenoloxidase in activation of prophenoloxidase system, and characterization of phenoloxidase from shrimp haemocytes induced by lipopolysaccharide or dopamine

The roles of serine protease, intracellular and extracellular phenoloxidase in activation of prophenoloxidase system, and characterization of phenoloxidase from shrimp haemocytes induced by lipopolysaccharide or dopamine

We investigated the effects of lipopolysaccharide (LPS) and dopamine (DA) on the activation of the prophenoloxidase (proPO) system of Litopenaeus vannamei. LPS and DA were shown with a negative dose-dependent effect on hyalne cells (HC), semi-granular cells (SGC), large granular cells (LGC), and total haemocyte count (THC). When haemocytes were treated with LPS or DA, serine proteinase activity and intracellular phenoloxidase (PO) activity were significantly reduced, but extracellular PO activity increased significantly. These findings indicated that the reduction in haemocyte counts was mainly because of the degranulation and activation of the proPO system from semi-granule and large granule cells. The PKC inhibitor, chelerythrine, and the TPK inhibitor, genistein, had an inhibitory effect on extracellular PO activity, while serine proteinase and intracellular PO activity increased. This suggests that the LPS and DA induce the activation of proPO in haemocytes via PKC and TPK-related signaling pathways, but serine proteinase may be activated only by PKC, as the genistein effects were not statistically significant. Electrophoresis analysis revealed that POs induced by LPS or DA have the same molecular mass and high diphenolase activity. Two PO bands at 526 kDa and 272 kDa were observed in PAGE, while in the haemocyte lysate supernatant (HLS), only a 272-kDa band was observed. This band was resolved after SDS-PAGE under non-reducing and reducing conditions into two groups of POs, 166 kDa and 126 kDa, and 78.1 kDa and 73.6 kDa, respectively, suggesting that PO in L. vannamei is an oligomer, which may have different compositions intra- and extracellularly.

Inhibitory effects of drought induced acidification on phenol oxidase activities in Sphagnum-dominated peatland

Extracellular phenol oxidases play an important role in the soil carbon cycle. How phenol oxidase activity in peatlands will respond to increased aeration induced by water-table drawdown or droughts if global warming continues remains unclear. We investigated the short-term effects of drought induced acidification on phenol oxidase activities of a Sphagnum-dominated peatland in central China. The results indicated a significant acidification induced by drought, with a decline in the average pH value from 5.67 ± 0.08 under waterlogging to 4.84 ± 0.08 in droughted peat. In addition, drought-treated peat had a lower average value of phenol oxidase activities compared to the waterlogging-treated peat (mean values of 0.1404 ± 0.0305 and 0.2623 ± 0.0324 μmol diqc min−1 g−1, respectively). The decreased phenol oxidase activities were mainly attributed to the acidification induced by drought. However, the declines in phenol oxidase activity did not lead to the reduction of the degradation in the deeper peat. Under drought conditions, a considerable part of the adsorbed phenolics, consisting primarily of high molecular-weight organic substances with phenolic groups, were further degraded into smaller-sized or lower molecular weight hydrophilic fractions in the vicinity of the water table. Our results highlight the important role of the water table position in peat decomposition processes. Even with lower activities of phenol oxidase, the decomposition processes were able to effectively keep on going as long as there was sufficient supply of water, oxygen and nutrients.

Phenoloxidase-producing halotolerant fungi from olive brine wastewater

The main aim of this study was to assess the ability of producing extracellular phenoloxidases (EPO) of fungi isolated from olive brine wastewater (OBW), the effluent from the debittering process of table olives. Five out of twenty isolates displayed EPO-producing ability as assessed by selective agar plate assays and were all halotolerant. Among them, Citeromyces matritensis (syn. Candida globosa) and Aspergillus fumigatus concomitantly produced laccase and Mn-peroxidase (MnP) activities. Candida boidinii and Candida bombi, instead, only produced laccase and Candida diddensiae only released MnP. Both C. matritensis and A. fumigatus, grown in shaken cultures on OBW, maintained their EPO-producing ability and removed phenols by 52.3 and 82.3%, respectively, after 10d incubation in the non-supplemented effluent. These results might suggest the possible use of these strains in the treatment of other saline phenol-rich effluents, such as pickling and tannery wastewater.

Experimental climate effect on seasonal variability of polyphenol/phenoloxidase interplay along a narrow fen–bog ecological gradient in Sphagnum fallax

AbstractExtracellular phenoloxidase enzymes play an important role in the stability of soil carbon storage by contributing to the cycling of complex recalcitrant phenolic compounds. Climate warming could affect peatland functioning through an alteration of polyphenol/phenoloxidase interplay, which could lead them to becoming weaker sinks of carbon. Here, we assessed the seasonal variability of total phenolics and phenoloxidases subjected to 2–3 °C increase in air temperature using open‐top chambers. The measurements were performed along a narrow fen–bog ecological gradient over one growing season. Climate warming had a weak effect on phenoloxidases, but reduced phenolics in both fen and bog areas. Multivariate analyses revealed a split between the areas and also showed that climate warming exacerbated the seasonal variability of polyphenols, culminating in a destabilization of the carbon cycle. A negative relationship between polyphenols and phenoloxidases was recorded in controls and climate treatments suggesting an inhibitory effect of phenolics on phenoloxidases. Any significant decrease of phenolics through repeatedly elevated temperature would greatly impact the ecosystem functioning and carbon cycle through an alteration of the interaction of polyphenols with microbial communities and the production of extracellular enzymes. Our climate treatments did not have the same impact along the fen–bog gradient and suggested that not all the peatland habitats would respond similarly to climate forcing.

Haemocyte phagocytosis, exocytosis and their signal transduction in white shrimp (&lt;I&gt;Litopenaeus vannamei&lt;/I&gt;) induced by lipopolysaccharide and dopamine

Effects of bacterial lipopolysaccharide ( LPS) and dopamine ( DA) on phagocytic activity, exocytosis and signal transduction of haemocytes in white shrimp (Litopenaeus vannamei) were studied. The haemocytes treated with LPS (1 - 10 mg/L) and dopamine (1 - 10 μmol/L) in vitro showed conspicuous dose-dependent decrease in cell count,and increase in extracellular phenoloxidase (PO) activity. Phagocytic activity of haemocytes was promoted by LPS and inhibited by dopamine. Meanwhile,the addition of chelerythrine (a PKA inhibitor) and genistein (a TPK inhibitor) to haemocytes can inhibit phagocytosis and exocytosis induced by LPS,and the inhibitory efficiencies on phagocytosis and exocytosis respectively were: genistein chelerythrine,chelerythrine genistein. Dopamine-induced cell phagocytosis was strenghtend by H-89 (a PKA inhibitor),its exocytosis was inhibited by chelerythrine and genistein,and the efficiency was chelerythrine genistein,and the two inhibitors had no impact on dopamine-induced phagocytosis.