Purification and characterization of a cysteine protease inhibitor from chum salmon (Oncorhynchus keta) plasma

A cysteine protease inhibitor (CPI) in chum salmon ( Oncorhynchus keta) plasma (CSP) was detected after performing inhibitory activity staining against papain under nonreducing condition. The CPI was purified from CSP by affinity chromatography with a yield and purification ratio of 0.94% and 30.36-fold, respectively. CSP CPI had a molecular mass of 70 kDa based on the results of SDS-PAGE and Sephacryl S-100 gel filtration. CSP CPI was a glycoprotein based on the periodic acid-Schiff (PAS) staining of the SDS-PAGE gel and classified as a kininogen. CSP CPI was stable in the pH range of 6.0-9.0 with maximal stability at pH 7.0. CSP CPI presented thermal stability at temperatures below 50 degrees C and exhibited maximal activity at temperatures of 20-40 degrees C. CSP CPI was determined to be a noncompetitive inhibitor against papain, with an inhibitor constant (Ki) of 105 nM.     

Purification and characterization of a mannan-binding lectin specifically expressed in corms of saffron plant (Crocus sativus L.)

Despite the economical interest of Crocus sativus, its biochemistry has been poorly studied. Herein, we have isolated a lectin present in saffron corm by gel-filtration, anion-exchange, and reversed-phase chromatography. One- and two-dimensional PAGE, MALDI-MS, and N-terminal amino acid sequence analyses indicated that the native protein forms noncovalently linked aggregates of about 80 kDa apparent molecular mass, mainly composed of two charged heterogeneous (pI's, 6.69-6.93) basic subunits of approximately 12 kDa. Their N-terminal sequences shared 25% similarity and were homologous to the N- and C-terminal domains of monocotyledonous mannose-binding lectins, respectively. An additional polypeptide of around 28 kDa apparent molecular mass was also detected, probably corresponding to a precursor processed into two mature subunits. In addition, the N-terminal domain subunit exhibited 56% similarity with curculin, a sweet protein with taste-modifying activity. The native lectin specifically interacts with a yeast mannan and is a major corm protein specifically expressed in this organ.     

Purification and characterization of hydrolase with chitinase and chitosanase activity from commercial stem bromelain

A hydrolase with chitinase and chitosanase activity was purified from commercial stem bromelain through sequential steps of SP-Sepharose ion-exchange adsorption, HiLoad Superdex 75 gel filtration, HiLoad Q Sepharose ion-exchange chromatography, and Superdex 75 HR gel filtration. The purified hydrolase was homogeneous, as examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme exhibited chitinase activity for hydrolysis of glycol chitin and 4-methylumbelliferyl beta-D-N,N',N' '-triacetylchitotrioside [4-MU-beta-(GlcNAc)(3)] and chitosanase activity for chitosan hydrolysis. For glycol chitin hydrolysis, the enzyme had an optimal pH of 4, an optimal temperature of 60 degrees C, and a K(m) of 0.2 mg/mL. For the 4-MU-beta-(GlcNAc)(3) hydrolysis, the enzyme had an optimal pH of 4 and an optimal temperature of 50 degrees C. For the chitosan hydrolysis, the enzyme had an optimal pH of 3, an optimal temperature of 50 degrees C, and a K(m) of 0.88 mg/mL. For hydrolysis of chitosans with various N-acetyl contents, the enzyme degraded 30-80% deacetylated chitosan most effectively. The enzyme split chitin or chitosan in an endo-manner. The molecular mass of the enzyme estimated by gel filtration was 31.4 kDa, and the isoelectric point estimated by isoelectric focusing electrophoresis was 5.9. Heavy metal ions of Hg(2+) and Ag(+), p-hydroxymercuribenzoic acid, and N-bromosuccinimide significantly inhibited the enzyme activity.     

Anti-inflammatory activities of mogrosides from Momordica grosvenori in murine macrophages and a murine ear edema model

Momordica grosvenori (Luo Han Guo), grown primarily in Guangxi province in China, has been traditionally used for thousands of years by the Chinese to make hot drinks for the treatment of sore throat and the removal of phlegm. The natural noncaloric sweetening triterpenoid glycosides (mogrosides) contained in the M. grosvenori fruits are also antioxidative, anticarcinogenic, and helpful in preventing diabetic complications. The aim of this study was to assess the anti-inflammatory properties of mogrosides in both murine macrophage RAW 264.7 cells and a murine ear edema model. The results indicate that mogrosides can inhibit inflammation induced by lipopolysaccharides (LPS) in RAW 264.7 cells by down-regulating the expression of key inflammatory genes iNOS, COX-2, and IL-6 and up-regulating some inflammation protective genes such as PARP1, BCL2l1, TRP53, and MAPK9. Similarly, in the murine ear edema model, 12-O-tetradecanoylphorbol-13-acetate-induced inflammation was inhibited by mogrosides by down-regulating COX-2 and IL-6 and up-regulating PARP1, BCL2l1, TRP53, MAPK9, and PPARδ gene expression. This study shows that the anticancer and antidiabetic effects of M. grosvenori may result in part from its anti-inflammatory activity.     

Purification and characterization of a thermostable alpha-galactosidase from Thermoanaerobacterium polysaccharolyticum

Food ingredients containing alpha-1,6-galactoside bonds elicit gastrointestinal disturbances in monogastric animals, including humans. Pretreatment of such ingredients with alpha-galactosidase (EC 3.2.1.22) has the potential to alleviate this condition. For this purpose, a thermostable alpha-galactosidase from Thermoanaerobacterium polysaccharolyticum was purified by a combination of anion exchange and size exclusion chromatographies. The enzyme has a monomeric molecular weight of approximately 80 kDa; however, it is active as a dimer. The optimum temperature for enzyme activity is 77.5 degrees C. Approximately 84 and 88% of enzyme activity remained after 36.5 h of incubation at 70 and 65 degrees C, respectively. Optimum activity was observed at pH 8.0, with a broad range of activity from pH 5.0 to 9.0. Different transition metals had weak to strong inhibitory effects on enzyme activity. The K(m) and V(max) of the enzyme are 0.29-0.345 mM and 200-232 micromol/min/mg of protein, respectively. Importantly, enzyme activity was only slightly inhibited by 75-100 mM galactose, an end product of hydrolysis. Enzyme activity was specific for the alpha-1,6-galactosyl bond, and activity was demonstrated on melibiose and soy molasses.     

Purification and characterization of a new endoglucanase from Aspergillus aculeatus

Endoglucanase has been isolated from Aspergillus aculeatus. The purified enzyme showed a single band and had a molecular weight of 45,000 Da as indicated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, with a specific activity of 1.4 units/mg. The purified enzyme was identified as endoglucanase, showing a high specific activity toward CM-cellulose and low specific activity toward Avicel. The activity of the isolated enzyme was optimum at a pH of 5.0 and temperature of 40 degrees C, respectively. The isoelectric point of the enzyme was 4.3. T(m) was found to be 57 degrees C. The treatment of the endoglucanase with diethylpyrocarbonate resulted in the modification of the histidine residues present in the enzyme, with a concomitant loss of 70% of the original enzymatic activity. However, carbodiimide completely inactivated the endoglucanase. The results show that the enzyme is able to sustain 50% of its activity even when heated at 90 degrees C for a period of 5 h. Endoglucanase can be used in the controlled hydrolysis of cellulose and other cellulose-rich foods. It can be used in the development of targeted functional foods from agrimaterials for value addition in the food chain.     

Purification and characterization of a peroxidase from corn steep water

Three cationic peroxidases have been detected in early, middle, and late corn steep water, with pI values of approximately 8.9, approximately 9.5, and >10.0. The major cationic corn steep water peroxidase (CSWP), with a pI >10, was purified 36400-fold with a 12% recovery from late steep water by a combination of acetone and ammonium sulfate precipitation and sequential chromatography on CM-cellulose, phenyl-Sepharose, and Sephadex G-75. The UV-vis spectrum of purified CSWP is typical of other plant class III peroxidases. The RZ (A(403)/A(280)) of CSWP was between 2.6 and 2.9. It is not glycosylated and exhibited an M(r) of 30662 +/- 7 by MALDI-TOF MS. The pH optimum of CSWP depends on the substrate, and it is active on 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid), guaiacol, ferulic acid, o-dianisidine, o-phenylenediamine, and pyrogallol but is not active on either syringaldazine or ascorbate. At 75 degrees C and pH 4.5, the enzyme has half-lives of 22.7 min (0 mM Ca(2+)) and 248 min (1 mM Ca(2+)). The enzyme is stable at room temperature (22-25 degrees C), losing <3% of the activity at pH 4.5 and <10% at pH 6.2 over 400 h in the presence of 1 mM Ca(2+).     

Purification and characterization of a trypsin inhibitor from Plathymenia foliolosa seeds

A novel trypsin inhibitor (PFTI) was isolated from Plathymenia foliolosa (Benth.) seeds by gel filtration chromatography on a Sephadex G-100, DEAE-Sepharose, and trypsin-Sepharose columns. By SDSPAGE, PFTI yielded a single band with a M(r) of 19 kDa. PFTI inhibited bovine trypsin and bovine chymotrypsin with equilibrium dissociation constants (K(i)) of 4 x 10(-8) and 1.4 x 10(-6) M, respectively. PFTI retained more than 50% of activity at up to 50 degrees C for 30 min, but there were 80 and 100% losses of activity at 60 and 70 degrees C, respectively. DTT affected the activity or stability of PFTI. The N-terminal amino acid sequence of PFTI showed a high degree of homology with various members of the Kunitz family of inhibitors. Anagasta kuehniella is found worldwide; this insect attacks stored grains and products of rice, oat, rye, corn, and wheat. The velvet bean caterpillar (Anticarsia gemmatalis) is considered the main defoliator pest of soybean in Brazil. Diatraea saccharalis, the sugar cane borer, is the major pest of sugar cane crops, and its caterpillar-feeding behavior, inside the stems, hampers control. PFTI showed significant inhibitory activity against trypsin-like proteases present in the larval midguts on A. kuehniella and D. saccharalis and could suppress the growth of larvae.     

Purification and characterization of a novel pyrethroid hydrolase from Aspergillus niger ZD11

The pyrethroid pesticides residues on foods and environmental contamination are a public safety concern. Pretreatment with pyrethroid hydrolase has the potential to alleviate the conditions. For this purpose, a fungus capable of using pyrethroid pesticides as a sole carbon source was isolated from the soil and characterized as Aspergillus niger ZD11. A novel pyrethroid hydrolase from cell extract was purified 41.5-fold to apparent homogeneity with 12.6% overall recovery. It is a monomeric structure with a molecular mass of 56 kDa, a pI of 5.4, and the enzyme activity was optimal at 45 degrees C and pH 6.5. The activities were strongly inhibited by Hg(2+), Ag(+), and rho-chloromercuribenzoate, whereas less pronounced effects (5-10% inhibition) were observed in the presence of the remaining divalent cations, the chelating agent EDTA and phenanthroline. The purified enzyme hydrolyzed various insecticides with similar carboxylester. trans-Permethrin is the preferred substrate.     

Purification and characterization of a carboxypeptidase from squid hepatopancreas (Illex illecebrosus)

The hepatopancreas of squid (Illex illecebrosus) extract contains a wide range of carboxypeptidase (CP) activities based on hydrolysis of N-CBZ-dipeptide substrates. SDS-PAGE zymograms with N-CBZ-Phe-Leu substrate revealed three activity zones (CP-I, 23 kDa; CP-II, 29 kDa; CP-III, 42 kDa). CP-I was purified 225-fold with 86.20% recovery based on N-CBZ-Ala-Phe activity by chromatography on DEAE-cellulose, gel filtration, and chromatofocusing. The purified enzyme had broad specificity toward N-CBZ-dipeptides; however, it preferred substrates with a hydrophobic amino acid at the C terminus. CP-I had greatest activity with N-CBZ-Ala-Phe (specific activity = 7104 units/mg of protein, K(m) = 0.40 mM, and physiological efficiency = 22863). CP-I had a pI of 3.4 and is a metalloprotease that is activated by Co(2+) and partially inhibited by Pefabloc, a serine protease inhibitor. With N-CBZ-Ala-Phe and Gly-Phe, it had optimum activity at pH 8 and 70 degrees C. The amino acid composition of squid CP-I is similar to that of CP A from other species.     

Purification and characterization of a beta-glucosidase from Citrus sinensis var. Valencia fruit tissue

A preliminary survey demonstrated activity for alpha-D-glucosidase, alpha-D-mannosidase, alpha-L-arabinosidase, beta-D-glucosidase, beta-D-xylosidase, and beta-D-galactosidase in orange fruit flavedo and albedo tissue. alpha-L-Rhamnosidase was not detected. Subsequently, a beta-glucosidase was purified from mature fruit rag tissue (composed of intersegmental septa, squeezed juice sacs, and fruit core tissue) of Citrus sinensis var. Valencia. The beta-glucosidase exhibited low levels of activity against p-nitrophenyl-beta-D-fucopyranoside (13.5%) and p-nitrophenyl-alpha-D-glucopyranoside (7.0%), compared to its activity against p-nitrophenyl-beta-D-glucopyranoside (pNPG, 100%). The enzyme was purified by a combination of ion exchange (anion and cation) and gel filtration (Superdex and Toyopearl HW-55S) chromatography. It has an apparent molecular mass of 64 kDa by denaturing electrophoresis or 55 kDa by gel filtration chromatography (BioGel P-100). Hydrolysis of pNPG demonstrated a pH optimum between 4.5 and 5.5. At pH 5.0 the temperature optimum was 40 degrees C. At pH 5.0 and 40 degrees C the K(m) for pNPG was 0.1146 mM and it had a V(max) of 5.2792 nkatal x mg(-1) protein (katal = 0.06 International Units = the amount of enzyme that produces, under standard conditions, one micromol of product per min). Of the substrates tested, the enzyme was most active against the disaccharide cellobiose (1-->4), but was not active against p-nitrophenyl-beta-D-cellobioside. High levels of activity also were observed with the disaccharides laminaribiose (1-->3), gentiobiose (1-->6), and sophorose (1-->2). Activity greater than that observed with pNPG was obtained with the flavonoids hesperetin-7-glucoside and prunin (naringenin-7-glucoside), salicin, mandelonitrile-beta-D-glucoside (a cyanogenic substrate), and sinigrin (a glucosinolate). The enzyme was not active against amygdalin, coniferin, or limonin glucoside.     

Preparation and characterization of chitosan-based nanocomposite films with antimicrobial activity

Four different types of chitosan-based nanocomposite films were prepared using a solvent-casting method by incorporation with four types of nanoparticles, that is, an unmodified montmorillonite (Na-MMT), an organically modified montmorillonite (Cloisite 30B), a Nano-silver, and a Ag-zeolite (Ag-Ion). X-ray diffraction patterns of the nanocomposite films indicated that a certain degree of intercalation was formed in the nanocomposite films, with the highest intercalation in the Na-MMT-incorporated films followed by films with Cloisite 30B and Ag-Ion. Scanning electron micrographs showed that in all of the nanocomposite films, except the Nano-silver-incorporated one, nanoparticles were dispersed homogeneously throughout the chitosan polymer matrix. Consequently, mechanical and barrier properties of chitosan films were affected through intercalation of nanoparticles, that is, tensile strength increased by 7-16%, whereas water vapor permeability decreased by 25-30% depending on the nanoparticle material tested. In addition, chitosan-based nanocomposite films, especially silver-containing ones, showed a promising range of antimicrobial activity.     

Purification and characterization of a corrinoid-compound in an edible cyanobacterium Aphanizomenon flos-aquae as a nutritional supplementary food

The vitamin B12 concentration of the dried cells of Aphanizomenon flos-aquae was determined by both microbiological method with Lactobacillus delbrueckeii ATCC7830 and chemiluminescence method with intrinsic factor. The Aphanizomenon cells contained 616.3 +/- 30.3 micro g (n = 4) of vitamin B12 per 100 g of the dried cells by the microbiological method. The values determined with the chemiluminescence method, however, were only about 5.3% of the values determined by the microbiological method. A corrinoid-compound was purified from the dried cells and characterized. The purified corrinoid-compound was identified as pseudovitamin B12 (an inactive corrinoid-compound for humans) by silica gel 60 TLC, C18 reversed-phase HPLC, ultraviolet-visible spectroscopy, and 1H NMR spectroscopy. The results suggest that the Aphanizomenon cells are not suitable for use as a vitamin B12 source, especially in vegans.     

Purification, crystallization, and preliminary X-ray characterization of a 36 kDa amaranth globulin.

The purpose of this study was to purify, crystallize, and characterize by X-ray diffraction an amaranth globulin for its subsequent structure elucidation. A 36-kDa amaranth globulin was extracted by sequential precipitation and purified by gel filtration and cationic exchange columns. It was crystallized at 18 degrees C from 4 M sodium formate. Suitable crystals for X-ray analysis were found to belong to the tetragonal crystal system with cell dimensions of a = b = 195.5 A and c = 164.14 A. Two possible tetragonal space groups P4(1)2(1)2 or P4(3)2(1)2 were determined. The crystals diffracted up to 2.5 A.     

Purification and characterization of a milk-clotting aspartic proteinase from globe artichoke (Cynara scolymus L.)

The study of proteinase expression in crude extracts from different organs of the globe artichoke (Cynara scolymus L.) disclosed that enzymes with proteolytic and milk-clotting activity are mainly located in mature flowers. Maximum proteolytic activity was recorded at pH 5.0, and inhibition studies showed that only pepstatin, specific for aspartic proteinases, presented a significant inhibitory effect. Such properties, in addition to easy enzyme inactivation by moderate heating, make this crude protease extract potentially useful for cheese production. Adsorption with activated carbon, together with anion exchange and affinity chromatography, led to the isolation of a heterodimeric milk-clotting proteinase consisting of 30- and 15-kDa subunits. MALDI-TOF MS of the 15-kDa chain determined a 15.358-Da mass, and the terminal amino sequence presented 96% homology with the smaller cardosin A subunit. The amino terminal sequence of the 30-kDa chain proved to be identical to the larger cardosin A subunit. Electrophoresis evidenced proteinase self-processing that was confirmed by immunoblots presenting 62-, 30-, and 15-kDa bands.     

Purification and characterization of a latent polyphenol oxidase from beet root (Beta vulgaris L.)

Polyphenol oxidase (PPO) has been extracted from beet root, in both soluble and membrane fractions. In both cases, the enzyme was in its latent state, and it was activated by sodium dodecyl sulfate. PPO was purified to apparent homogeneity. The soluble PPO purification was achieved by hydrophobic interaction chromatography and gel filtration chromatography, with apparent molecular mass of 55 kDa. The membrane PPO purification was achieved by anion exchange chromatography and gel filtration with apparent molecular mass of 54 kDa. A totally denaturing SDS-PAGE indicated the presence of a single polypeptide with an apparent molecular mass of 60 kDa for both fractions, with the band also revealed by Western blot. A partially denaturing SDS-PAGE stained a single active 36 kDa band for both fractions. Under native isoelectric focusing, a major acidic band of pH 5.2 was detected in both fractions. Kinetic characterization of PPO on the natural substrate l-dopa was carried out.     

Purification and characterization of a stable cysteine protease ervatamin B, with two disulfide bridges, from the latex of Ervatamia coronaria

Latex of the medicinal plant Ervatamia coronaria was found to contain at least three cysteine proteases with high proteolytic activity, called ervatamins. One of these proteases, named ervatamin B, has been purified to homogeneity using ion-exchange chromatography and crystallization. The molecular mass of the enzyme was estimated to be 26 000 Da by SDS-PAGE and gel filtration. The extinction coefficient (epsilon(1%)(280 nm)) of the enzyme was 20.5 with 7 tryptophan and 10 tyrosine residues per molecule. The enzyme hydrolyzed denatured natural substrates such as casein, azoalbumin, and azocasein with a high specific activity. In addition, it showed amidolytic activity toward N-succinyl-alanine-alanine-alanine-p-nitroanilide with an apparent K(m) and K(cat) of 6.6 +/- 0.5 mM and 1.87 x 10(2) s(-)(1), respectively. The pH optima was 6.0-6.5 with azocasein as substrate and 7.0-7.5 with azoalbumin as substrate. The temperature optimum was around 50-55 degrees C. The enzyme was basic with an isoelectric point of 9.35 and had no carbohydrate content. Both the proteolytic and amidolytic activity of the enzyme was strongly inhibited by thiol-specific inhibitors. Interestingly, the enzyme had only two disulfide bridges versus three as in most plant cysteine proteases of the papain superfamily. The enzyme was relatively stable toward pH, denaturants, temperature, and organic solvents. Polyclonal antibodies raised against the pure enzyme gave a single precipitin line in Ouchterlony's double immunodiffusion and typical color in ELISA. Other related proteases do not cross-react with the antisera to ervatamin B showing that the enzyme is immunologically distinct. The N-terminal sequence showed conserved amino acid residues and considerable similarity to typical plant cysteine proteases.     

Purification and characterization of a polyphenol oxidase from the seeds of field bean (Dolichos lablab)

The polyphenol oxidase from field bean (Dolichos lablab) seeds has been purified to apparent homogeneity by a combination of ammonium sulfate precipitation, DEAE-Sephacel chromatography, phenyl agarose chromatography, and Sephadex G-200 gel filtration. The purified enzyme has a molecular weight of 120 +/- 3 kDa and is a tetramer of 30 +/- 1.5 kDa. Native polyacrylamide gel electrophoresis of the purified enzyme revealed the presence of a single isoform with an observed pH optimum of 4.0. 4-Methyl catechol is the best substrate, followed by catechol, and L-3,4-dihydroxyphenylalanine, all of which exhibited a phenomenon of inhibition by excess substrate. No activity was detected toward chlorogenic acid, catechin, caffeic acid, gallic acid, and monophenols. Tropolone, both a substrate analogue and metal chelator, proved to be the most effective competitive inhibitor with an apparent K(i) of 5.8 x 10(-)(7) M. Ascorbic acid, metabisulfite, and cysteine were also competitive inhibitors.     

HPLC-PDA-MS and NMR characterization of C-glycosyl flavones in a hydroalcoholic extract of Citrus aurantifolia leaves with antiplatelet activity

A hydroalcoholic extract of lime ( Citrus aurantifolia) leaves has been developed in Cuba to be used as a nutritional supplement and phytomedicine in the form of tincture (TLL). A HPLC-PDA-ESI/MS/MS method has been used for the comprehensive analysis of C-glycosyl flavones in TLL. Six C-glycosyl flavones were characterized and, to confirm the proposed structures and to elucidate the nature of the sugar units, a preparative procedure was applied, and isolated compounds were characterized by NMR. Apigenin-6,8-di-C-beta-D-glucopyranoside (vicenin II) (1), diosmetin-6,8-di- C-beta- d-glucopyranoside (2), apigenin-8-C-beta-D-glucopyranoside (vitexin) (3), apigenin-8-C-[alpha-L-arabinopyranosyl-(1-->6)]-O-beta-D-glucopyranoside (4), apigenin-6-C-[alpha-l-arabinopyranosyl-(1-->6)]-O-beta-D-glucopyranoside (5). and apigenin-6-C-beta-D-glucopyranoside (isovitexin) (6) were identified in TLL and quantified by HPLC-PDA. Compounds 4 and 5 were two new arabinosyl derivatives of vitexin and isovitexin. Inhibitor effect of TLL on platelet aggregation induced by physiological agonists of platelets was evaluated in human plasma. TLL inhibited significantly ADP and epinephrine-induced platelet aggregation in a concentration-dependent manner (IC 50=0.40 and 0.32 mg/mL, respectively).     

Purification of a lectin from the marine red alga Gracilaria cornea and its effects on the cattle tick Boophilus microplus (Acari: Ixodidae)

A lectin was purified from the seaweed Gracilaria cornea by hydrophobic interaction chromatography on phenyl-Sepharose CL-4B followed by affinity chromatography on immobilized mucin. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of G. cornea lectin (GCL) revealed a single protein band of approximately 60 kDa, whereas by gel filtration on Sephadex G-100 its native molecular mass was 66 kDa. GCL exhibited a single isoeletric point of 4.3 and a 52.5% content of neutral sugars. Furthermore, the EDTA-treated lectin did not show any significant decrease in its ability to agglutinate trypsin-treated chicken erythrocytes. These data suggest that GCL is an acidic, monomeric glycoprotein that probably does not require divalent metal ions for its hemagglutinating activity. GCL hemagglutinating activity was not inhibited by any of the mono-, di-, and trisaccharides tested but was by the complex glycoproteins fetuin and porcine stomach mucin. Exposure of engorged females of the cattle tick (Boophilus microplus) to 0.1 mg mL(-1) GCL significantly (P < 0.05) reduced the female weight after the oviposition period, the egg mass weight, the hatching period, and the mean larvae survival time.