ME Extracts Research Articles

Fijian medicinal plants and their role in the prevention of Type 2 diabetes mellitus.

Medicinal plants (MPs) are natural sources of active compounds with potential therapeutic benefits in alleviating various illnesses for decades. Fijian people also are using these MPs for the management/prevention of Type 2 diabetes mellitus (T2DM) and associated complications. However, till date, none of these Fijian MP's antidiabetic potential have been explored or evaluated. Here, we investigated the antidiabetic potential of Fijian MPs scientifically. Phytochemicals such as polyphenols were detected to inhibit the activity of α-amylase and α-glucosidase, the two key carbohydrate enzymes linked to T2DM. Therefore, in the present study, the total phenolic content (TPC), α-amylase and α-glucosidase inhibitory activity of five Fijian MPs: Vobo (Mussaenda raiateensis, MR), Vula walu (Blechnum orientale, BO), Gasau (Miscanthus floridulus, MF), Molikaro (Citrus limon, CL) and Beki ni sina (Dicranopteris caudate, DC) collected from mainland region of Vitilevu, Fiji Islands, were evaluated in vitro. The hydromethanolic (ME) and dichloromethane (DM) extracts of these selected MPs were investigated. The ME extracts of BO (0.102 ± 0.009 mM CE) and DC (0.098 ± 0.09 mM Catechin Equivalence [CE]) showed a higher TPC compared with the control [vanillic acid (0.052 ± 0.003 mM CE, *P value < 0.05)]. However, the TPC of MF, MR and CL were found in the range of 0.020 ± 0.009 to 0.009 ± 0.01 mM CE. The ME extracts of MF and MR inhibited α-glucosidase significantly in comparison with acarbose as evidenced from the IC50 values (IC50 of MF = 1.58 ± 0.03 ng/µl; IC50 of MR = 1.87 ± 0.43 ng/µl and IC50 of acarbose = 3.34 ± 0.15 ng/µl). Moreover, DM extracts of MR (IC50 = 1.31 ± 0.29 ng/µl) also showed significantly higher α-glucosidase inhibitory activity. In contrary, MR (IC50 = 16.18 ± 0.16 ng/µl) and CL (IC50 = 9.21 ± 0.51 ng/µl) also showed significant α-amylase inhibitory activity in ME and DM extracts, respectively. These, results suggest that Fijian MPs could be a potential source of natural inhibitors of enzymes involved in carbohydrate digestion and thus may possibly be used in managing T2DM.

Free Radical Scavenging, Reducing Power and Lipid Peroxidation Inhibition Activities of M. communis Berries Methanol Extract and its Fractions

Free radicals or highly reactive oxygen species are capable of inducing oxidative damage to the human body Plants containing phenolic compounds have potent antioxidant capacity. The present study was undertaken to investigate the in vitro antioxidant activities of Myrtus communis L. (Myrtaceae), a plant widely used as natural remedy for digestive disorders in folk medicine. Total polyphenol contents were determined using Folin Ciocalteu's reagent; flavonoids were quantified employing the AlCl3 Method. EAE extract showed the highest polyphenolic and flavonoids contents (358.37±2.28 GAE/g of dry extract and 105.44±3.48 QE/g of dry extract) respectively. The EAE had the highest antioxidant activity as measured by DPPH radical and hydroxyl radical scavenging activity. EAE and ME extracts exhibited the highest reducing power. EAE possess an IC50 close to BHT (0.074 mg/ml) as reference drug All extracts exhibited antioxidant activity in the linoleic acid emulsion system (76.81% -86.93 %). EAE showed an inhibition ratio of (86.93 %) close to that of BHT (94.9±1.52 %). These findings provide evidence that Myrtus communis L. berries are a potential source of antioxidant which have many benefits towards human health.

Investigating the Antioxidant and Cytocompatibility of Mimusops elengi Linn Extract over Human Gingival Fibroblast Cells.

Background—chlorhexidine (CHX) is most commonly used as a chemical plaque control agent. Nevertheless, its adverse effects, including teeth discoloration, taste alteration and calculus build-up, limit its use and divert us to medicinal herbs. The purpose of the study was to evaluate the phytochemical composition, antioxidant potential, and cytotoxic effects of Mimusops elengi Linn extract (ME) over normal human cultured adult gingival fibroblasts (HGFs). Methods—in vitro phytochemical screening, total flavonoid content, antioxidant potential by DPPH and Nitric Oxide (NO) radical scavenging activity, and cytotoxic effects of ME extracts over HGF were explored. The viability of HGF cells was determined using 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT), neutral red uptake, and trypan blue assay after treatment with different concentrations of CHX and ME (0.3125 to 10 µg/mL). Results—ME showed some alkaloids, glycosides, saponins and flavonoids exhibited relatively moderate-to-good antioxidant potential. Increasing the concentration of CHX and ME from 0.3125 to 10 µg/mL reduced cell viability from 29.71% to 1.07% and 96.12% to 56.02%, respectively. At higher concentrations, CHX reduced the viability of cells by 52.36-fold compared to ME, revealed by MTT assay. At 10 µg/mL concentration, the mean cell viability of CHX and ME-treated cells was 2.24% and 57.45%, respectively, revealed by a neutral red assay. The viability of CHX- and ME-treated HGF cells estimated at higher concentrations (10 µg/mL) using trypan blue assay was found to be 2.18% and 47.36%, respectively. A paired t-test showed significance (p < 0.05), and one-way ANOVA difference between the mean cell viability of CHX- and ME-treated cells at different concentrations. One-way ANOVA confirmed the significant difference between the viability of CHX- and ME-treated cells. Conclusions—The cytoprotective and antioxidant effects of ME emphasize its potential benefits. Therefore, it could emerge as a herbal alternative and adjunct to conventional oral hygiene methods, that can diminish periodontal tissue destruction.

Antimicrobial Potential and Phytochemical Screening of Clathria sp. 1 and Tedania (Tedania) stylonychaeta Sponge Crude Extracts Obtained from the South East Coast of South Africa.

Scientists have continuously searched for novel bioactive compounds to overcome the inherent problems associated with drug resistance, the evolution of unknown diseases, and the toxicity of currently used compounds. The ocean has been considered a rich source of compounds that possess unique chemical structures and novel biological capabilities. Biologically active molecules isolated from marine flora and fauna have shown significant advancement over the past century in the pharmaceutical industry. Marine natural products (MNPs) have been used as nanomedicine, cosmetics, wound healing, antimicrobial agents, anticancer agents, and anti-inflammatory agents. The physicochemical parameters of the collection site were also recorded. This study's marine sponge species were collected from Phillip's Reef, South Africa, at 12 m during the spring season. Ethyl acetate (EA) and dichloromethane : methanol (DCM : ME, 1 : 1) were used as extraction solvents. Crude extracts of the marine sponges were tested against MRSA, P. aeruginosa, C. difficile, A. fumigatus, and C. albicans. Phytochemical screening was conducted to identify seven critical phytochemical groups. A pH reading of 8.01 and a temperature of 15.45°C were recorded at the sampling site. Clathria sp. 1 and Tedania (Tedania) stylonychaeta EA crude extracts showed bioactivity against all five test pathogens. The DCM : ME crude extract of Clathria sp. 1 was the only bioactive crude extract from DCM : ME extracts. This crude extract was only bioactive against C. albicans as no activity was observed against the other four pathogens. EA crude extracts of Clathria sp. 1 yielded more significant inhibition zones against both fungal pathogens. These EA crude extracts performed better than fluconazole as inhibition zones of 35 ± 0 mm at 24 mg/ml, 31 ± 0 mm at 19 mg/ml, 31 ± 0 mm at 14.4 mg/ml, 30 ± 0 mm at 9.6 mg/ml, and 25 ± 0 mm at 7.2 mg/ml were recorded. Clathria sp. 1 crude extracts exhibited higher inhibition zones compared to Tedania (Tedania) stylonychaeta. The antibiotic imipenem (26 ± 0.7 mm at 10 μg) and ciprofloxacin (30 ± 0.3 mm at 5 μg) exhibited higher zones of inhibition than EA crude extracts of Tedania (Tedania) stylonychaeta at all test concentrations. In this study, Clathria sp. 1 was observed to have broad-spectrum bioactivity as EA crude extracts were bioactive against MRSA, P. aeruginosa, C. difficile, A. fumigatus, and C. albicans. In addition to this, the EA crude extract of Clathria sp. 1 was bacteriostatic (9.6 mg/ml). Clathria sp. 1 DCM : ME crude extract only tested positive for the presence of terpenoids. In contrast, EA crude extracts did not test positive for the existence of any of the seven phytochemicals. Our study has revealed that Tedania (Tedania) stylonychaeta and Clathria sp. 1 sponge species collected from Phillip's Reef in South Africa can produce bioactive compounds useful against bacterial and fungal species.

Effect of plant extracts and citric acid on phytoremediation of metal-contaminated soil

Application of activating agents can significantly improve efficiency of plants for remediation of soils contaminated by heavy metals, however, damage to soil and plants limits application of traditional activating agents. The aim of our experiments is to select an efficient,green and low-cost activating agent to improve efficiency of plant extraction technology. In this study, contaminated soil was remediated by Sedum alfredii. The effects of two plant extracts (i.e., Oxalis corniculata,OX and Medicago sativaextract, ME) in addition to citric acid (CA) were studied in oscillatory activation experiment and pot experiment. The oscillation activation experiment revealed that extraction quantity of heavy metals in the soil was enhanced significantly with concentration of plant extract. The extraction quantity of Zn from 100% OX extract and ME extract were significantly higher than 10 mmol/L CA (54.04% and 33.09%, respectively). The 10 mmol/L CA has best extraction efficiency for Cd, up to 41.36 µg/kg, which is significantly higher than CK (control) (p < 0.05). The pot experiment exhibited that application of CA has significantly reduced soil pH and organic matter content by 8.63% and 28.21%, respectively, however the two extracts have no significant effect on soil properties. The study indicated that application of CA has negative effects on root morphological parameters and chlorophyll fluorescence parameters of Sedum alfredii.The addition of extracts of two plants have not caused any harm to Sedum alfredii. The application of three activating agents was beneficial for purification of Cd and Zn in soils, and its repairing efficiency was improved by 3.92, 3.37, 3.33 times and 0.44, 0.20, 0.86 times, respectively. The combination of plant extracts and hyperaccumulators can effectively remove heavy metals from contaminated soils, which provided a theoretical basis for mitigation of pollution in soils.

Zizyphus lotus (L.) extracts as prebiotics in the aggregation and adhesion of probiotic and inhibition of pathologic bacteria from patients with colorectal cancer

The mucosal surfaces of the intestinal tract harbor a complex microbiota. Bacteria enter in contact with intestinal cells and bestow important nutritional, metabolic and protective functions which benefit the host. Various factors are able to alter the balances between different intestinal bacteria. Dysbiosis has been described in various pathologies and metabolic diseases in humans, such as cancer. Colorectal cancer pathology can benefit from gut microbial imbalance. Its overgrowth may lead to acute symptoms. An alternative strategy to restore this balance is the use of plant extracts which exhibit a prebiotic activity by stimulating of probiotic bacteria and antimicrobial activity against pathogenic bacteria colonization. The present study aims to evaluate the in vitro antioxidant and antibacterial activity of phenolic extracts (Aqueous extract "AE" and Methanolic extract "ME" ) from Zizyphus lotus. It aims also to investigate the effect of this extract as a prebiotic on the aggregation of probiotic and inhibitory effects of pathogenic bacteria isolated from faeces of patients with colorectal cancer. Phytochemical analysis of different extracts from Zizyphus lotus by HPLC showed that all are rich in phenolic compounds (225.40 mgAGE/gDW for AE and 63.04 mgAGE/gDW for ME extracts). Moreover, all extracts showed an important antioxidant activity (227 µg/ml for AE and 195 µg/ml for ME). These extracts also exhibited a significant prebiotic effect and antibacterial activity. Characterization of the in vitro effect of the aqueous extract showed that the percentage of autoaggregation and adhesion of probiotic and inhibitory effects of pathogenic bacteria increased in the presence of aqueous extracts.

Moringa oleifera leaf extract ameliorated high-fat diet-induced obesity, oxidative stress and disrupted metabolic hormones

BackgroundObesity is a health problem in many countries, and maintaining a perfect weight is challenging. Moringa oleifera leaf extract (ME) is rich in polyphenols with antioxidant and pharmaceutical potential. The present study investigated the potential protective effect of Moringa oleifera leaf extract against obesity induced from a high-fat diet (HFD), oxidative stress and disruption of metabolic hormones compared to simvastatin (SIM) or their combination.ResultsRats fed a HFD for 6 weeks exhibited a significant increase in body weight and levels of serum glucose and lipid fractions, verifying an obesity state. There were also higher levels of insulin and leptin and lower gherlin in sera of HFD rats compared to the levels in control rats. Homeostasis model assessment for insulin resistance (HOMA-IR), quantitative insulin sensitivity check index (QUICKI) and the atherogenic index were elevated, indicating the development of insulin resistance and dyslipidaemia in these rats. These changes were accompanied with a significant increase in oxidative stress, as indicated by elevated lipid peroxidation and protein oxidation with low levels of antioxidants in liver. The activities of liver function enzymes, including aspartate amino transferase, alanine amino transferase, alkaline phosphatase and gamma glutamyltransferase, were also significantly increased in serum. Concurrent treatment with 300 mg/kg ME for 6 weeks ameliorated the increase in body weight and improved the levels of glucose, lipid fractions and metabolic hormones, indicating the anti-obesity effect and amelioration of tissue insulin resistance potential of ME. ME treatment also normalized oxidative stress and antioxidants in liver and improved liver function enzymes, indicating the antioxidant potential of ME. The effects of ME were similar to SIM, and the combination of these agents was better than each agent alone.ConclusionWe propose that ME extract has anti-obesity and antioxidant potential and may be used as a lipid-lowering drug to control weight, obesity and its pathophysiological consequences.

Wound healing of diabetic rats treated with Moringa oleifera extract.

To evaluate if Moringa oleifera leaf aqueous extract (ME) influences the healing of skin wounds of diabetic rats. Wistar rats were used (6 rats/group). Group 1 received normal saline (NS) v.o. Group 2 received moringa extract (100mg/kg v.o) for 3 weeks. Groups 3 and 4: Streptozotocin (STZ) induced diabetes. Group 3 received NS; Group 4 received aqueous ME (100mg/kg) v.o.The wounds of groups 1 and 3 rats were topically treated with NS; wounds of groups 2 and 4 treated with 200µL of 10% ME. After anesthesia, all rats had skin square excision wounds 1.5cm2. Wound percent contractions were measured. On 10th day, blood glucose and serum cytokines were measured. Histometry of wounds was studied using ImagePro6.0 software. Glycemia was significantly reduced in ME treated rats. These rats had higher percent contraction of the wounds on 2nd, 5th and 10th days, then controls (p<0.05). Diabetic rats treated with NS had TNF-α, IL-1β and IL-6 expression higher than in rats receiving ME. The histopathological score of ME treated diabetic rats (198±13.7) was significantly higher than treatment with NS (145±10.5). ME extract positively influenced healing of wounds in diabetic rats after systemic and topical treatment.

A potential use of vine-shoot wastes: The antioxidant, antifeedant and phytotoxic activities of their aqueous extracts

One of the biggest challenges for wine producing regions is to create alternatives to increase the added value of the large amount of vine-shoot wastes generated. The antioxidant, antifeedant and phytotoxic activities of vine-shoot aqueous extracts obtained by different extraction methods (Conventional Solid-Liquid Extraction, CSLE; Solid-Liquid Dynamic Extraction, SLDE-Naviglio; Microwave Extraction, ME) have been studied. Because of their higher content of phenolic compounds, CSLE and ME extracts showed a higher DPPH radical-scavenging activities compared with SLDE-Naviglio extracts. Significant antifeedant effects against Leptinotarsa decemlineata were observed for CSLE and ME. None of the extracts had phytotoxic effect against Lactuca sativa germination and radicle growth, but SLDE-Naviglio extracts stimulated the radicule elongation. ME extracts were the most active inhibitors of Lolium perenne germination. These results suggest some potential applications of vine-shoot extracts including cosmetics, nutraceuticals or pharmaceuticals linked to their antioxidant effects, or in organic agriculture because of their antifeedant and allelopathic activities.

Synergistic and suppressive effects of dietary phenolic acids and other phytochemicals from cereal extracts on nuclear factor kappa B activity

In this study the combinatorial action of the main phenolic acids (ferulic, caffeic, p-coumaric and sinapic acids) found in extracts of free (ME) and bound phenolic acids (HE) from oat, barley and wheat flour on the modulation of NF-κB activity was investigated. The results show that combination of phenolic acids in low concentrations (<0.1 μg/ml) has a significant synergistic, or enhanced effect, on NF-κB activity, while their combination in high concentrations (0.3–70 μg/ml) helps to suppress the strong effect obtained by individual solutions of ferulic and p-coumaric acids. The modulation of NF-κB activity observed by HE extracts is the effect of combinatorial action of the phenolic acids present therein, while the modulation of NF-κB activity observed with ME extracts is the result of combinatorial action of phenolic acids together with other phenolic compounds present in the extracts. These results increase the knowledge about the health promoting effect from cereal consumption and dietary phenolic acids.

Analgesic and antiinflammatory activity of Cyclamen repandum S. et S.

According to folk medicine some species belonging to the genus Cyclamen were used for their biological activities. Early investigation of the different species of the genus resulted in the isolation of triterpenic saponins. No phytochemical and biological data are available on C. repandum. As part of a series of phytochemical investigations for bioactive compounds from medicinal plants, Cyclamen repandum S. et S. was investigated. The present study sought to find the antiinflammatory and antinociceptive activities of C. repandum tubers in rats and mice. A preliminary screening was conducted with three different extracts in the tests used, particularly the paw edema and the writhing tests. Subsequently some saponins isolated from the ME extract, the more effective one, have been identified. This paper also describes the results of fractionation and bioassay guided chemical studies. Chemical investigation of the active extract afforded the isolation and characterization of six triterpenic saponins. The possible antiinflammatory and analgesic properties were investigated as the saponin content of the fractions allows to speculate on such aspect.

Cell wall protein and glycoprotein constituents of Aspergillus fumigatus that bind to polystyrene may be responsible for the cell surface hydrophobicity of the mycelium.

Cell surface hydrophobicity (CSH) of Aspergillus fumigatus grown both in complex medium (yeast extract/peptone/dextrose; YPD) and minimal (Vogel's N) medium was monitored by assessing attachment of polystyrene microspheres to the cell surface. It was found that mature mycelium was hydrophobic. Treatment of intact mycelium with beta-mercaptoethanol (beta ME) abolished binding of the microspheres to hyphal elements, and coating of the microspheres with beta ME extracts from mycelium inhibited their attachment to intact mycelial cells. A. fumigatus mycelium was tagged in vivo with biotin and treated with beta ME. The beta ME extracts were analysed by SDS-PAGE and Western blotting with both peroxidase-conjugated-ExtrAvidin and concanavalin A (ConA). This procedure allowed identification of cell wall surface proteins and glycoproteins. Rabbit polyclonal antisera were raised against beta ME extracts obtained from cells grown in YPD and Vogel's N media. These antisera defined some major cell-wall-bound antigens. SDS-PAGE and Western blotting analysis of the cell wall material released by beta ME and adsorbed on polystyrene microspheres revealed about 19 protein species with apparent molecular masses ranging from 20 to 70 kDa, and two high-molecular-mass glycoproteins of 115 and 210 kDa. Treatment of cells grown in YPD, but not those grown in Vogel's N medium, with beta ME released a 55 kDa polypeptide able to adsorb to polystyrene microspheres that was detectable with the antisera. The ability to bind to polystyrene particles exhibited by several protein and glycoprotein species released by beta ME treatment suggested that these cell wall moieties possess exposed hydrophobic domains that could be responsible for the CSH of mycelium.

Diagnosis of systemic candidiasis by enzyme immunoassay detection of specific antibodies to mycelial phase cell wall and cytoplasmic candidal antigens.

Diagnosis of systemic Candida infections was attempted by the use of an enzyme-linked immunosorbent assay (EIA) to detect IgG antibodies towards cell wall-bound and cytoplasmic candidal antigens. Cell wall antigens were sequentially solubilized by treatment of germinated blastoconidia of Candida albicans (ATCC 26555 strain) with beta-mercaptoethanol (beta ME extract) and digestion with Zymolyase 20T, a beta-glucanase preparation (Zymolyase extract). Protoplasts obtained after treatment with Zymolyase were osmotically lysed (cytoplasmic antigens). Sera were obtained from patients with systemic (n = 28) and superficial (n = 46) candidiasis. Control sera were obtained from normal healthy individuals (n = 31) and from hospitalized patients at low (n = 36) and at high (n = 13) risk of developing systemic candidiasis yet showing no symptoms of candidal infection. Detection of antibodies in crude sera samples by EIA using all of these antigenic extracts was highly specific (98-100%), but sensitivity of the method was low (3.5-17.8%). However, adsorption of sera with latex microspheres coated with purified Candida mannan in order to selectively remove anti-mannan antibodies prior to EIA improved the diagnostic efficiency of this test. Improvement was particularly noticeable when the beta ME extract was used as antigenic preparation, yielding a sensitivity of 89.2% and a specificity of 98.6%.

LH-releasing activity of hypothalamic extracts not attributable to GnRH.

LH secretion is under hypothalamic control, mostly through GnRH. Hypothalamic extracts stimulate LH release in vitro, an action usually attributed to the presence of GnRH. However, the presence of other LH-releasing factors cannot be ruled out since there are other neuromediators endowed with the capacity to influence LH release. We analyzed the presence of an LH-releasing activity in bovine median eminence extracts (ME extracts) different from GnRH using cultured rat pituitary cells. Both, GnRH and ME extracts caused a dose-dependent stimulation of LH release. The maximum response obtained with ME extracts (up to 10-fold the respective baseline) was significantly greater than the maximum response obtained with GnRH (up to 5-fold the respective baseline). Monoclonal or polyclonal GnRH antibodies in the solid phase completely eliminated the LH-releasing activity of 10(-7) M GnRH; in contrast, they only partially removed the activity present in ME extracts. The GnRH receptor blocker, Nal-Glu, 10(-7) M, which completely blocked the effect of GnRH could not totally suppress the LH-releasing activity of ME extracts. These findings indicate that ME extracts contain, in addition to GnRH, an LH-releasing activity not attributable to GnRH because it differs in its immunoreactivity and in its ability to stimulate LH release when GnRH receptors are blocked.

Identification of a 58-kilodalton cell surface fibrinogen-binding mannoprotein from Candida albicans

Treatment of both yeast (blastoconidia) and hyphal (blastoconidia with germ tubes) cells of Candida albicans with beta-mercaptoethanol (beta ME) releases a complex array of cell wall-bound proteins and glycoproteins. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western immunoblotting with fibrinogen-anti-fibrinogen antibody allowed the identification of a 58-kDa mannoprotein (mp58) in both extracts which specifically interacts with human fibrinogen. Treatment of intact cells with low concentrations of beta-glucanase (Zymolyase 20T) for short periods or with beta ME abolished or significantly reduced binding of fibrinogen. A rabbit polyclonal antiserum was raised against the purified mp58 species released by beta ME from germinated blastoconidia (PAb anti-mp58). By Western blotting, the antiserum cross-reacted with the homologous 58-kDa fibrinogen-binding mannoprotein present in beta ME extracts from blastoconidia, and by indirect immunofluorescence, the antiserum labelled both yeast cells and hyphae, yet reactivity was found primarily on the cell surface of filamentous forms. Immunostaining of human infected tissue sections with PAb anti-mp58 showed that the mp58 species is also expressed in vivo; in this case, the species is in the forms of both yeast and hyphal elements similarly labelled by the antiserum. Purified immunoglobulin G fraction from the antiserum did not alter the binding of fibrinogen as determined by a modified enzyme-linked immunosorbent assay and Western blotting. The N- and O-glycosidically linked carbohydrates represent 18 to 20% and 3 to 4%, respectively, of the molecular mass of the mp58. O-linked sugar residues may be involved in the interaction of the molecule with fibrinogen.

Cell wall glycoproteins of Candida albicans as released by different methods

Different methods of extraction frequently used in other studies were used to release glycoproteins from both intact cells and isolated cell walls of yeast and hyphal forms of Candida albicans. Extracts were obtained from whole cells by treatment (i) with 2-mercaptoethanol (beta ME) at pH 8.6 and 37 C degrees and (ii) with zymolyase after treatment with beta ME. Extracts were obtained from isolated and washed cell walls (i) by boiling with beta ME and sodium dodecyl sulphate (SDS), (ii) by boiling with SDS and (iii) by treatment with zymolyase after SDS. The extracts were separated by SDS-polyacrylamide gel electrophoresis and analysed by Western blotting with four reagents. Analysis with concanavalin A (ConA) revealed different glycoprotein populations depending on the treatment. Three possible germ-tube-specific constituents were observed; and 80 kDa component released by beta ME from both intact cells and cell walls, and 47 kDa and 43 kDa moieties released by zymolyase only from intact cells. MAb 4C12, specific for the protein portion of a large germ tube constituent, recognized polydisperse material which just entered the gel in beta ME extracts and in the region extending up from 200 kDa to near the top of the gel in zymolyase extracts. MAb 24.17, specific for a carbohydrate determinant of yeast phase cells, reacted with disperse material in the region from the top of the gel to one-third to two-thirds the distance to the 220 kDa mass marker. Antiserum specific for the serotype A determinant of mannan reacted with large disperse component(s) migrating in the region from the top of the gel to about two-thirds the distance to the 220 kDa mass marker and with a 180 kDa component. The components recognized by MAb 4C12, but not those recognised by MAb 24.17 and serotype A antiserum, were effected by treatment with endo-beta-N-acetylglucosamidase H. The various analyses revealed that the method of extraction affected the composition and size of the constituents recognized by the reagents.