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Drug discovery from complex mixture like Chinese herbs is a challenge and extensive false positives make the obtainment of specific bioactive compounds difficult. In the present study, a novel sample preparation method was proposed to rapidly reveal the specific bioactive compounds from complex mixtures using α-glucosidase as a case. Firstly, aqueous and methanol extracts of 500 traditional Chinese medicines were carried out with the aim of finding new sources of α-glucosidase inhibitors. As a result, the extracts of fruit of Terminalia chebula (FTC), flowers of Rosa rugosa (FRR) and Eugenia caryophyllata (FEC) as well as husk of Punica granatum (HPG) showed high inhibition on α-glucosidase. On-line liquid chromatography-diode array detection-tandem mass spectrometry and biochemical detection (HPLC-DAD-MS/MS-BCD) was performed to rapidly screen and characterize α-glucosidase inhibitors in these four extracts. After tentative identification, most of compounds with inhibitory activity in the investigated crude extracts were found to be tannins commonly recognized as non-specific enzyme inhibitors in vitro. Subsequently, the four extracts were treated with gelatin to improve specificity of the on-line system. Finally, two compounds with specific α-glucosidase inhibition were identified as corilagin and ellagic acid. The developed method could discover specific α-glucosidase inhibitors in complex mixtures such as plant extracts, which could also be used for discovery of specific inhibitors of other enzymes., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Alpha-glucosidase inhibitors play a potential role in the treatment of type 2 diabetes by delaying glucose absorption in the small intestine. Ecklonia maxima, a brown alga which grows abundantly on the west coast of South Africa, is used to produce alginate, animal feed, nutritional supplements and fertilizer. The crude aqueous methanol extract, four solvent fractions and three phlorotannins: 1,3,5-trihydroxybenezene (phloroglucinol) (1), dibenzo [1,4] dioxine-2,4,7,9-tetraol (2) and hexahydroxyphenoxydibenzo [1,4] dioxine (eckol) (3) isolated from E. maxima were evaluated for antiradical and alpha-glucosidase inhibitory activities. All the phlorotannins tested had strong antioxidant activities on DPPH free radicals with EC50 values ranging from 0.008 to 0.128μM. Compounds 2 and 3 demonstrated stronger antioxidant activity and an alpha-glucosidase inhibitory property than positive controls. These results suggest that E. maxima could be a natural source of potent antioxidants and alpha-glucosidase inhibitors. This study could facilitate effective utilization of E. maxima as an oral antidiabetic drug or functional food ingredient with a promising role in the formulation of medicines and nutrition supplements., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Objective: To investigate the presence of cysteine-rich secretory protein 1 (CRISP1) in seminal plasma as a means of distinguishing between obstructive azoospermia (OA) and nonobstructive azoospermia (NOA)., Design: Seminal plasma from normospermic donors (n = 45) and azoospermic donors (n = 80) was examined to determine CRISP1 levels. Neutral alpha-glucosidase (NAG) enzymatic activity was measured for comparison with CRISP1 levels., Setting: Research unit of an academic medical center., Patient(s): Normospermic and azoospermic donors from the clinical andrology laboratory of the centre hospitalier universitaire de Québec and from Mount Sinai Hospital., Intervention(s): Seminal CRISP1 measurement by Western blot analysis. Neutral alpha-glucosidase activity was evaluated by a photometric method., Main Outcome Measure(s): Seminal plasma CRISP1 levels, NAG activity, cutoff value, sensitivity, and specificity., Result(s): All seminal plasma samples from normospermic and nonobstructive azoospermic donors were CRISP1 positive, whereas CRISP1 was absent or present at low levels in samples from patients with OA. A significant correlation between seminal CRISP1 levels and NAG activity was found in azoospermic semen samples. The cutoff point to distinguish between donors with NOA or OA was established at 0.655 (relative intensity). At this threshold, specificity was 85% and sensitivity was 92%., Conclusion(s): Seminal CRISP1 combined with NAG activity can potentially distinguish between OA and NOA., (Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Inhibitors of the enzymes dipeptidyl peptidase (DPP)-IV and α-glucosidase are two classes of pharmacotherapeutic agents used for the treatment of type 2 diabetes. In the present study, whey protein isolate (WPI), α-lactalbumin, β-lactoglobulin, serum albumin, and lactoferrin hydrolysates obtained by peptic digestion were investigated for their potential to serve as natural sources of DPP-IV and α-glucosidase inhibitors. Although inhibition of DPP-IV activity was observed in all pepsin-treated whey proteins studied, the α-lactalbumin hydrolysate showed the greatest potency with an IC50 value of 0.036 mg/mL. Conversely, only WPI, β-lactoglobulin, and α-lactalbumin hydrolysates displayed some inhibitory activity against α-glucosidase. This study suggests that peptides generated from whey proteins may have dual beneficial effects on glycemia regulation and could be used as functional food ingredients for the management of type 2 diabetes.

Biodegradation of organic wastewater constituents by activated sludge microorganisms is based on enzymatic processes. It is supposed that wastewater treatment plants (WWTP) differ in their enzymatic fingerprints. To determine such fingerprints, activated sludges from nine aerated tanks of six WWTPs were repeatedly sampled and analyzed for the activities of l-alanine aminopeptidase, esterase, α- and β-glucosidase, alkaline phosphatase, phosphodiesterase, phosphotriesterase, and sulfatase. In one WWTP the enzymatic activities and their variations within 1 week were assayed in various process stages. Mostly the enzymatic profiles were dominated by l-alanine aminopeptidase, followed by alkaline phosphatase. They differed in variable contributions of esterase, phosphodiesterase, α- and β-glucosidase. The sulfatase activity was generally low. For the first time phosphotriesterase activity was detected in various samples, but with limited analytical validity. Particle mass-related activities of individual enzymes varied between plants by factors 2-4 and up to 11, when related to suspension volumes., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

The present study evaluates the influence of drying and cooking processes on the health properties of two bell Capsicum annuum L. cultivars Roggiano and Senise compared with fresh peppers. The content of phytochemicals decreased in the order fresh>dried>dried frying processes. HPLC analysis was applied to quantify five flavonoids from peppers. Apigenin was identified as main constituent. Its content was affected by drying and dried frying processes. The antioxidant activity was evaluated by DPPH, ABTS, β-carotene bleaching test and Fe-chelating activity assay. A comparable radical scavenging activity was observed for both cultivars. Interestingly, frying process did not influenced this property. Roggiano peppers exhibited the highest antioxidant activity using β-carotene bleaching test with IC(50) values of 38.1 and 24.9 μg/mL for total extract and n-hexane fraction, respectively. GC-MS analysis of lipophilic fraction revealed the presence of fatty acids and vitamin E as major components. In the inhibition of the carbohydrate-hydrolyzing enzymes fresh Senise peppers exerted the strongest activity against α-amylase with an IC(50) value of 55.3 μg/mL. Our results indicate that C. annuum cultivars Roggiano and Senise have an interestingly potential health benefits not influenced by processes that are used before consumption., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

We determined the total and dissolved extracellular enzymatic activity (EEA) of α-glucosidase and β-glucosidase (AGase and BGase), alkaline phosphatase (APase) and leucine aminopeptidase (LAPase) activities in the epi-, meso- and bathypelagic waters of the subtropical Northeast Atlantic. EEA was also determined in treatments in which bacterial EEA was inhibited by erythromycin. Additionally, EEA decay experiments were performed with surface and deep waters to determine EEA lifetimes in both water masses. The proportion of dissolved to total EEA (66-89 %, 44-88 %, 57-82 % and 86-100 % for AGase, BGase, APase and LAPase, respectively) was generally higher than the cell-associated (i.e., particulate) EEA. The percentage of dissolved to total EEA was inversely proportional to the percentage of erythromycin-inhibited to total EEA. Since erythromycin-inhibited plus dissolved EEA equaled total EEA, this tentatively suggests that cell-associated EEA in the open oceanic water column is almost exclusively of bacterial origin. The decay constants of dissolved EEA were in the range of 0.002-0.048 h(-1) depending on the type of extracellular enzyme, temperature and depth in the water column. Although dissolved EEA can have different origins, the major contribution of Bacteria to cell-associated EEA and the long life-time of dissolved EEA suggest that Bacteria-and not mesophilic Archaea-are essentially the main producers of EEA in the open subtropical Northeast Atlantic down to bathypelagic layers.

Five phenolic compounds were isolated from the seeds of Perilla (Perilla frutescens L.) using gradient solvent fractionation, silica gel column chromatography, and preparative high-performance liquid chromatography (HPLC). Their chemical structures were identified as caffeic acid-3-O-glucoside (1), rosmarinic acid-3-O-glucoside (2), rosmarinic acid (3), luteolin (4), and apigenin (5) using NMR spectroscopy and HPLC-ESI/MS analysis. Among them, luteolin (4) inhibited α-glucosidase (EC 3.2.1.20) with IC(50) value of 45.4μM. The inhibition kinetic analysed by Dixon plot indicate that luteolin is a noncompetitive inhibitor, and the inhibition constant K(I) was calculated at 45.0μM. Moreover, rosmarinic acid (3) and luteolin (4) inhibited recombinant human aldose reductase (EC 1.1.1.21) with IC(50) values of 11.2 and 0.6μM, respectively. Notably, the inhibition kinetic of luteolin (4) follows a hyperbolic dependence on aldose reductase inhibition by Dixon plot. Thus, inhibition kinetic indicates that luteolin (4) is a mixed-type inhibitor., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Blueberries have been extensively researched, but there are limited studies on other parts of the plant. Here we report the first phytochemical examination of highbush blueberry (Vaccinium corymbosum) flowers, which yielded 21 phenolics. The compounds were identified from extensive NMR and mass spectral analyses and included five caffeic acid (1-5), three coumaric acid (6-8), and two cinnamyl alcohol (9-10) derivatives, eight flavonol glycosides (11-18), and three phenylpropanoid-substituted catechins (19-21). The isolates were evaluated for antioxidant and α-glucosidase inhibitory activities. Overall, the flavonol glycosides and phenylpropanoid-substituted catechins showed superior antioxidant activities compared to the positive controls, vitamin C (IC(50)=63μM) and butylated hydroxytoluene (IC(50)=1548μM). Similarly, these phenolic sub-classes were more potent α-glucosidase inhibitors than the clinical drug, acarbose (IC(50)=200μM). Thus, non-consumed parts of food plants may be exploited as sources of bioactive compounds beyond their edible parts alone for nutraceutical and functional food applications., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

The objectives of this study were to identify novel peptides from albumin, and to evaluate and validate the anti-diabetic activity of peptides against α-glucosidase and α-amylase. In the research, albumin hydrolysate was purified and identified, tandem MS was adapted to characterise the amino acid sequences of peptides from the hydrolysate. In addition, anti-diabetic effects of the peptides with α-glucosidase and α-amylase inhibitory activity have been performed. The present work found eight novel peptides from albumin. Results also suggested that peptide KLPGF had α-glucosidase inhibitory activity with an IC(50) of 59.5±5.7μmoll(-1) and α-amylase inhibitory activity with an IC(50) of 120.0±4.0μmoll(-1). In conclusion, the results revealed that the peptide KLPGF was a potential anti-diabetic inhibitor., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Electrospray ionisation-mass spectrometry (ESI-MS) was applied to analyse the water-soluble extract of red ginseng (RG). Several new compounds were produced from the Maillard reaction during the steaming and drying process for preparing RG. Both the tandem electrospray ionisation (ESI-MS(n)) and Fourier transform ion cyclotron resonant mass spectrometric (FT-ICR-MS) data of these products proved that they were the initial Maillard reaction products (MRPs) of maltose with glutamic acid/aspartic acid, which were specific components in RG. In addition, their anti-diabetic and antioxidant activities were examined in vitro. The anti-diabetic activities were evaluated by studying the α-glucosidase inhibition using ultrafiltration LC-MS/MS techniques, while the antioxidant activities were investigated by UPLC-ESI-MS method. The results demonstrated that four initial MRPs in RG were identified as α-glucosidase inhibitors, and showed marked scavenging effect on the hydroxyl radical ((·)OH). Based on these studies, the processing method of RG was improved to generate more active compounds., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Pompe disease is a lysosomal storage disorder caused by a deficiency of acid alpha glucosidase (GAA). Diagnosis of Pompe disease is typically based on an enzyme analysis of blood or tissues, such as fibroblasts, followed by confirmation through molecular testing. The advent of fluorometric and mass spectrometry methods for enzyme analysis in dried blood spots (DBS) has simplified the diagnostic approach for Pompe disease, facilitating high-throughput screening of at-risk populations and newborn infants. The following unit will provide the detailed analytical protocol for measurement of GAA activity in DBS using tandem mass spectrometry.

Pompe disease can be treated effectively, if immune tolerance to enzyme replacement therapy (ERT) with acid α-glucosidase (GAA) is present. An adeno-associated viral (AAV) vector carrying a liver-specific regulatory cassette to drive GAA expression (AAV-LSPhGAA) established immune tolerance in GAA knockout (KO) mice, whereas ubiquitous expression with AAV-CBhGAA provoked immune responses. Therefore, we investigated the hypothesis that immune tolerance induced by hepatic-restricted expression was dominant. AAV-LSPhGAA and AAV-CBhGAA were administered singly or in combination to groups of adult GAA-KO mice, and AAV-LSPhGAA induced immune tolerance even in combination with AAV-CBhGAA. The dual vector approach to GAA expression improved biochemical correction of GAA deficiency and glycogen accumulations at 18 weeks, and improved motor function testing including wire-hang and grip-strength testing. The greatest efficacy was demonstrated by dual vector administration, when both vectors were pseudotyped as AAV8. T cells from mice injected with AAV-LSPhGAA failed to proliferate at all after an immune challenge with GAA and adjuvant, whereas mock-treated GAA-KO mice mounted vigorous T cell proliferation. Unlike AAV-LSPhGAA, AAV-CBhGAA induced selective cytokine and chemokine expression in liver and spleen after the immune challenge. AAV-CBhGAA transduced dendritic cells and expressed high-level GAA, whereas AAV-LSPhGAA failed to express GAA in dendritic cells. The level of transduction in liver was much higher after dual AAV8 vector administration at 18 weeks, in comparison with either vector alone. Dual vector administration failed to provoke antibody formation in response to GAA expression with AAV-CBhGAA; however, hepatic-restricted expression from dual vector expression did not prevent antibody formation after a strong immune challenge with GAA and adjuvant. The relevance of immune tolerance to gene therapy in Pompe disease indicates that hepatic expression might best be combined with nonhepatic expression, achieving the benefits of ubiquitous expression in addition to evading deleterious immune responses.

Background and Aims: The "gold standard" for the diagnosis of celiac disease (CD) is the small intestinal biopsy. A significant number of biopsies are inadequate for interpretation. Furthermore, the labeling of a biopsy as a Marsh I or II is somewhat subjective and may vary with the experience of the pathologist. Our hypothesis is that patients with intact villi undergoing biopsies frequently have associated disaccharidase deficiencies (DSD)., Methods: We reviewed 220 charts of pediatric patients with CD and selected those with a duodenal biopsy Marsh score of I/II. The disaccharidase (DS) levels of these patients were compared with a randomly selected, age-matched control group. DSD is defined as levels below the lower limits of normal., Results: Lactase (mean lactase = 18.8 in the control group vs. 4.2 in the diseased group, p = 0.004); sucrase (mean sucrase = 46.4 in the control group vs. 21.4 in the diseased group, p = 0.001); maltase (mean maltase = 138 in the control group vs. 52.5 in the diseased group, p = 0.001); palatinase (mean palatinase = 9.6 in the control group vs. 3.3 in the diseased group, p < 0.001)., Conclusion: There is a profound deficiency of DS levels in pediatric patients with CD who have intact villi.

The aim of this study was to assess the use of functional microbial ecological techniques in detailing processes during composting. Using starch as an amendment for a co-composting process, small-scale individual mesocosms were investigated at two temperatures, 30 degrees C and 60 degrees C, over a five-week period. In order to determine the more exact processes occurring during degradation, extracellular enzyme analysis (EEA) and community-level physiological profiling (CLPP) were used. The EEA demonstrated that, although assay temperature effects were present, no significant difference between the two in situ composting temperatures was observed for the two enzyme substrates, alpha- and beta-glucosidase, used. The CLPP profiling was only performed on the 30 degrees C compost samples. These analyses indicated a dynamic, but broadly predictable, environment, suggesting that the use of this approach may be further applicable to these types of study. By investigating both total dry and organic weights, no difference in organic matter degradation between the two temperatures was observed. These results showed that either starch degradation acted independently of temperature or compensatory effects occurred. The former hypothesis was supported by a second experiment investigating starch degradation at both temperatures with appropriate negative controls. Trying to reconcile the EEA results with the changes in total matter was difficult without making speculative assumptions. The study highlights the need to further develop and evaluate these microbial ecological techniques to address such problems.

Active compounds with antidiabetic potential were isolated from silk peptide E5K6 by consecutive ultrafiltration and gel filtration using Biogel P-2 and RS-HPLC using a YMC-Pack Pro C18 column. The highest α-glucosidase inhibitory activity of silk peptide E5K6 resulted from fractions with MW <1 kDa. The activities of gel-filtered fractions from silk peptide E5K6 of <1 kDa were assayed in vitro, demonstrating that the fourth peak (F4) had the highest α-glucosidase inhibitory activity (IC(50) = 37.1 mg/mL). F4 of silk peptide E5K6 was separated by HPLC into two peaks. Moreover, the purified compounds were identified as Gly-Glu-Tyr (GEY, MW = 367 Da) and Gly-Tyr-Gly (GYG, MW = 295 Da) according to amino acid sequences, and their α-glucosidase inhibitory activities (IC(50)) were 2.7 and 1.5 mg/mL, respectively.

Saprotrophic cord-forming basidiomycetes are important decomposers of lignocellulosic substrates in soil. The production of extracellular hydrolytic enzymes was studied during the growth of two saprotrophic basidiomycetes, Hypholoma fasciculare and Phanerochaete velutina, across the surface of nonsterile soil microcosms, along with the effects of these basidiomycetes on fungi and bacteria within the soil. Higher activities of α-glucosidase, β-glucosidase, cellobiohydrolase, β-xylosidase, phosphomonoesterase and phosphodiesterase, but not of arylsulphatase, were recorded beneath the mycelia. Despite the fact that H. fasciculare, with exploitative hyphal growth, produced much denser hyphal cover on the soil surface than P. velutina, with explorative growth, both fungi produced similar amounts of extracellular enzymes. In the areas where the mycelia of H. fasciculare and P. velutina interacted, the activities of N-acetylglucosaminidase, α-glucosidase and phosphomonoesterase, the enzymes potentially involved in hyphal cell wall damage, and the utilization of compounds released from damaged hyphae of interacting fungi, were particularly increased. No significant differences in fungal biomass were observed between basidiomycete-colonized and noncolonized soil, but bacterial biomass was reduced in soil with H. fasciculare. The increases in the activities of β-xylosidase, β-glucosidase, phosphomonoesterase and cellobiohydrolase with increasing fungal:bacterial biomass ratio indicate the positive effects of fungal enzymes on nutrient release and bacterial abundance, which is reflected in the positive correlation of bacterial and fungal biomass content., (© 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.)

The aim of this study was to evaluate the gastroprotective efficacy of andrographolide isolated from Andrographis paniculata in rats induced with duodenal ulcers. Duodenal ulcers were induced by cysteamine administration in rats pretreated with 3 mg kg⁻¹ BW day⁻¹ of andrographolide for 30 days. Ulcer score, myeloperoxidase activity, TBARS level, GSH/GSSG ratio and enzyme antioxidants were measured in the duodenal tissue. Brush border and basolateral membranes were isolated to assay sucrase, maltase, alkaline phosphatase and total ATPases. Ulcer score was significantly minimised in rats pretreated with andrographolide. Elevation in myeloperoxidase and TBARS levels were found to be minimised significantly due to andrographolide treatment. Membrane-bound enzyme activities and the thiol redox status of glutathione were significantly maintained in duodenal mucosa of rats that received andrographolide. This study reveals that the major component of A. paniculata, andrographolide, has potent antiulcer properties that are most likely caused by minimising inflammatory changes, counteracting free radical formation and maintaining the thiol redox status in the duodenum.

Since the past decade consumption of certain foods has been reported to have a positive effect on health. The object of the study was to determine for the first time the chemical composition and the antioxidant, anti-inflammatory and hypoglycemic potential of Citrus medica L. cv Diamante flowers, leaves and fruits (endocarp and mesocarp) at two maturity stages. Flowers and leaves were characterized by the highest total phenols and flavonoids content. A declining trend was observed during maturity of fruits for both phenols and flavonoids. The antioxidant activity evaluated by the β-carotene bleaching test showed a strong activity for flowers and endocarp of mature fruits with IC50 values of 2.8 μg/mL and 3.5 μg/mL, respectively, after 30 min of incubation. Interestingly, the mature fruits endocarp (IC50 value of 426.0 μg/mL) could inhibit α-amylase with an IC50 value 2-fold higher than immature fruits. None of the tested extracts affected the proliferation of human skin fibroblasts 142BR. The obtained results suggest a potential use of C. medica L. cv Diamante as new valuable Citrus species with functional properties for food or nutraceutical product on the basis of high content of phytochemicals., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Spices are extensively used to enhance the taste and flavor of foods and are known to possess several medicinal properties. Myristica fragrans, Parmelia perlata, Illicium verum, Trachyspermum copticum and Myristica malabarica, the commonly used spices in India were assessed for antidiabetic activity in streptozotocin induced diabetic rats. In the in vitro insulin secretion studies on isolated islets of Langerhans, M. fragrans, T. copticum and M. malabarica showed dose dependent insulin secretion. At 1 mg/ml, P. perlata showed significant in vitro alpha-glucosidase inhibitory activity with IC(50) value of 0.14 mg/ml followed by M. malabarica (0.64 mg/ml), I. verum (0.67 mg/ml), M. fragrans (0.85 mg/ml) and T. copticum (0.92 mg/ml). The DPPH free radical scavenging activity of the extracts at a concentration of 1 mg/ml was as M. malabarica (90.45%), M. fragrans (89.89%), I. verum (87.22%), P. perlata (76.70%) and T. copticum (38.14%). P. perlata showed the highest phenolic content (i.e., 118.5 mg gallic acid equivalents/g) followed by M. malabarica (84.13 mg gallic acid equivalents/g). M. malabarica showed the highest flavonoid content (i.e., 38.35 mg quercetin equivalents/g). Regular use of these spices may prevent postprandial rise in glucose levels through inhibition of intestinal alpha-glucosidase and may maintain blood glucose level through insulin secretagogue action.

Pompe disease is caused by the deficiency of acid α-glucosidase (GAA), which degrades glycogen into glucose. Its manifestation is characterized by a broad and continuous spectrum of clinical severity ranging from severe infantile to relatively benign adult form. We describe a 12-year-old girl diagnosed at a presymptomatic stage of late form Pompe disease due to fortuitous detection of an elevated level of serum creatine kinase (CK) at the age of 4. Biopsies were taken from the quadriceps muscle and studied with histological and histochemical techniques, as well as in electron microscope. Sporadic muscle cells showed the accumulation of lysosomal glycogen, suggesting Pompe disease. Interestingly, we found lysosomal bound glycogen, located in the axons of intramuscular nerves. The diagnosis was confirmed by deficient GAA activity in leukocytes. Mutation analysis revealed changes IVS1-13T>G and p.C103G in the GAA gene. The patient was able to obtain enzyme replacement therapy in the early asymptomatic stage of the disease., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Contraceptive techniques which target vas deferens have been paid great attention for their good efficacy, safety and reversibility. We have made a filtering-type intra-vas device (IVD) using nano-copper complex/polymer composites. Twenty male adult Beagle dogs and 40 male rabbits were randomly assigned to four groups (sham-operation, IVD, reversal and vasectomy groups). Dogs' semen parameters, concentration of α-glucosidase, copper and zinc ions were tested pre-operation and 1, 3, 6 and 12 months post-operation. The pregnancy rates of the rabbits were evaluated by mating trials after the IVDs were implanted. The histology of testis, epididymides and vas deferens of the animals was examined using an electron microscope. Apoptosis of the cells in the testes, epididymides and vas deferens was detected by TUNEL method. There was no sperm in the semen of dogs, which had been inserted IVD and vasectomized at 1, 3, 6 and 12 months post-operation. The concentration of α-glucosidase in the IVD group, reversal group and sham-operation group was not significantly different between pre- and post-operation. The pregnancy rates of the female rabbits in the vasectomy, IVD and reversal groups were all zero, but the pregnancy rate in the reversal group, after taking out IVD, and that of the sham-operation group was 60% and 80%, respectively. The ultrastructures of the testes, epididymides and vas deferens of the male animals in the IVD group and sham-operation group were in normal ranges compared with the vasectomy group. The apoptosis of the cells in the testes, epididymides and vas deferens in the vasectomy group of both dogs and rabbits was obvious compared with the other groups. No significant changes in the quantities of copper and zinc ions were found in semen of the male dogs both pre- and post-operation. Our studies demonstrated that the filtering-type nano-copper complex/polymer composites intra-vas device may be an efficacious, safe and reversible male contraceptive device., (© 2010 The Authors. International Journal of Andrology © 2010 European Academy of Andrology.)

Objective: To study the active compounds from the rhizomes of Musa basjoo., Method: Antioxidant and alpha-glucosidase inhibitory activity of different extracts were tested. Using bioassay-guided fractionation, the chemical constituents in EtOAC extracts were isolated by column chromatography and identified by MS and NMR spectroscopy., Result: Five compounds were isolated and identified as 2',3, 4'-trihydroxyflavone (1), 3,3'-bis-hydroxyanigorufone (2), irenolone (3), 4-dihydroxy-9-(4'-hydroxyphenyl)-phenalenone (4) and 3,4-dihydroxybenzaldehyde (5). Compound 1(IC50 8.61 mg x L(-1)), 3 (IC50 19.55 mg x L(-1)) and 5 (IC50 1.1 mg x L(-1)) had antioxidant activity. Compound 2 (IC50 24.15 mg x L(-1)) and 4(IC50 2.81 mg x L(-1)) had alpha-glucosidase inhibitory activity. Compound 5 showed MIC of 0.078, 0.313, 0.039 microg/disc against SA, MRSA and ESBLs, respectively., Conclusion: Compound 1-5 were isolated from this plant for the first time. Compound 5 was isolated from the genus Musa for the first time. All compound except 5 were first reported about activity.

Background: The definitive biochemical test for the diagnosis of disaccharidase deficiencies is the determination of enzyme activities in small bowel mucosa. Traditional methods of tissue homogenization are limited by low throughput. Lack of tissue-based quality control materials is a limitation for clinical laboratories that perform tests of the intestinal disaccharidases. The objectives of this study were to develop a method for determining disaccharidase activities using a high-throughput homogenization method and to create matrix-appropriate quality control materials., Methods: Intestinal mucosa was homogenized using three different methods. Activities of lactase, maltase, palatinase, and sucrase were determined by measuring glucose liberated from a hydrolysis reaction with disaccharide substrates and normalized to total protein concentration. Tissue-based quality control materials were developed from pig intestine and pooled clinical specimens., Results: A high-throughput homogenization method processed 24 specimens in 2 min compared to 1 specimen per 2 min for traditional methods. There were no significant differences (p>0.90) in enzyme activities as a function of the homogenization method used. 92 determinations of enzyme activities from 23 clinical specimens showed acceptable agreement (y=1.13x-6.6) and were highly correlated (r=0.95) as compared to results obtained by an external laboratory. Enzyme activities were variably inactivated by temperatures > or = 60 degrees C. Pig intestinal mucosa produced non-deficient enzyme activities while deficient activities were produced by combining equal volumes of unheated and heat-inactivated homogenates pooled from clinical specimens., Conclusions: A high-throughput method of tissue homogenization was more efficient than traditional methods and did not affect enzyme activities. Matrix-appropriate quality control materials can be created from pig intestine and pooled human homogenates., (Copyright 2010 Elsevier B.V. All rights reserved.)

Pompe disease is an autosomal recessive lysosomal storage disorder (LSD) caused by deficiency of lysosomal acid alpha-glucosidase (GAA) activity. This is the first LSD in which newborn screening has been shown to improve clinical outcomes. Newborn screening also identified multiple rare gene variants in this population. Among 132,538 newborns screened, 107 babies (1 in 1239) who had low dried blood spot GAA activity were genotyped. Sixty-nine (64.5%) babies had a total of 54 mutations and 35 novel predictably pathogenic mutations; 36 babies (33.6%) who had no mutation were homozygous for the c.[1726A; 2065A] pseudodeficiency allele. Because 81% of the chromosomes (14% in the controls) were in haplotype *03, we found a link between the pseudodeficiency allele and other mutated alleles. The newborns with Pompe disease detected by screening had lymphocyte GAA activities 0.45 to 1.65 nmol/mg/h (normal 66.7+/-33.8), while only 2 of the 100 false-positive cases had GAA activity less than 2.00 nmol/mg/h (or 3% of the normal mean). Therefore, newborn screening for Pompe disease could be successfully conducted by including genotyping and lymphocyte GAA assay, even in a population with mutation heterozygosity and pseudodeficiency., (Copyright 2009 Elsevier Inc. All rights reserved.)

An HPLC-UV method has been developed for the determination of valibose, miglitol, voglibose and acarbose, the four anti-diabetic drugs. The separation was accomplished successfully by using reversed phase chromatography (Prevail carbohydrate column, 250 mm x 4.6 mm, 5 microm) with a gradient acetonitrile-phosphate buffer solution (pH 8.0) at a wavelength of 210 nm. Furthermore, the method of a high-performance liquid chromatography coupled with ESI-MS in positive ionization mode has been established. These two methods were successfully applied to the assay and qualitative detection of four alpha-glucosidase inhibitors in the potential counterfeit anti-diabetic drugs.

1. Effects of early feeding with a diet containing added putrescine on duck intestinal development and growth performance were examined by a 2 x 2 factorial arrangement with two different feeding times (6 and 48 h) and two levels of putrescine (0 and 025%). 2. A significant main effect of early feeding on increasing body weight (BW) was observed from hatch to 35 d, whereas dietary putrescine had no significant effect on BW. 3. In the first week posthatch, enhanced small intestinal weight and intestinal density (weight of intestinal tissue/unit length of intestine), increased villus length and reduced crypt depth were observed in the early feeding group, while no effect was observed when putrescine was added to the diet. 4. Maltase and sucrase activity and protein/DNA ratio in jejunum were increased by early feeding in the first week, while decreased by putrescine supplementation. 5. In conclusion, early feeding methods have great potential for small intestine development and thereafter enhanced the growth performance of ducks, but dietary putrescine used during this period should be used cautiously to avoid toxicity.

Colored grains including red, purple, and black rice, purple corn, black barley, and black soybean contain anthocyanins. The present study was designed to (i) identify and quantify the individual anthocyanins and measure the total phenolic content (TPC), (ii) evaluate the antioxidant and alpha-glucosidase inhibitory activity, and (iii) correlate the TPC with total antioxidant activity and alpha-glucosidase inhibitory potency in these colored grains. The TPC was measured using a Folin-Ciocalteu assay, while the total antioxidant activity was determined by a method based on the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity. Among all of the studied colored grains, black rice possessed the highest TPC, which was 86 times greater than that of red rice. In addition, black rice had the highest total anthocyanin contents and alpha-glucosidase inhibitory activity. A significant positive correlation of the antioxidant activity and alpha-glucosidase inhibitory activity with total anthocyanin content and TPC was observed in this study. It is concluded that black rice possesses the highest antioxidant activity and alpha-glucosidase inhibitory among all of the colored grains tested and can be further explored as a functional food.

The organic extract of the roots of Broussonetia papyrifera showed extremely high alpha-glucosidase inhibitory activity with an IC50 of around 10 microg/mL. Due to its potency, subsequent bioactivity-guided fractionation of the chloroform extract led to 12 polyphenols, 1-12, 4 of which were identified as chalcones (1-4), another 4 as flavans (5-8), 2 as flavonols (9 and 10), and 2 others as the novel species benzofluorenones (11 and 12). Broussofluorenone A (11) and broussofluorenone B (12) emerged as new compounds possessing the very rare 5,11-dioxabenzo[b]fluoren-10-one skeleton. These compounds (1-12) were evaluated for alpha-glucosidase inhibitory activity to identify their inhibitory potencies and kinetic behavior. The most potent inhibitor, 10 (IC50=2.1 microM, Ki=2.3 microM), has an inhibitory activity slightly higher than that of the potent alpha-glucosidase inhibitor deoxynojirimycin (IC50=3.5 microM). The novel alpha-glucosidase inhibitors 11 (IC50=27.6 microM) and 12 (IC50=33.3 microM) are similar in activity to sugar-derived alpha-glucosidase inhibitors such as voglibose (IC50=23.4 microM). Interestingly, major constituents (1, 2, 6, 7, 9, and 10) of B. papyrifera displayed significant inhibitory activity with IC50 values of 5.3, 11.1, 12.0, 26.3, 3.6, and 2.1 microM, respectively. In kinetic studies, chalcones (1-4) exhibited noncompetitive inhibition characteristics, whereas the others (5-12) showed mixed behavior.

This study compared the enzymatic activity of clinical isolates of Cryptococcus neoformans, Cryptococcus gattii, environmental isolates of C. neoformans and non-neoformans Cryptococcus. Most of the cryptococcal isolates investigated in this study exhibited proteinase and phospholipase activities. Laccase activity was detected from all the C. neoformans and C. gattii isolates, but not from the non-neoformans Cryptococcus isolates. There was no significant difference in the proteinase, phospholipase and laccase activities of C. neoformans and C. gattii. However, significant difference in the enzymatic activities of beta-glucuronidase, alpha-glucosidase, beta-glucosidase and N-acetyl-beta-glucosaminidase between C. neoformans and C. gattii isolates was observed in this study. Environmental isolates of C. neoformans exhibited similar enzymatic profiles as the clinical isolates of C. neoformans, except for lower proteinase and laccase activities.

This study was designed to evaluate the effect of mycoplasma contamination on acid hydrolase activity and the action of the mycoplasma removal agent (MRA), in cultures of human fibroblasts from individuals with lysosomal diseases. For this purpose, we measured the activity of the b-galactosidase, arylsulphatase B (ASB), hexosaminidase A and a-glucosidase enzymes. The activity of the above mentioned enzymes in fibroblasts contaminated by mycoplasma was measured before and after the addition of the MRA. The results were then compared to the enzymatic activity in contamination-free cultures. Only the ASB enzyme showed significant alteration in activity both in the presence of mycoplasma and MRA. The remaining enzymes did not suffer significant interference by the presence of the two agents. Of the four enzymes tested, three did not suffer significant alterations by the presence of the mycoplasma nor from the MRA. However, the activity measured in the ASB enzyme increased significantly in the presence of mycoplasma and MRA and could lead to a doubtful diagnosis. Therefore, we suggest that contamination should be prevented by using aseptic techniques as well as the MRA in those fibroblast cultures that cannot be discarded.

Alpha-glucosidase from Aspergillus niger is an enzyme that catalyzes hydrolysis of alpha-1,4 linkages and transglucosylation to form alpha-1,6 linkages. In this study, an analytical method of oligosaccharides by nuclear magnetic resonance (NMR) was used to provide quantitative estimation of the fractions of each sugar unit and was applied to characterize the alpha-glucosidase reaction. Our data indicated that alpha-glucosidase reacts with the nonreducing end of oligosaccharides to form an alpha-1,6 linkage, and then a sugar unit with two alpha-1,6 linkages is gradually produced. Data from mass spectrometry suggested that the sugar unit with two alpha-1,6 linkages originates mainly from a 3mer and/or 4mer when oligosaccharides are used as substrates.

An experiment was conducted to investigate the effects of diets containing soybean meal (SBM), soybean protein concentrate (SPC), and soybean protein isolate (SPI) on growth performance and gut function of the young turkey. A total of 812 one-day-old male turkey poults were randomly assigned to 4 dietary treatments, with 7 pens per treatment and 29 birds per pen. The 4 experimental diets contained SBM, SBM-SPC, SPC, and SPI and were fed throughout the two 4-wk experimental periods. In each period, the diets were isonitrogenous and isocaloric and contained similar amounts of total and water-soluble nonstarch polysaccharides. The content of oligosaccharides differed among the diets and averaged 2.4, 1.9, 0.9, and 0.1% for SBM, SBM-SPC, SPC, and SPI, respectively. When compared with SBM, birds consuming the SBM-SPC and SPC diets had higher (P<0.05) final BW (4.32 vs. 4.45 and 4.46 kg, respectively). Incorporation of SPI as a substitute for SBM resulted in improved (P<0.05) feed utilization (from 1.76 to 1.67) but did not affect the final BW. Significant changes in cecal concentrations of short-chain fatty acids were observed and averaged 130, 103, and 89 micromol/g of digesta for the SBM, SBM-SPC, and SPC diets, respectively. This coincided with the proportional decrease in dietary oligosaccharide content (from 2.4 to 0.9%) and was further substantiated by a significant decrease in ileum weights. Feeding the SPI diet resulted in the lowest ileal and cecal tissue weights as well as the lowest cecal short-chain fatty acids concentration. There was no effect of diet on digesta pH, viscosity, and mucosal sucrase and maltase activities. Bacterial beta-glucuronidase activity was decreased (P=0.08) in the cecum (from 0.98 to 0.60 U/g) with decreased dietary oligosaccharide content. In conclusion, partial or almost complete substitution of SBM with SPC suppressed the fermentation processes in the ceca but enhanced the growth rate. Substitution of SBM with SPI significantly improved feed utilization but decreased BW of 4-wk-old turkeys with no effect on growth rate of older 8-wk-old birds.

Objective: To study the chemical constituents and bioactivity of Teurium pilosum., Method: Various column chromatographic techniques were used to isolate the constituents. A combination of EI-MS, NMR spectroscopy and X-Ray were applied to identify the structures. The anti-microorganism was accomplished by disk diffusion method, the antioxidant activity was assayed by the DPPH microanalysis models and the inhibitory activity of alpha-glucosidase was screened In vitro., Result: Eight compounds were isolated and identified as: glyceryl tristearate (1), 2,5-dioxolanone (2), fernenol (3), stigmasta-5,22-dien-3P-ol (4), 24-nor cholesta-5,22 (E)-dien-3beta-ol (5), ca-spinasterol (6), (3,4-dihydroxyphenyl) acrylate (7), 3,4-dihydroxy phenyl acrylic acid (8)., Conclusion: All compounds have been isolated from the genus for the first time. Compound 3 [IC50 = (37.63 +/- 3.45) mg +/- L(-1)], 6 [IC50 = (178.92 +/- 4.99) mg x L(-1)] and 8 [IC50 = (44.32 +/- 7.02) mg x L(-1)] are of higher inhibitory alpha-glucosidase activity than that of acarbose [IC50 = (1081.27 +/- 12.3) mg x L(-1)]. Compound7 [IC50 = (4.81 +/- 0.96) mg x L(-1)] and 8 [IC50 = (4.16 +/- 0.11) mg L(-1)] showed higher antioxidant activity than that of BHT [IC50 = (35.64 +/- 0.36) mg x L(-1)] and BHA [IC50 = (8.74 +/- 0.39) mg x L(-1)]. Compound 5-8 exhibited inhibitory activity against Fusarium graminearum. Compound 5 and 8 showed inhibitory activity against Botrytis cinerea.

In this study, we have investigated sedimentation velocity ultracentrifugation (AUC-SV), size exclusion chromatography (SEC), and circular dichroism (CD) methods for the detection and quantitation of protein aggregates using recombinant acid alpha-glucosidase (rhGAA) as a model. The results of this study showed that the formation and molecular weight distribution of rhGAA aggregated species were dependent upon the formulation conditions as well as the storage or stress conditions used to induce aggregation. The utility of CD as a probe for non-native, aggregated species was affirmed, as this method was sensitive to rhGAA aggregation levels of < or = 4%. An extensive evaluation of AUC-SV variability was performed using nine levels of spiked rhGAA aggregate that were analyzed on six occasions. Based on our data, the precision of the AUC-SV results increased with increasing levels of aggregate, with a mean RSD of 37.2%. The limit of quantitation (LOQ) for the AUC-SV method, which was based on a Precision criterion of RSD < 20%, was determined to be > or = 3% aggregated rhGAA. The Precision and LOQ of the SEC method, determined using the same rhGAA sample set, was found to be 3.8% and > or = 0.2%, respectively. In general, there was good agreement between the levels of aggregated rhGAA determined using the AUC-SV and SEC methods, with a slight positive bias noted for the AUC-SV results. These studies emphasize the value of applying multiple, well-characterized analytical tools in the evaluation of therapeutic protein aggregation.

The aim of this study was to evaluate the correlation between the secretory function of the male accessory glands and sperm parameters in normospermic controls and infertile patients. One hundred and fifty-nine men were investigated: they were composed of two groups: normospermic (n = 37) and infertile (n = 122) men with altered sperm characteristics. These infertile men were divided into the following groups: asthenozoospermia (n = 38), teratozoospermia (n = 40) and asthenoteratozoospermia (n = 44). The patients underwent semen analysis and measurements of fructose, neutral alpha-glucosidase and citric acid. The level of fructose was significantly decreased in asthenozoospermic and increased in asthenoteratozoospermic men. It was significantly correlated with semen volume, sperm count, motility and morphology. The seminal alpha-glucosidase levels were significantly correlated with semen volume and pH and citric acid was significantly correlated with pH. Thus, alpha-glucosidase and citric acid levels were associated with semen pH. The significant correlation between semen parameters, accessory glands and epididymal functions highlights the relationship between semen and normal genital tract function.

Introduction: Studies that compare the impact of different infectious entities of the male reproductive tract (MRT) on the male accessory gland function are controversial., Materials and Methods: Semen analyses of 71 patients with proven infections of the MRT were compared with the results of 40 healthy non-infected volunteers. Patients were divided into 3 groups according to their diagnosis: chronic prostatitis NIH type II (n = 38), chronic epididymitis (n = 12), and chronic urethritis (n = 21)., Results: The bacteriological analysis revealed 9 different types of microorganisms, considered to be the etiological agents, isolated in different secretions, including: urine, expressed prostatic secretions, semen and urethral smears: E. Coli (n = 20), Klebsiella (n = 2), Proteus spp. (n = 1), Enterococcus (n = 20), Staphylococcus spp. (n = 1), M. tuberculosis (n = 2), N. gonorrhea (n = 8), Chlamydia tr. (n = 16) and, Ureaplasma urealyticum (n = 1). The infection group had significantly (p < 0.05) lower: semen volume, alpha-glucosidase, fructose, and zinc in seminal plasma and, higher pH than the control group. None of these parameters was sufficiently accurate in the ROC analysis to discriminate between infected and non-infected men., Conclusion: Proven bacterial infections of the MRT impact negatively on all the accessory gland function parameters evaluated in semen, suggesting impairment of the secretory capacity of the epididymis, seminal vesicles and prostate. These findings were associated with an infectious related significant increase of semen pH. None of the semen parameters evaluated can be suggested as a diagnostic tool for infection.

The diagnosis of glycogenosis type II is often complicated by the rarity of the condition and the heterogeneity of the clinical manifestations of the disease. It is a progressive, debilitating, and often fatal neuromuscular disorder that manifests as a continuum of clinical phenotypes, which vary with respect to organ involvement, age at onset, and severity. Early diagnosis requires both increased awareness among physicians regarding the clinical characteristics of the disease and fast and reliable acid alpha-glucosidase (GAA) enzyme activity assays to confirm the GAA deficiency. The clinical diagnosis of glycogenosis type II is confirmed by virtual absence (found in infants) and marked reduced activity (found in juveniles and adults) of GAA enzyme in blood samples, cultured fibroblasts, and muscle biopsies. This article specifically highlights the need for early recognition of the clinical manifestation of the disease in infants, juveniles, and adults. Descriptions of the main clinical features of the condition, as well as differential diagnosis are included. In addition, the tests required for a confirmed diagnosis are described, and use of muscle imaging to evaluate muscle pathology is reviewed.

The larvicidal action of the entomopathogen Bacillus sphaericus towards Culex quinquefasciatus is due to the binary (Bin) toxin present in crystals, which are produced during bacterial sporulation. The Bin toxin needs to recognize and bind specifically to a single class of receptors, named Cqm1, which are 60-kDa alpha-glucosidases attached to the apical membrane of midgut cells by a glycosylphosphatidylinositol anchor. C. quinquefasciatus resistance to B. sphaericus has been often associated with the absence of the alpha-glucosidase Cqm1 in larvae midgut microvilli. In this work, we aimed to investigate, at the ultrastructural level, the midgut cells from C. quinquefasciatus larvae whose resistance relies on the lack of the Cqm1 receptor. The morphological analysis showed that midgut columnar cells from the resistant larvae are characterized by a pronounced production of lipid inclusions, throughout the 4th instar. At the end of this stage, resistant larvae had an increased size and number of these inclusions in the midgut cells, while only a small number were observed in the cells from susceptible larvae. The morphological differences in the midgut cells of resistant larvae found in this work suggested that the lack of the Cqm1 receptor, which also has a physiological role as being an alpha-glucosidase, can be related to changes in the cell metabolism. The ultrastructural effects of Bin toxin on midgut epithelial cells from susceptible and resistant larvae were also investigated. The cytopathological alterations observed in susceptible larvae treated with a lethal concentration of toxin included breakdown of the endoplasmic reticulum, mitochondrial swelling, microvillar disruption and vacuolization. Some effects were observed in cells from resistant larvae, although those alterations did not lead to larval death, indicating that the receptor Cqm1 is essential to mediate the larvicidal action of the toxin. This is the first ultrastructural study to show differences in the cell morphology of resistant larvae and further investigation is needed to understand the impact of the lack of expression of midgut enzymes on the physiology of resistant insects.

Glycosylation is considered to be an important reaction for the chemical modification of compounds with useful biological activities. Glycoside hydrolases are biotechnologically attractive enzymes which can be used in synthetic reactions for assembling glycosidic linkages with absolute stereoselectivity at an anomeric centre. Most of these enzymes are commercially available but there is great interest in the search for new biocatalysts with original catalytic characteristics. The marine environment has shown to be a very interesting source for new glycosyl hydrolases for both hydrolytic and synthetic aspects. In particular, Aplysia fasciata a marine herbivorous mollusc has been shown to be a potent producer of a library of glycoside hydrolases applied to the synthesis of glycosidic bonds. The impressive assortment of glycosidases in marine organisms clearly indicates that the potential biodiversity of these enzymes is still largely unexplored and that potential applications of biocatalysts from the sea will increase in the near future.

Objective: The objective of the present study was to examine the effect of glucagon-like peptide-2 (GLP-2) administration in a piglet, juvenile model of short bowel syndrome., Materials and Methods: Four-week-old piglets underwent either a sham operation or 75% small bowel resection. Postoperatively, piglets received either polymeric infant formula diet or the diet and subcutaneous human recombinant GLP-2 (1600 microg/day for 7 days, 800 microg/day thereafter). Food intake was monitored throughout the experiment, and stool and serum samples obtained fortnightly. After the piglets were killed, tissues were obtained from the duodenum, jejunum, ileum, and terminal ileum, and used for morphological and functional analysis., Results: Treatment with GLP-2 resulted in significantly increased numbers of proliferating and apoptotic cells in the ileum of sham and small bowel resection piglets (P < 0.05). GLP-2 administration resulted in decreased weight gain, serum albumin, and disaccharidases in both sham and small bowel resection piglets (P < 0.001 compared with polymeric infant formula diet alone)., Conclusions: This is the first study to our knowledge to examine the effect of GLP-2 administration in a juvenile short bowel syndrome model. Contrary to adult rodent studies, administration of GLP-2 resulted in adverse outcomes including reduced ability to gain weight; decreased serum albumin, tissue maltase, and sucrase; and villous atrophy. We anticipate this information will have important implications for future paediatric clinical trials.

Objective: To detect the changes of biochemical markers in the semen of premature ejaculation patients and investigate the correlation of the markers with premature ejaculation., Methods: Fifty-six premature ejaculation patients and 60 males with normal sexual behavior were enrolled in this experiment. Acid phosphatase, alpha- glucosidase and fructose were assayed by the methods of glucose oxidase, disodium phenyl phosphate and disodium phenyl phosphate respectively., Results: The contents of acid phosphatase, alpha-glucosidase and fructose were (36.37 +/- 31.33) U/ml, (39.97 +/- 22. 09) U/ml and (3.40 +/- 1.92) mg/ml in the premature ejaculation patients and (54. 27 +/- 20. 96) U/ml, (55.71 +/- 16.19) U/ml and (2.55 +/- 1.12) mg/ml in the normal control, respectively, with significant differences in the former two markers between the two groups. The rate of the abnormal content of both acid phosphatase and alpha- glucosidase was 31% and 13% (P < 0.05) , while that of the normal content of the three markers was 10% and 33% in premature ejaculation group and the control, respectively (P < 0. 05 )., Conclusion: The abnormality of both acid phosphatase and alpha-glucosidase is one of the causes of premature ejaculation. Because acid phosphatase and alpha- glucosidase reflect the functions of the prostate and epididymis, we should pay attention to the status of these two organs in the treatment of premature ejaculation.

Objective: To establish a method of determining alpha-glucosidase activity in seminal plasma by semi-automatic biochemistry analyzer., Methods: Alpha-glucosidase activity in seminal plasma from 51 men with normal semen parameters in routine semen analysis were detected by semi-automatic biochemistry analyzer and manual glucose oxidase method, respectively. Then, the intra-assay and inter-assay coefficient variation (CV) and normal reference value were calculated. In the meanwhile, the correlation between the two methods was analyzed., Results: The intra-assay CVs of 2 seminal plasma samples with different alpha-glucosidase activity detected by semi-automatic biochemistry analyzer were 12.63% and 9.13%, and the inter-assay CVs were 10.67% and 13.49%, respectively. The normal reference value for seminal alpha-glucosidase activity detected with semi-automatic biochemistry analyzer ranged from 102.28 to 555.08 U/L. There was a significantly positive correlation between the semi-automatic biochemistry analyzer and the manual glucose oxidase method (r = 0.792, P < 0.01)., Conclusion: The method of determining alpha-glucosidase activity in seminal plasma by semi-automatic biochemistry analyzer, with its simplicity, less cost of time and reagents, and more reliable result, could be applied to clinical laboratory medicine.

Objective: To evaluate the correlation of the level of free L-carnitine with accessory gland markers and its clinical significance., Methods: Semen samples from 30 fertile men and 222 infertile patients were collected by masturbation. The measurement of the semen quality was carried out by computer-assisted semen analysis system. The seminal plasma components of free L-carnitine, alpha-glucosidase, fructose and acid phosphatase were determined. The results obtained were statistically calculated with an SPSS 12.0 program to evaluate the difference between the control group and the infertile one and the correlation of the free L-carnitine levels with the seminal plasma components of alpha-glucosidase, fructose and acid phosphatase., Results: The concentration of free L-carnitine (P < 0.01) and the activity of alpha-glucosidase (P < 0.05) were significantly reduced in the infertile group as compared with the control, with no significant difference in the concentration of fructose and the activity of acid phosphatase between the two groups. There was a statistically significant positive correlation between seminal plasma free L-carnitine level and alpha-glucosidase activity (r = 0.504, P < 0.001., Conclusion: The determination of free L-carnitine level in seminal plasma is a useful test in the evaluation of epididymal function, which may serve as a guidance for the clinical treatment of male infertility as well as for the study on the mechanisms of male reproduction.

Objective: To investigate and analyze the results of the determination of sperm concentration, fructose concentration, alpha-glucosidase and acid phosphatase (ACP) activities in the seminal plasma from different hospitals in the city of Nanjing, so as to provide a basis for the external quality control (EQC) of semen analysis within Jiangsu Province or even the whole country., Methods: Eight samples of quality control products for low and high concentrations sperm count, fructose, alpha-glucosidase and ACP determination were prepared and divided, each detected for the sperm concentration, fructose, alpha-glucosidase and ACP activity, and the coefficient variances (CVs) were calculated. The products were then distributed to 11 hospitals in the city, and the results were collected and analyzed. In addition, the total relative errors (REs) for each product was calculated based on the results after dividing as reference values., Results: The CVs from the 8 samples after dividing were 3.83% - 11.16%. Collected from the 11 hospitals attending EQC were 11 reports of the results of sperm concentration, and 5 the results of fructose, alpha-glucosidase and ACP in seminal plasma. Among the results from different laboratories, those of fructose determination showed the minimal difference (CVs: 8.99% and 3.95% for low and high concentrations, respectively) , next came alpha-glucosidase (CVs: 16.66% and 18.41% for low and high activities, respectively), and ACP determination showed the maximal difference (CVs: 54.12% and 65.58% for low and high activities, respectively). Moreover, the same trend was observed in RE values, as shown in the total REs, which were 11.99% (low concentration) and 20.31% (high concentration) for the determination of fructose in seminal plasma, 22.92% and 27.26% for alpha-glucosidase, 7.34% and 318.35% for ACP in different laboratories, and the maximal RE value was detected in the result of the high-activity ACP sample. Of the 11 hospitals, 6 determined sperm concentration with the computer-assisted semen analysis (CASA) system, and the other 5 with the modified hemocytometer. RE values (148.47% and 187.59% for low and high concentration samples, respectively) and sperm concentrations ([62.74 +/- 16.63] x 10(6)/ml and [163.32 +/- 36.24] x 10(6)/ml) counted with the hemocytometer were significantly higher than those with the CASA system (REs 13.97% and 10.48%; sperm concentrations [24.88 +/- 4.16] x 10(6)/ml and [54.24 +/-23.06] x 10(6)/ml )., Conclusion: The methods of seminal alpha-glucosidase and fructose determination were relatively stable in current andrology laboratories, and the variance range could be accepted. However, the method of seminal ACP determination might be unadaptable to clinical application, and needs to be further improved. Hemocytometer, which significantly overestimated sperm concentration, could not be applied to the assay of sperm concentration.

Objective: To assess the diagnostic value of alpha-glycosidase for epididymal lump induced obstructive azoospermia., Methods: Seventy-six infertile men with normal spermatogenic function were divided according to sperm density into a normal density group (n = 27), an oligospermia group (n = 21) and an obstructive azoospermia group (n = 28), and another 30 fertile males were included as normal controls. Semen plasma alpha-glycosidase, leucocyte count, sexual hormone levels and the diameters of the epididymal lumps were measured and their correlations were analyzed., Results: alpha-Glycosidase in the obstructive azoospermia group was significantly lower than that of the other groups (P <0.05), and negatively correlated with epididymal volume (r = -0.417, P <0.05) and leucocyte count (r = -0.342, P <0.05)., Conclusion: Alpha-Glycosidase has been proved of diagnostic value for epididymal obstructive azoospermia.

Neutral alpha-glucosidase (NAG) activity is considered a functional epididymal marker in several species. Unlike the rat, no NAG activity has been detected in mice. The aims of the present study were to evaluate NAG secretory activity (the supernatant of the incubated tissue) in mouse epididymis and to determine whether it could be used as a functional epididymal marker. Epididymides (whole or in parts) were incubated in the presence or absence of testosterone (10(-5) m) and secretory NAG activity was compared with known positive controls. Furthermore, we compared enzyme activity in epididymides from well-fed and undernourished mice (50% food restriction for 21 days), a model that alters the epididymal maturation processes. Spectrophotometric analysis revealed NAG activity in mouse epididymis (22.6 +/- 3.7 mU g(-1) tissue; n = 4), being higher in the caput. NAG activity was statistically higher in the caput than in the corpus and in the cauda. No significant differences existed between the caput NAG activity and complete epididymis NAG activity. In undernourished mice, we confirmed changes in epididymal maturation observed previously (i.e. increased number of immature spermatozoa and diminution of the sperm concentration). Concordantly, the epididymides of undernourished mice exhibited decreased enzyme secretory activity, which increased to values similar to those seen in controls following incubation in the presence of testosterone (22.5 +/- 2.6, 12.5 +/- 1.0 and 22.4 +/- 3.7 mU g(-1) tissue, n = 9 in control (n = 7), undernourished (n = 9) and undernourished + testosterone groups (n = 9), respectively). In conclusion, NAG activity was detected in mouse epididymis. Although the present study supports the possibility of using NAG as an epididymal marker, more studies are necessary to effectively prove that NAG activity can be used as an epididymal marker.

We attempted to provide an instruction for the standardization of alpha-glucosidase level determination and quality controls within the same laboratory and among different laboratories. Each of 51 semen samples was divided into two aliquots, centrifuging at 1000 g for 10 min or 3000 g for 15 min. The alpha-glucosidase level and sperm concentration of each sample were assayed. The alpha-glucosidase level in seminal plasma obtained at 3000 g for 15 min centrifugation was significantly lower than that at 1000 g for 10 min (p = 0.001). An additional 6 samples of seminal plasma with or without phenylmethylsulfonyl fluoride (PMSF), obtained at 3000 g for 15 min centrifugation, were frozen for 20 days, and each of 6 samples was determined for their alpha-glucosidase levels after thawing every other day. There was no significant difference between alpha-glucosidase levels in seminal plasma regardless of the presence of PMSF. The alpha-glucosidase level increased with the length of abstinence period. In conclusion, centrifugal velocity and abstinence time should be standardized in the alpha-glucosidase determination. Frozen seminal plasma may serve as the sample for quality control among clinical laboratories.

To evaluate the effect of chymotrypsin on the examination of alpha-glucosidase activity in seminal plasma, thirty-nine samples of fresh liquefied semen with or without chymotrypsin and forty-eight samples of fresh un-liquefied semen with chymotrypsin were determined for the total alpha-glucosidase activity in seminal plasma. The total alpha-glucosidase level of each sample was assayed by the method of glucose oxidase. The correlations between alpha-glucosidase level and semen parameters, including semen volume, pH, sperm concentration, grade a and b motility and total motility, were analyzed with SPSS 11.0 software. The results showed that chymotrypsin had no effect on seminal alpha-glucosidase activity determination. Chymotrypsin could improve the liquefaction for un-liquefied semen, and there was no significant difference of alpha-glucosidase activity between liquefied and un-liquefied semen samples. There were significantly positive correlations between seminal alpha-glucosidase activity (U/ml) and sperm concentration (r = 0.338, p = 0.015) and between total alpha-glucosidase activity (U/ejaculate) and semen volume (r = 0.677, p = 0.000). However, there was no significant correlation between alpha-glucosidase level (U/ml) and semen volume, pH, sperm motility or grade a and b motility (r = -0.234 approximately 0.077, p = 0.099 approximately 0.993). The data indicated that chymotrypsin could be added into the un-liquefied semen samples for alpha-glucosidase activity determination, and there were different correlations between seminal alpha-glucosidase level and various semen parameters.