This research communication reports the effects of a compound enzyme preparation consisting of fibrolytic (cellulase 3500 CU/g, xylanase 2000 XU/g, β-glucanase 17 500 GU/g) and amylolytic (amylase 37 000 AU/g) enzymes on nutrient intake, rumen fermentation, serum parameters and production performance in primiparous early-lactation (47 ± 2 d) dairy cows. Twenty Holstein–Friesian cows in similar body condition scores were randomly divided into control (CON, n = 10) and experimental (EXP, n = 10) groups in a completely randomized single-factor design. CON was fed a basal total mixed ration diet and EXP was dietary supplemented with compound enzyme preparation at 70 g/cow/d. The experiment lasted 4 weeks, with 3 weeks for adaptation and then 1 week for measurement. Enzyme supplementation significantly increased diet non-fibrous carbohydrates (NFC) content as well as dry matter intake (DMI) and NFC intake (P < 0.05). EXP had increased ruminal butyrate and isobutyrate percentages (P < 0.01) but decreased propionate and valerate percentages (P < 0.05), as well as increased serum alkaline phosphatase activity and albumin concentration (P ≤ 0.01). Additionally, EXP had increased milk yield (0.97 kg/d), 4% fat corrected milk yield and energy corrected milk yield, as well as milk fat and protein yield (P < 0.01). In conclusion, dietary supplementation with a fibrolytic and amylolytic compound enzyme preparation increased diet NFC content, DMI and NFC intake, affected rumen fermentation by increasing butyrate proportion at the expense of propionate, and enhanced milk performance in primiparous early-lactation dairy cows.

We previously reported that the addition of a specified mulberry fruit extract (MFE) to rice consistently reduces post-prandial glycaemic (PPG) and post-prandial insulinemic (PPI) responses. This research tested whether this effect generalises to a broad range of rice types, reflecting the wide variation in rice characteristics known to influence glycaemic responses. In a randomised, balanced, partial factorial crossover design, Sona Masoori (SM), Bora Saul (BS), Gobindobogh (Gb) and Banskati (Bn) rices were tested with and without 0·37 g MFE. Healthy, normal-weight Indian adults (N 120) each consumed four of the eight possible boiled rice meals, all containing about 50 g available carbohydrate. The primary outcome was the effect of MFE on PPG, expressed as the percentage change in the positive, incremental AUC over 2 h. The mean effect of MFE on PPG for all rice types combined was −11·4 % (P < 0·003). The reduction in PPG was in a qualitatively similar range for all rice types (–9·8 to −15·1 %), and this was statistically significant for Bn. MFE also reduced the corresponding PPI response to all rice types combined by a mean of 10·1 % (P < 0·001; range −6·1 to −13·4 %), and the reduction in PPI was statistically significant for SM, Gb and BS. In conclusion, addition of 0·37 g MFE modestly reduced PPG and PPI responses to rices in general, and the effects were statistically significant for specific rice types.

The fish paste product, fish balls ‘tsumire’, is a traditional type of Japanese food made from minced fish as well as imitation crab, kamaboko and hanpen. Although tsumire is known as a high-protein and low-fat food, there is a lack of scientific evidence on its health benefits. Hence, we aimed to investigate the effects of tsumire intake on organ weight and biomarker levels in Sprague–Dawley rats for 84 d as a preliminary study. Six-week-old male Sprague–Dawley rats were divided into two groups: group I, fed normal diets, and group II, fed normal diets with 5 % dried tsumire. Throughout the administration period, we monitored their body weight and food intake; at the end of this period, we measured their organ weight and analysed their blood biochemistry. No significant differences were observed with respect to body weight, food intake, organ weight and many biochemical parameters between the two groups. It was found that inorganic phosphorus and glucose levels were higher in group II rats than in group I rats. On the other hand, sodium, calcium, amylase and cholinesterase levels were significantly lower in group II than in group I. Interestingly, we found that the levels of aspartate aminotransferase, alanine transaminase, lactate dehydrogenase and leucine aminopeptidase in group II were significantly lower than in group I, and that other liver function parameters of group II tended to be lower than in group I. In conclusion, we consider that the Japanese traditional food, ‘tsumire,’ may be effective as a functional food for human health management worldwide.

An oral starch administration trial was used to evaluate glucose homoeostasis in grass carp (Ctenopharyngodon idella) and Chinese longsnout catfish (Leiocassis longirostris Günther). Fish were administered with 3 g of a water and starch mixture (with 3:2 ratio) per 100 g body weight after fasting for 48 h. Fish were sampled at 0, 1, 3, 6, 12, 24 and 48 h after oral starch administration. In grass carp, plasma levels of glucose peaked at 3 h but returned to baseline at 6 h. However, in Chinese longsnout catfish, plasma glucose levels peaked at 6 h and returned to baseline at 48 h. The activity of intestinal amylase was increased in grass carp at 1 and 3 h, but no significant change in Chinese longsnout catfish was observed. The activity of hepatic glucose-6-phosphatase fell significantly in grass carp but change was not evident in Chinese longsnout catfish. The expression levels and enzymic activity of hepatic pyruvate kinase increased in grass carp, but no significant changes were observed in the Chinese longsnout catfish. Glycogen synthase (gys) and glycogen phosphorylase (gp) were induced in grass carp. However, there was no significant change in gys and a clear down-regulation of gp in Chinese longsnout catfish. In brief, compared with Chinese longsnout catfish, grass carp exhibited a rapid increase and faster clearance rate of plasma glucose. This effect was closely related to significantly enhanced levels of digestion, glycolysis, glycogen metabolism and glucose-induced lipogenesis in grass carp, as well as the inhibition of gluconeogenesis.

Mungbean seeds, despite being protected inside the pod, are susceptible to pre-harvest sprouting (PHS) following rainfall due to lack of fresh seed dormancy (FSD), which deteriorates the quality of the seed/grain produced. Therefore, development of mungbean cultivars with short (10–15 days) period of FSD has become important to curtail losses incurred by PHS. In this study, we investigated variations in PHS, fresh seed germination (FSG) and activity of α-amylase enzyme in diverse mungbean genotypes. There was a wide variation in PHS tolerance and FSG among 163 genotypes examined and 14 genotypes were found to be tolerant (<20%) to PHS. Seed germination in a pod, a measure used to evaluate PHS, varied from 7.14% in germplasm accession Chamu 4 to 82.52% in cultivated variety IPM 2–3. There was a marked increase in α-amylase activity in genotypes showing high FSG and PHS, especially at 48 and 72 h after germination as compared with PHS tolerant genotypes. Therefore, α-amylase can be used as an effective biochemical marker to evaluate a large number of mungbean genotypes for FSD and PHS. Also, the variation in seed germinability as found in this study could be further used for mungbean improvement programme.

An enzymatic (amylase) pretreatment was developed that allows quantitative recovery of the parent compound from soil samples receiving the pesticide in starch-encapsulated formulation. The pretreatment involves adding amylase, dissolved in a phosphate buffer solution, to soil samples and incubating the resulting slurry at 50 C for 1 h. Efficiency of this pretreatment was tested on two soils, a silty clay and a loamy sand, receiving atrazine and alachlor in the starch-encapsulated formulation. The inexpensive method adds approximately 1 h to the extraction procedure and a twofold increase in percentage recovery of starch-encapsulated atrazine. The enzyme pretreatment is not necessary for quantitative recovery of pesticides that have rapid rates of release such as alachlor. The method will be useful for studying starch encapsulation pesticide behavior in the soil environment.

Giardia lamblia localize and multiply in the small intestine and may cause acute or chronic diarrhoea with malabsorption of fat, protein and other nutrients. Abnormal pancreatic function has been documented in giardiasis and trophozoites directly inhibit pancreatic lipase activity in vitro. The aim of this study was to examine the effect of Giardia trophozoites on pancreatic trypsin, chymotrypsin and amylase activity in vitro. Axenically cultured Giardia trophozoites (Portland-1 stock) were incubated with a range of concentrations of trypsin, chymotrypsin and amylase and enzyme activity assayed over time. Tryptic activity was decreased after incubation with Giardia trophozoites. This reduction was time dependent and linear over the incubation period of 2 h. At a trypsin concentration of 18 BAEE units/ml, there was a 35.5±4% reduction in enzyme activity after 2 h compared to controls. The total amount of activity lost was proportional to the initial trypsin concentration up to 185 BAEE units/ml. At this initial concentration, the activity was reduced by 46.5±3 units/ml after 2 h. Above this concentration, little further loss of enzyme activity was seen. To investigate the nature and specificity of this effect, similar experiments were conducted using killed trophozoites and with a related protozoan. Trichomonas vaginalis. No loss of enzyme activity was evident. Media previously incubated for 2 h with trophozoites did not diminish tryptic activity. Trophozoites had no effect on chymotrypsin or amylase activities over the range of concentrations tested. Giardia trophozoites directly diminish pancreatic tryptic activity in vitro, by an active process which appears specific and not mediated via a secretory product. This mechanism may enhance parasite survival in the small intestine by allowing trophozoites to escape destruction by luminal proteases and may also contribute to malabsorption.

Seeds of the common bean, Phaseolus vulgaris, contain two inhibitors of mammalian and insect α-amylases (αAls) that show specificity towards the amylases of different insect species. Expression in pea (Pisum sativum) and azuki bean (Vigna angularis) of a chimeric gene consisting of the cDNA of bean αAl-1 and a seed-specific promoter makes the seeds of these legumes resistant to three species of Old World bruchids whose amylases are inhibited by αAl-1. This was the first successful genetic engineering of insect resistance in seeds. To understand the basis of the specificity between amylases and inhibitors we cloned a second bean inhibitor (αAl-2) with different specificity, and we cloned the cDNA of the New World bruchid, Zabrotes subfasciatus. The amylase of this bruchid is inhibited by αAl-2, but not by αAl-1. Knowledge of the amino acid sequences and of the three-dimensional structure of the pancreatic α-amylase–αAl-1 complex allows us to predict the peptide domains and amino acids of the proteins that are important for protein–protein recognition and inhibition of enzyme activity.

Insecticide resistance is frequently associated with fitness disadvantages in the absence of insecticides. However, intense past selection with insecticides may allow the evolution of fitness modifier alleles that mitigate the cost of insecticide resistance and their consequent fitness disadvantages. Populations of Sitophilus zeamais with different levels of susceptibility to insecticides show differences in the accumulation and mobilization of energy reserves. These differences may allow S. zeamais to better withstand toxic compounds without reducing the beetles' reproductive fitness. Enzymatic assays with carbohydrate- and lipid-metabolizing enzymes were, therefore, carried out to test this hypothesis. Activity levels of trehalase, glycogen phosphorylase, lipase, glycosidase and amylase were determined in two insecticide-resistant populations showing (resistant cost) or not showing (resistant no-cost) associated fitness cost, and in an insecticide-susceptible population. Respirometry bioassays were also carried out with these weevil populations. The resistant no-cost population showed significantly higher body mass and respiration rate than the other two populations, which were similar. No significant differences in glycogen phosphorylase and glycosidase were observed among the populations. Among the enzymes studied, trehalase and lipase showed higher activity in the resistant cost population. The results obtained in the assays with amylase also indicate significant differences in activity among the populations, but with higher activity in the resistant no-cost population. The inverse activity trends of lipases and amylases in both resistant populations, one showing fitness disadvantage without insecticide exposure and the other not showing it, may underlay the mitigation of insecticide resistance physiological costs observed in the resistant no-cost population. The higher amylase activity observed in the resistant no-cost population may favor energy storage, preventing potential trade-offs between insecticide resistance mechanisms and basic physiological processes in this population, unlike what seems to take place in the resistant cost population.

Podisus nigrispinus (Dallas) is a common predator in agricultural and natural systems in Neotropical America. Its feeding strategy involves extra-oral digestion and to better understand this process its salivary glands were extracted and subjected to morphological and preliminary enzyme characterization. The salivary glands of P. nigrispinus are formed by a pair of main and accessory gland complexes. The main salivary glands are further divided into an anterior and a posterior lobe. The compartmentalization of the salivary gland complex is likely to be important for the production, activation and release of the digestive enzymes used in the extra-oral digestion of prey items. Proteases and lipase, important digestive enzymes involved in zoophagy, were detected in the salivary glands of P. nigrispinus. The prevailing trypsin-like protease activity was characterized by using the serine-protease substrate N-α-benzoyl-L-Arg-p-nitroanilidine (L-BApNA) and the trypsin inhibitors tosyl-L-lysine chloromethyl ketone (TLCK) and benzamidine. The KM value obtained for trypsin-like activity was 1.57 mm and the different peaks of optimum pH and temperature activity suggest the presence of multiple forms of this enzyme in P. nigrispinus. Detection of amylase activity in the salivary glands of this predator suggests its ability to digest starch and obtain nutrients from plants, which may have adaptative value under prey scarcity.

Cereal grain germination and early seedling growth involve the co-ordinated action of endosperm and embryo tissues to mobilize the storage reserves of the starchy endosperm. This mobilization is accomplished by hydrolases secreted from the aleurone and scutellar tissues. The breakdown products are then transported to the growing seedling by the scutellum. This resource-harvesting system is regulated at multiple levels. One well-defined aspect of control is brought about by the hormone gibberellin (GA). Gibberellin is released from the embryo upon imbibition and activates the aleurone cells. The secretory apparatus of the aleurone then proliferates, supporting increased hydrolase synthesis and secretion to degrade the starchy endosperm. The molecules that regulate this response to GA are now being increasingly characterized. Elements such as cGMP, calcium, calmodulin and protein kinases are well known as regulators in other eukaryotic cell types and are emerging as key control factors in the aleurone hormone response. However, superimposed upon this molecular regulatory system is another level of control, the structural pattern of tissues and stored macro- molecules that was laid down during grain development. It is the interaction of these structural motifs combined with the molecular regulatory mechanisms that ensure the appropriate timing and positioning of hydrolase production and endosperm reserve mobilization. This integrated control system ensures an extended release of nutrients to fuel early seedling growth.

Zymography is an electrophoretic technique by which enzyme activity can be visualized directly on a polyacrylamide gel as discrete bands. A modified, more rapid technique for amylase zymography is described and compared with previously published methods. Whereas previous methods are based on 0.1 M acetate buffer as substrate buffer, our method utilizes 50mM Tris buffer containing Ca2+, Na+, NaN3 and Triton X-100 which helps rapid hydrolysis of the starch and stabilization of the enzyme. The staining procedure, previously requiring overnight incubation of the gel in iodine solution at 4 °C, has been reduced to 5 min at room temperature. Both methods gave rise to comparable levels of enzyme activity on polyacrylamide gels. Our modified method requires 8 h to complete the whole zymographical procedure instead of 18-20 h as in previous methods.

The range of processes regulated by gibberellins (GAs) covers all aspects of the life history of the plant from seed germination to vegetative growth and flowering. In seeds there has been an intensive search, using the techniques of both biochemistry and cell biology, for the regulatory molecules linking GA perception to gene regulation and the events of germination. Although a GA receptor has yet to be identified, the site of perception has been localized to the plasma membrane. Calmodulin, Ca2+ and cGMP have also been identified as elements of the GA signal transduction pathway. These regulators parallel many of the signalling elements identified in the transduction of other signals such as phytochrome and ABA. Studies of GA-regulated gene expression, principally of the α-amylases of cereal aleurone, have identified core GA-responsive promoter elements, such as the gibberellin response element (GARE), box-1 and pyrimidine boxes, as well as elements that may lend specificity to GA-regulated expression, such as the Opaque-2-similar element (O2S), and TRE and CRE motifs. One of the most striking features of all of these studies of the molecular basis of GA action is the interaction of GA-dependent regulatory elements with those of other factors such as ABA. GA-response elements also appear to be conserved between disparate GA-response systems. For example, Myb transcription factors appear to regulate a multitude of GA-induced genes in cereal aleurone as well as to alter GA responses when expressed in Arabidopsis. Thus the study of GA signal transduction and response systems is highlighting the conservation of regulatory elements used by plants. These common factors, used by distinct signal transduction systems, provide a molecular basis for the integration of the GA signal with other growth regulators that is the hallmark of plant growth and development.

Malnutrition in infancy is a global problem which leads to retardation of childhood growth and development. There is a pressing need to improve weaning strategies for infants of the developing world. Traditional Gambian weaning foods are watery and of low energy density, but addition of energy in the form of fat and carbohydrate leads to thick, viscous gruels which are difficult to ingest. Partial digestion with amylase (EC3·2·1·1)-rich flour reduces their viscosity while retaining their energy density. The aim of the present study was to measure the digestibility of a maize-based weaning food, before and after amylase digestion, in malnourished children using a 13C breath test. Ten children (aged 7–16 months; mean weight-for-age Z score −0·8) received isovolumetric and isoenergetic quantities of a maize-based weaning food naturally abundant with 13C. Breath samples were collected at intervals of 30 min for 5 h thereafter and 13CO2 enrichment was measured by isotope-ratio mass spectrometry. Percentage dose of 13C recovered increased from a mean 13·7 (SD 3·7)% before, to 18·3 (SD 5·6)% after ingestion of amylase-treated weaning foods (P < 0·1). There was a significant inverse relation between age and weight, and percentage dose of 13C recovered in children receiving amylase-treated feeds. There were no differences in concentrations of amylase in saliva of infants or breast milk of their mothers. Partial digestion of supplementary foods may improve the nutrition of undernourished weaning children, not only by reducing their viscosity, thereby increasing ingestion, but also by improving their digestion and thereby their absorption.

Activity of digestive enzymes (trypsin, chymotrypsin and amylase) was studied in relation with food in Penaeus vannamei larvae first stages. Microparticles containing casein involved the decrease of soluble protein content and enzyme activities. Specific trypsin activity but not chymotrypsin was correlated (p < 1%) to casein level between 10 and 60% in microparticles. Cellulose fibres in food seemed to have little effect on digestive enzymes. Starch, between 1 and 20% in microparticles, had no influence on specific amylase activity. The effect of different feeding conditions were also tested on growth. No clear relationship between growth and enzyme is established since only trypsin is concerned in the case of a casein dose effect in the food. The protein source (casein, gelatin, squid meal or fish protein soluble concentrate) as well as the carbohydrate quality (soluble starch, standard corn starch, amylopectin corn starch or pregelatinized corn starch) were tested. Squid meal stimulated significantly chymotrypsinactivity (p < 2%) while trypsin activity decreased with fish protein soluble concentrate p < 5%). Source of carbohydrate seemed very important, amylase activity increased significantly ( p < 2%) with corn starch. These adaptations are not correlated to growth performances since squid meal or fish protein soluble concentrate improved larval development until zoe 3 substage. These results suggest a specific induction of digestive enzymes to the food quality in larvae independent of growth performances.

Some properties of the gut amylase of Sophrorhinus insperatus Faust have been studied. The enzyme has maximum activity at pH 5.0 and 45°C. It is activated by C−, Ca2+ and Cd2+ tons and inhibited by Cu2+ and Hg2+ lons. Low Michaelis constants of 0.26 mg/ml and 032 mg/ml were obtained for the larva and adult gut amylases. Enzymatic activity was found to be consistently higher in the larva than in adult stage, probably suggesting the larval stage as the most destructive stage. Extracts of Cola nitida (Vent) Schott and Endlicher and C. acuminata (Pal de Beauv) Schott and Endlicher, inhibited amylase activity in vitro at varying magnitudes. The higher level of inhibitors in C. acuminata is an index of its low susceptibility to weevil attack both on the field and at storage.

Amylase, sucrase and cellulase were detected in the alimentary canals of workers and soldiers of Amitermes evuncifer Silvestri, but only sucrase and cellulase were found in the alimentary canal of the alates. Enzyme activities varied in different parts of the alimentary canal. That high enzyme activities were observed in the guts of the workers (than those of the soldiers) is due to the fact that the workers forage for food which they digest partially before feeding other members of the colony through trophallaxis. The presence of enzymes in the alimentary canals of alates and soldiers indicates that the partially digested food, from the workers, undergoes further digestion in their systems.