Total Protease Activity Research Articles

Effect of musical stimuli and white noise on rainbow trout (Oncorhynchus mykiss) growth and physiology in recirculating water conditions

Two musical stimuli transmissions (Mozart and Romanza) as compared with white noise treatment or control, both resulted in significantly higher growth performance in juvenile (6.7±0.12g) rainbow trout (Oncorhynchus mykiss) reared for 14 weeks. Carcass chemical composition and fatty acid composition (% of total fatty acids and mg/g carcass wet weight) did not differ among experimental treatments. The same was observed with regard to liver composition. Brain serotonin (5-HT) and its metabolite (5-HIAA) levels were increased in Mozart fish groups compared to all other treatments. However, serotonergic activity (as defined by the 5-HIAA: 5-HT ratio) for the Mozart groups was similar to control groups and was increased in Romanza and white noise fish groups. Brain dopaminergic activity (as defined by the DOPAC: DA ratio, i.e. dopamine metabolite to dopamine levels) was lower in Mozart compared to control fish groups. Differences were also observed as regards total carbohydrase and protease activity in several parts of the digestive tract.In conclusion, the results of the present data indicate that the musical stimuli transmitted were beneficial for the growth performance of rainbow trout. The fact that white noise treatment presented no major differences from control fish groups suggests that this specific stimulus was neither beneficially nor negatively perceived by rainbow trout, while it further supports the hypothesis that it is the musical stimuli per se that make all the difference.

Programmed cell death begins earlier in the mesophyll cells of tulip petals than in the epidermal cells

Based on histological observations, the onset of programmed cell death (PCD) occurs earlier in the mesophyll cells of petals than in the epidermal cells, but few biochemical studies have addressed the onset of PCD in mesophyll cells. We sampled epidermal cells and, separately, mesophyll cells from tulip petals, and then used biochemical methods to examine the onset of PCD in the two different tissue types. DNA laddering and collapse of cells had begun in mesophyll cells before petals had visibly wilted, but DNA laddering was not evident in epidermal cells at that point. Changes in protein content and total proteinase activity during senescence also supported the conclusion that the onset of PCD occurred in mesophyll cells before it occurred in epidermal cells. This study is the first to use multiple biochemical methods of analysis, as well as microscopic observations, to demonstrate that PCD begins earlier in the mesophyll cells than in the epidermal cells of tulip petals.

Response of Lymantria dispar (Lepidoptera: Lymantriidae) larvae from differently adapted populations to allelochemical stress: Effects of tannic acid

The effects of tannic acid on mean values and genetic variation in fitness-related traits (mass, relative growth rate) and specific activities of digestive enzymes (total proteases, a-glucosidase and lipase), and genetic variation in their plasticity, were investigated in fifth instar larvae of Lymantria dispar L. (Lepidoptera: Lymantriidae) originating from two populations with different host use histories (oak and locust-tree). The two populations did not differentiate with respect to fitness-related traits, i.e. adverse effects of tannic acid were similar in both populations. However, Robinia larvae, which originated from the locust-tree forest, were characterized by higher total protease and lipase activity and lower a-glucosidase activity than Quercus larvae, which originated from the oak forest. Higher plasticity of lipase and lower plasticity of a-glucosidase in response to tannic acid were also recorded. Quantitative genetic analysis revealed mostly significant expression of genetic variation in the examined traits and trait plasticity, suggesting the potential for evolution of adaptive plastic responses to new environmental conditions and presence of a stressor. The genetic correlations observed between the environments significantly differed from one, which indicates there are no constraints on the evolution of trait plasticity.

Enzymatic Characteristics and Growth Parameters of Ornamental Koi Carp (Cyprinus carpio var. Koi) Larvae Fed by Artemia nauplii and Cysts

The purpose of this study was to compare the larval growth and digestive enzyme activities of koi carp (Cyprinus carpio var. Koi) larvae fed on Artemia nauplii (AN) and decapsulated cyst (ADC). The experiment was carried out in triplicate and lasted 27 days. Specific growth and survival rates, specific activities of total protease, lipase, amylase and chitinase were examined in experimental groups. At the end of the experiment for group fed with AN total length, weight and SGR were 20.56±3.23 mm, 0.9±0.05 g and 2.03 % d−1 , respectively and for group fed with ADC it was 28.16±4.81 mm, 0.28±0.16 g and 6.02 d−1 (P 0.05). Besides, there were significant differences on specific enzyme activities (total protease, amylase, chitinase, lipase) (P<0.05). Obtained results suggested that koi larvae fed by ADC presented relatively higher growth parameters.

An in-vitro evaluation of the efficacy of garlic extract as an antimicrobial agent on periodontal pathogens: A microbiological study

With the rise in bacterial resistance to antibiotics, there is considerable interest in the development of other classes of antimicrobials for the control of infection. Garlic (Allium sativum Linn.) has been used as medicine since ancient times and has long been known to have antibacterial, antifungal, and antiviral properties. This study was undertaken to assess the inhibitory effect of garlic on Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans, to assess the time-kill curve of P. gingivalis and A. actinomycetemcomitans, and to determine the antiproteolytic activity of garlic on P. gingivalis. Ethanolic garlic extract (EGE) and aqueous garlic extract (AGE) were prepared and the inhibitory effects of these extracts for two periodontal pathogens (P. gingivalis and A. actinomycetemcomitans) were tested. Antiproteolytic activity on protease of P. gingivalis was determined. 25 microliter (μl), 50 μl, and 75 μl of AGE showed 16 mm, 20 mm, and 25 mm zone of inhibition, respectively, on P. gingivalis. The AGE showed greater bacteriostatic activity against the P. gingivalis with minimum inhibitory concentration determined at 16.6 μl/ml. The time-kill assay of AGE and EGE were compared for P. gingivalis and A. actinomycetemcomitans. AGE showed better antiproteolytic activity on total protease of P. gingivalis compared to the EGE. Thus, the study concludes the antimicrobial activity of garlic extract against periodontal pathogens, P. gingivalis, A. actinomycetemcomitans. Its action against P. gingivalis includes inhibition of total protease activity, and this raises the possibility that garlic may have therapeutic use for periodontitis and possibly other oral infections.

A new quorum-sensing inhibitor attenuates virulence and decreases antibiotic resistance in Pseudomonas aeruginosa

Quorum sensing (QS) has been a novel target for the treatment of infectious diseases. Here structural analogs of Pseudomonas aeruginosa autoinducer N-acyl homoserine lactone (AHL) were investigated for QS inhibitor (QSI) activity and a novel QSI was discovered, N-decanoyl-L-homoserine benzyl ester (C2). Virulence assays showed that C2 down-regulated total protease and elastase activities, as well as the production of rhamnolipid, that are controlled by QS in P. aeruginosa wild-type strain PAO1 without affecting growth. C2 was also shown to inhibit swarming motility of PAO1. Using a microdilution checkerboard method, we identified synergistic interactions between C2 and several antibiotics, tobramycin, gentamycin, cefepime, and meropenem. Data from real-time RT-PCR suggested that C2 inhibited the expression of lasR (29.67%), lasI (21.57%), rhlR (28.20%), and rhlI (29.03%).

Regulation of pericellular proteolysis by hepatocyte growth factor activator inhibitor type 1 (HAI-1) in trophoblast cells

Hepatocyte growth factor activator inhibitor type 1/serine protease inhibitor Kunitz type 1 (HAI-1/SPINT1) is a membrane-bound Kunitz-type serine protease inhibitor that is abundantly expressed on the surface of cytotrophoblasts, and is critically required for the formation of the placenta labyrinth in mice. HAI-1/SPINT1 regulates several membrane-associated cell surface serine proteases, with matriptase being the most cognate target. Matriptase degrades extracellular matrix protein such as laminin and activates other cell surface proteases including prostasin. This study aimed to analyze the role of HAI-1/SPINT1 in pericellular proteolysis of trophoblasts. In HAI-1/SPINT1-deficient mouse placenta, laminin immunoreactivity around trophoblasts was irregular and occasionally showed an intense punctate pattern, which differed significantly from the linear distribution along the basement membrane observed in wild-type placenta. To explore the molecular mechanism underlying this observation, we analyzed the effect of HAI-1/SPINT1 knock down (KD) on pericellular proteolysis in the human trophoblast cell line, BeWo. HAI-1/SPINT1-KD BeWo cells had increased amounts of cellular laminin protein and decreased laminin degradation activity in the culture supernatant. Subsequent analysis indicated that cell-associated matriptase was significantly decreased in KD cells whereas its mRNA level was not altered, suggesting an enhanced release and/or dislocation of matriptase in the absence of HAI-1/SPINT1. Moreover, prostasin activation and pericellular total serine protease activities were significantly suppressed by HAI-1/SPINT1 KD. These observations suggest that HAI-1/SPINT1 is critically required for the cell surface localization of matriptase in trophoblasts, and, in the absence of HAI-1/SPINT1, physiological activation of prostasin and other protease(s) initiated by cell surface matriptase may be impaired.

The Th2 cytokine, interleukin‐4, abrogates the cohesion of normal stratum corneum in mice: implications for pathogenesis of atopic dermatitis

There is mounting evidence that Th2 cytokines adversely affect skin barrier functions and contribute to the pathogenesis of atopic dermatitis (AD). AD is also characterized by abnormal cohesion in the stratum corneum (SC). However, the contribution of Th2 cytokines to this abnormality remains unknown. This study examined the effects of IL-4, a prototypic Th2 cytokine, on the cohesion of the SC. Structural and physiological assessments revealed that repeated intradermal injections of IL-4 compromised the cohesion of the SC of normal hairless mice. Two potential mechanisms were explored to account for the altered cohesion. First, IL-4 decreased the amount of corneodesmosomes and down-regulated the expression of desmoglein 1, but not of corneodesmosin (CDSN) or loricrin expression, in murine skin and in cultured human keratinocytes (KC). IL-4 did not affect the skin surface pH, and in situ zymography revealed no net change in total serine protease activity in the IL-4-treated SC. Yet, IL-4 enhanced expression of kallikrein (KLK)7, while simultaneously down-regulating KLK5 and KLK14. Finally, IL-4 did not alter the expression of the lympho-epithelial Kazal-type inhibitor (LEKTI) in KC. This study suggests that IL-4 abrogates the cohesion of SC primarily by reducing epidermal differentiation.

Digestive enzyme activity in juvenile Nile tilapia (Oreochromis niloticus, L) submitted to different dietary levels of shrimp protein hydrolysate

The effect of different dietary concentrations of shrimp protein hydrolysate (SPH) on digestive enzyme activity of Nile tilapia juveniles was evaluated. SPH concentrations in diets were 0, 15, 30 and 60 g kg−1 (treatments SPH0, SPH15, SPH30 and SPH60, respectively). Hemoglobin, azocasein, BApNA (Nα-benzoyl-dl-arginine-p-nitroanilide), SApNA (Suc-Ala-Ala-Pro-Phe p-nitroanilide), aminoacyl of β-naphthylamide and starch were used as substrates for enzyme activity determinations. The activity of total alkaline protease was significantly higher (P < 0.05) in fish under SPH15 and SPH60 treatments than in the control (SPH0). However, the effect was not dose-dependent. Substrate-SDS-PAGE was also performed to evaluate changes in the profile of Nile tilapia digestive proteases caused by SPH. Substrate-SDS-PAGE revealed 12 active proteolytic bands, eight of which responded to SPH dietary incorporation. Inhibition substrate-SDS-PAGE indicated a decrease in the activity of three enzymes, with trypsin activity decreasing with the increase of SPH concentration, whereas the opposite occurred for two aminopeptidases. Distinct protease profiles were also found for each treatment, suggesting adaptability of digestive proteases from Nile tilapia to the different diets.

Proteolytic activities in Phaseolus vulgaris cotyledons under copper stress

The changes in the protease activities of bean cotyledons were investigated in response to copper stress. Assays using synthetic substrates and specific protease inhibitors followed by activity measurements and electrophoresis analysis allowed to study the classes of enzymes involved in the storage protein mobilization during the germination of bean (Phaseolus vulgaris L) seeds, and then identify which ones were affected in the presence of 200μM CuCl2 in the imbibition medium. Copper treatment affected embryo growth and total protease activity. The results of SDS-gelatin-PAGE show that Cu excess led to a decrease in protease activity of 45 to 66kDa. Moreover, cysteine-, aspartic- and metallo-protease activities were markedly lowered under copper stress, while serine-protease one was enhanced as well as its activity dependent abundance in comparison with control. However, the relative distribution of major cysteine protease in H2O-germinated seeds was significantly diminished after Cu exposure. Thus, copper excess can disturb the nitrogen freeing from reserve tissues at enzymatic level; differential responses of protease classes are discussed, notably, cysteine protease in the way of storage protein mobilization and serine protease in protective mechanism one.

Isolation and selection of some Bacillus strains capable of nattokinase production

Nattokinase enzyme, found in Natto in 1980 by Dr. Sumi (Japan), can dissolve blood clots. In this paper, two strains of Bacillus sp.7.2 and Bacillus sp.NP3 were selected for nattokinase production. On the steamed soybean substrate, at room temperature, after 40 hours of fermentation, the nattokinase activity was about 470 FU/g, which accounted for about 70-85% as compared to the total protease activity. These results show that we can produce the nattokinase enzyme at low cost for healthcare of the Vietnamese people, and are not dependent on imported functional foods containing nattokinase enzyme.

Expression of a nematode symbiotic bacterium-derived protease inhibitor protein in tobacco enhanced tolerance against Myzus persicae

Fusion proteins of a protease inhibitor from an entomopathogenic nematode symbiotic bacterium (PIN1) and green fluorescent protein (GFP) were expressed in tobacco (Nicotiana tobacum cv. Samsun NN). The PIN1-GFP protein expressed under the control of the CaMV-35S promoter was detected in leaves of transgenic tobacco plants. The effect of PIN1 on anti-pest activity for Myzus persicae was tested by feeding neonate aphids on three independent homozygous lines. For nymphs fed on PIN1-GFP-expressing plants, no effects on insect survival were observed but average insect weight and fecundity were significantly reduced. The aphid biomass was decreased by 30-35% compared to those reared on control plants. The effects of PIN1 on M. persicae were correlated with the decrease of the leucine aminopeptidase and total protease activities of whole insect extracts. Furthermore, an increase in polyphenoloxidase activity was observed in PIN1-GFP-expressing plants. These results revealed that the transgenic expression of PIN1 in tobacco enhanced tolerance against aphids. Key message This study suggests that entomopathogenic nematode symbiotic bacterium is another valuable resource of protease inhibitors which can be engineered into plants for insect pest management.

Elevated CO2 Reduces the Resistance and Tolerance of Tomato Plants to Helicoverpa armigera by Suppressing the JA Signaling Pathway

Both resistance and tolerance, which are two strategies that plants use to limit biotic stress, are affected by the abiotic environment including atmospheric CO2 levels. We tested the hypothesis that elevated CO2 would reduce resistance (i.e., the ability to prevent damage) but enhance tolerance (i.e., the ability to regrow and compensate for damage after the damage has occurred) of tomato plants to the cotton bollworm, Helicoverpa armigera. The results showed that elevated CO2 reduced resistance by decreasing the jasmonic acid (JA) level and activities of lipoxygenase, proteinase inhibitors, and polyphenol oxidase in wild-type (WT) plants infested with H. armigera. Consequently, the activities of total protease, trypsin-like enzymes, and weak and active alkaline trypsin-like enzymes increased in the midgut of H. armigera when fed on WT plants grown under elevated CO2. Unexpectedly, the tolerance of the WT to H. armigera (in terms of photosynthetic rate, activity of sucrose phosphate synthases, flower number, and plant biomass and height) was also reduced by elevated CO2. Under ambient CO2, the expression of resistance and tolerance to H. armigera was much greater in wild type than in spr2 (a JA-deficient genotype) plants, but elevated CO2 reduced these differences of the resistance and tolerance between WT and spr2 plants. The results suggest that the JA signaling pathway contributes to both plant resistance and tolerance to herbivorous insects and that by suppressing the JA signaling pathway, elevated CO2 will simultaneously reduce the resistance and tolerance of tomato plants.

Allometric relationships in feeding and digestion in the Chilean mytilids Mytilus chilensis (Hupé), Choromytilus chorus (Molina) and Aulacomya ater (Molina): A comparative study

As a first attempt to explain the functional basis of the co-occurrence of three sympatric species of native mussels frequently found at the cold-temperate coasts of Chile, we tested the existence of potential morpho-functional differences operating at the feeding and digestive levels of Mytilus chilensis, Choromytilus chorus and Aulacomya ater. Clearance rate (CR (l h–1)), gill area (GA: mm2), dry weight of labial palps (WLP: mg), gut contents (GC: mg), weight of digestive gland (WDG: mg), specific (mg of maltose or tyrosine mg prot−1h−1) and total (mg of maltose or tyrosine digestive gland−1h−1) amylase, cellulase, laminarinase, xylanase and protease activities were measured in different size individuals of the three species collected from Corral Bay (Chile). Hydrolytic activity of digestive extracts towards natural substrates elaborated from different macrophyte was also tested for the 3 species. No significant differences between species were found for mass-exponents (b) scaling CR (b=0.744) and GA (b=0.803). However, size-specific CR and GA were found to vary between species, ranking as follows: M. chilensis>C. chorus>A. ater. WLP fitted to an inter‐specific common function (b=0.77). Regarding digestive parameters, no significant inter‐specific differences were found between mass-exponents for GC (b=0.434), WDG (b=1.067) and total enzyme activities (common b ranged from 0.904 to 1.212), except total amylase which was significantly higher in A. ater (b=1.838). Specific activities of digestive enzymes were size-independent except A. ater amylase, which increased with body-size, and protease in the three species which decreased with size. For a common body-size, GC and carbohydrase activities were significantly higher in M. chilensis than in C. chorus and A. ater. Inter‐specific differences were found in the composition of carbohydrase pool: amylase was the prevalent enzyme in M. chilensis (up to 44.5% of total carbohydrase) whereas the three carbohydrases had a similar contribution in C. chorus and A. ater. Sugar release from vascular plants was significantly higher in C. chorus.

Developmental and moult cycle related biochemical changes in larvae of the spider crab, &lt;i&gt;Maja brachydactyla&lt;/i&gt; (Brachyura: Majidae)

Ontogenetic changes in dry weight, elemental composition (CHN), and digestive enzyme activities (total protease, amylase) were studied during the moulting cycle of all larval instars (zoea I-II, megalopa; ZI, ZII, M) of the spider crab Maja brachydactyla Balss, 1922 reared in the laboratory. A gradual but significant increase in biomass was observed throughout the zoeal instars, followed by steeper growth in the megalopa. Maximum values were reached in moult stage D 2 . Digestive enzyme activities also increased significantly throughout larval development. The ZI showed a continuous increase in total protease activity during its moult cycle, suggesting that newly hatched larvae have a gradually increasing need to take up proteins to provide amino acids, which are prime materials for growth and development, and possibly to use them as a metabolic energy source. The largest variations in digestive enzyme activites in relation to the moult cycle were observed in the megalopa instar. After an initial increase in postmoult, enzyme activities decreased to low levels similar to those measured in the zoeal instars at intermoult, followed by another increase to a maximum level in premoult. Metamorphosis involves great physiological and behavioural changes, which could explain the large variations in the megalopa moult cycle. We therefore suggest that the moult cycle stages should be taken into account when nutritional condition indices are used to estimate the physiological performance of decapod crustacean larvae.

Proteolytic profile in the larval midgut of Chilo suppressalis Walker (Lepidoptera: Crambidae)

AbstractChilo suppressalis is a key constraint on production of rice. The current research was conducted to study the types of digestive proteases in the larval midgut of C. suppressalis. It was found that activity of total digestive proteases increased from the first to the fifth larval instars, which showed different nutritional requirements. Four types of proteinases and two types of exopeptidase were identified so that their activities from the highest to the lowest activities is trypsin‐like, chymotrypsin‐like and elastase for proteinases, and amino and carboxypeptidases for exopeptidases. Meanwhile, just one type of cysteine protease, cathepsin D, was determined in the fourth and fifth instar larvae. The optimal pH for activity of total protease was found to be pH 9–10 and optimal temperature was observed to be 35–40°C, where there was the highest proteolytic activity. Some specific inhibitors of proteases including PMSF, TLCK, TPCK, DTT, E‐64, cystatin, phenanthroline and EDTA were used to confirm the types of proteases in the midgut of C. suppressalis.

Identification and biochemical characterization of 20S proteasome in wheat roots under salt stress

In order to gain a better understanding of proteases state in wheat roots under salt stress, the roots of wheat (Triticum aestivum L., H6756) exposed to 300 mM NaCl were investigated at different days after treatment (DAT). The results showed that the content of soluble proteins decreased continuously, but the activity of total proteases increased gradually, and five root endopeptidase isoenzymes (RE1-5) were detected by natural gradient polyacrylamide gel electrophoresis (GPAGE) with supplement of gelatin as a substrate. Among five REs, RE1 was not only detected in salt-treated roots, but also obviously possessed the higher activity, whereas its protein amount decreased gradually during the salt treatment process. The biochemical characters of RE1 were examined afterwards. The results showed that its molecular mass was about 740 kDa and its optimal temperature was about 40°C. But the optimal pH was 5 to substrate gelatin or 7–8 to substrate casein. Its activity was obviously inhibited by 10 mM PMSF and 20 μM MG115, and RE1 was further confirmed as 20S proteasome by Western blotting. The results in this study suggested that 20S proteasome of wheat roots might be an important protease accumulated in roots in response to salt stress.

Effects of dietary crude protein levels on development, antioxidant status, and total midgut protease activity of honey bee (Apis mellifera ligustica)

The effects of different levels of dietary crude protein on the development, antioxidant enzymatic activity, and total midgut protease activity of honey bees were investigated in the study. A total of 30 colonies of bees with sister queens were used in the test. Dietary treatments were pure rape pollen (Control) and pollen substitutes (PS) with crude protein (CP) levels at 15%, 20%, 25%, 30%, and 35% (designated as PS15, PS20, PS25, PS30, and PS35), respectively. We compared the effects of these diets on honey bees by measuring diet consumption, bee development (egg hatch, pupation success, and pupal weight), and the protein content of emergent adult bees, their antioxidant status and the activity of their midgut digestive proteases. Bees consumed significantly more (P < 0.001) natural pollen than any PS, and bees fed PS had similar diet consumption over the entire experimental period. However, the total protein intake was varied (P < 0.05). PS with a protein level about 30% was recognized as excellent quality diet for maximum body weight, highest protein content and antioxidant enzymatic activity, and PS with a protein rate about 35% exerted the greatest effect on increasing percentage of hatch and percentage of pupation. All these results indicate that PS appeared to be a valuable proteinaceous food approximated to the pollen, and 30∼35% of dietary protein level was optimal to maintain the colony development.

Related antipodes: a comparative study on digestive endopeptidases from Northern krill and Antarctic krill (Euphausiacea)

The Antarctic krill, Euphausia superba, and the Northern krill, Meganyctiphanes norvegica, are closely related species but occupy significantly different trophic and climatic environments. E. superba holds a key position as a phytoplankton grazer in the Southern Ocean. The omnivorous M. norvegica is an important member of plankton communities in the Northeast Atlantic. Both species expressed high proteolytic activities which were dominated by serine proteinases. In the stomachs of Antarctic krill, activities of total proteinase, trypsin, and chymotrypsin were significantly higher than in Northern krill. In the midgut glands, however, total proteinase and trypsin activities were similar in both species, but chymotrypsin activity was significantly higher in Antarctic krill. Moreover, Antarctic krill expressed four trypsin isoforms while only one isoform appeared in Northern krill. Chymotrypsin was present in either species as one single isoform. Antarctic krill adapted to the low and patchy distribution of food by elevated enzyme activities and the expression of trypsin isoforms with slightly different catalytic properties. Presumably, these enzymes facilitate in concerted action the efficient utilization of proteins from phytoplankton, the major food. Northern krill, in contrast, seems not to be equipped to face food limitation. It expresses a “simple” or “basic” set of digestive enzymes for utilizing abundant and easily digestible prey.

Comparative studies on the changes of total soluble proteins and protease activity in Georgian commercial peanuts contaminated by Aspergillus flavus

The presence of Aspergillus species is an indicator of storage conditions, which also suggests the possibility of several biochemical changes in grains. A comparative change in total soluble proteins and protease activity was determined in commercial peanut seeds collected from Georgia State. Protein contents of healthy peanuts, naturally contaminated peanuts and then artificially inoculated peanut seeds with A. flavus were estimated by Bradford method, and protease activity was also determined by using the Protease Detection Kits. Protein contents and the protease activity of the peanuts varied from sample to sample. The soluble protein content of seeds was significantly higher in healthy peanuts than in artificially inoculated or naturally infected peanuts with A. flavus. Protease activity was found to be higher in artificially inoculated seeds than in either naturally infected or healthy peanuts. Level of soluble proteins in buffer extracts of contaminated seeds decreased with incubation time, and protease activity increased with incubation time. These changes may be attributed to host response due to infection, contribution by A. flavus or due to biochemical alterations that occur naturally during the transition from endosperm to seedling during incubation period.