ПРИКЛАДНАЯ БИОХИМИЯ И МИКРОБИОЛОГИЯ, 2013, том 49, № 6, с. 600-605
UDC 631.445.41
DETERMINATION OF HYDRATION PROPERTIES AND THERMAL BEHAVIOR OF Paecilomyces variotii BY DIFFERENTIAL SCANNING
CALORIMETRY
© 2013 R. S. Canel*, **, V. Ludemann*, O. De la Osa* and J. R. Wagner*, **
*National University of Quilmes, Buenos Aires, 1876Argentina **National Scientific and Research Council, Buenos Aires, 1033 Argentina e-mail: rcanel@unq.edu.ar Received Novembar 20, 2012
Due to the structure and the composition of Paecilomyces variotii, the mycelia of this fungus could have potential applications as ingredients in wettable foods. For this use, drying could be employed, justifying the study of thermal behavior of P. variotii. The objectives of this work were to perform a study of thermal behavior of P. variotii isolates, to evaluate the hydration properties of these mycelia and to analyze the effect of different technological parameters on the latter properties. Wet cultures exhibited a wide endothermic transition, with mean values of peak temperature of 61°C and denaturation enthalpy of 4 J/g dry matter. Initial (50°C) and final (80°C) temperatures of the endothermic transition were used to dry the mycelia. Freeze-drying was also assayed. For all dried mycelia, a decrease in denaturation enthalpy between 40 and 50% was observed for drying at 50°C and freeze-drying, and a drastic decrease of almost 100% for drying at 80°C. According to the hydration properties, wet mycelia exhibited water holding capacity (WHC) value of 45 g water/g dry matter. Significant differences among dried mycelia, resulting WHC values in order: 50°C > freeze-dried > 80°C (p < 0.05) were revealed for each P. variotii strain. Fungi obtained by drying at 50°C and by freeze-drying, showed a rapid water absorption (ti/2 < 0.1 min). Ionic strength, pH and particle size of dried mycelia influenced the hydration properties.
DOI: 10.7868/S0555109913060044
The genus Paecilomyces was split from Penicillium by Bainier in 1907 [1] on the basis of their differences in phialide shape and conidial color. Paecilomyces species are important as soil fungi and insect pathogens. Only two species are commonly isolated from foods: Paecilomyces variotii and Paecilomyces lilacinus [2]. P. variotii has not been reported as a mycotoxin producer [2], and is used in the food industry as a producer of enzymes. This fungus produces a thermostable glucoamylase [3]. a-amylases have the widest range of industrial applications [4, 5] including brewing, baking, textile and detergent production [6, 7]. P. variotii also produces tannase [8], an enzyme that cleaves ester linkages in hydrolyzable tannins.
Single cell protein (SCP) is a term coined in the 1960's to embrace microbial biomass products of fermentation [9]. P. variotii has been used for the production of microbial protein because of its excellent ability to grow in a variety of highly-polluting industrial effluents, such as molasses, hydrolyzed wood, spent sulfite liquor and vinasse [10—13]. P. variotii was the first fungus used in an industrial process for the production of microbial protein for animal feed, known as "Pekilo" process [10].
The successful adoption and use of microbial biomass depend on the properties of the fungus, which determine its usefulness as a functional food ingredi-
ent. Functional properties are defined as the physico-chemical properties that contribute with the rheologi-cal parameters and stability and sensory features of a food product. These properties can be classified as hydration, structuring, surface and organoleptic properties [14]. The water holding capacity (WHC) and the water absorption capacity (WAC) expresses the hydration properties. WHC refers to the ability of a hydrated material to retain water against the action of an external force of centrifugal gravity or compression, whereas WAC refers to the ability of a material to absorb water in its structure spontaneously when it comes in contact through a surface that remains wet. Both WHC and WAC are relevant for each food system [15].
The possible inclusion of Pekilo biomass has been studied in different food systems, such as sausages and meat balls [16]. P. variotii has been found to increase the hardness of sausages up to 30% substitution level, probably owing to its good water imbibing and gelling properties reported earlier [16]. Pekilo has also been studied in doughs, particularly in white wheat bread and soft and hard rye bread. This biomass has also been observed to increase the water binding of the dough from about 62% to approximately 72% when the flour substitution level is increased from 0 to 8%. The mild taste of Pekilo powder is distinctly found in bread, even at the lowest substitution levels [17].
In our previous work [18], we isolated and identified filamentous fungi from several sources. Non-toxic fungal strains, characterized by the Artemia salina bio-assay [19], were used to study their composition (protein, total dietary fiber and P-glucan) and hydration properties (WAC and WHC). We found that these mycelia give the following composition: protein content ranged from 30 to 47% of dry weight, total dietary fiber content changed from 16 to 53% of dry weight and P-glucans content varied from 1 to 26% of dry weight. The highest values of these parameters were found in P. variotii. According to hydration properties, this fungus showed the highest values for WHC and WAC, with a rapid water absorption rate (mycelia dried at 50°C) [18]. These results suggest the potential application of this fungus as ingredient in wettable foods. For this use, P. variotii may require drying, which justifies the study of its thermal behavior.
Then, the objectives of this work were to perform a study of thermal behavior of P. variotii isolates, to evaluate the hydration properties of these mycelia and to analyze the effect of different technological parameters on the latter properties.
MATERIALS AND METHODS
Fungal strains and culture conditions. Four strains of P. variotii isolated from pepper were used. The isolates were identified according to Pitt and Hocking [2]. The cultures were inoculated by 1 x 105 conidia/mL and grown in liquid media containing (g/L): yeast extract — 20 and sucrose — 40 by shaking at 135 rpm and 25 °C for 7 days. Mycelia samples were collected by filtration through Whatman N° 1 filter paper (USA) under vacuum and washed twice with distilled water.
Drying process. Wet mycelia of P. variotii were fractionated to be subjected to different drying conditions: a) drying in stove with air circulation at 50°C and 80°C until constant weight (moisture content <0.5%), and b) freeze-drying. For this latter process, samples were frozen at —80°C for 5 days. Freeze-drying was performed using HETO FD4 (Denmark) coupled to a Rotary Vane Vacuum Pump RZ5, for 30 h.
Protein, total dietary fiber and p-glucans determination. Dry mycelia were analyzed to determine protein [20], total dietary fiber (TDF) [21, 22] and P-glu-can. The latter was analyzed by the Megazyme TM commercial kit instruction (Ireland).
Milling and fractionation. Dried mycelia were ground in an FW100 Model Chincan miller (China) and fractionated to obtain different particle sizes (>850 ^m, 500-850 ^m and <500 ^m) using IRAM 1501/76 ASTM-E-11-81 sieves No. 35 and No. 20 (Argentina)
Thermal behavior. The thermal behavior of wet and dried mycelia of P. variotii was studied by modulated differential scanning calorimetry (MDSC) using a
Q200 calorimeter (TA Instrument, USA, amplitude: +/- 1°C, frequency: 60 sec). The samples were brought to 80% of water either by partial dehydration in the case of wet mycelia by Whatman No. 1 paper or by addition of water in the case of dry mycelia. Samples (12-15 mg) were hermetically sealed in standard aluminum pans and heated from 10 to 90°C at a heating rate of 5°C/ min. An empty pan was used as reference. Enthalpies and peak temperatures (Tp) of en-dotherms were obtained from thermograms. Enthalpies (H) were expressed as J/g dry matter by perforating the pans and heating overnight at 105°C [23]. All assays were performed in duplicate.
Hydration properties. WAC was determined on dried mycelia. Water absorption kinetics was followed using the Baumann equipment [24], which consists of a funnel connected to a horizontal capillary. A 50 mg sample was dusted on a wetted filter paper which was fastened to a glass filter placed on top of the funnel filled with water. The apparatus was kept at 20°C. The amount of water uptaken by the sample at equilibrium was read in the graduated capillary. The maximum amount of water absorbed and the time to reach WAC were determined [25]. The half time (t1/2) was defined as the time of the samples to absorb half of the maximum amount of water.
WHC was determined on both wet and dried mycelia of P. variotii. For WHC determination on wet mycelia, a full pellet was dispersed in distilled water and then shaken (for 2 h at 25°C and 100 rpm) and centrifuged (for 20 min at 15°C and 15000 x g. The pellet was dried in a stove with air circulation at 50°C until constant weight and finally weighed. For WHC determination of dry mycelia, 100 mg of sample was stirred with 9 mL of distilled water in an orbital shaker at 100 rpm for 2 h and then centrifuged for 30 min at 800 x g. The supernatant decanted and the pellet finally weighed. WHC and WAC were expressed as g of wa-ter/g of dry matter. Both determinations were performed using distilled water or 0.01 M phosphate buffer (pH 5.0 or 7.0) with different concentrations of NaCl (0, 1, 2 and 3%). Besides, hot water (80°C) was used to determine WHC. To this end, 100 mg of sample was stirred with 1 mL of distilled water at room temperature, in an orbital shaker for 1 min and then 8 mL of hot distilled water (80°C) was added. In the second assay, 100 mg of sample was stirred with 9 mL of hot distilled water (80°C). Both experiments were performed as prev
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