The effects of some storage conditions on viability of Lecanicillium lecanii conidia to whitefly (Homoptera: Trialeurodes vaporariorum)

A study on the survival of Lecanicillium lecanii conidia in storage at room temperature was carried out. Firstly, drying methods of conidia powder were compared. Vacuum-freeze drying (VFD) was more suitable for drying conidia as compared to vacuum drying (VD) at room temperature. Vacuum-freeze drying for 24-h resulted in a water content of 5.4%, and a viability, determined as germination of conidia in 2% glucose solution after16 h, was 90.3% and the infection in greenhouse whitefly, Trialeurodes vaporariorum was about 94.7% at a dose of 1×10⁸ conidia/mL. Secondly, the factors influencing viability of conidia stored at room temperature were evaluated in the laboratory. Temperature was the most critical factor influencing conidial storage stability, among the tested factors affecting survival of conidia stored at room temperature for 6 months. Both conidial germination and infection of hosts decreased with storage temperature increasing from 15 to 35°C, and at 35°C the survival of stored conidia for 6 months was near zero. The moisture content of the conidial powder was another major factor influencing viability of stored conidia at room temperature. Conidial powder dried to about 5% moisture content showed higher viability than non-dried conidial powder. For the carriers, clay and charcoal were more suitable for storage of L. lecanii conidia at room temperature. At a room temperature of 25°C, L. lecanii conidia which were dried to 5% water content and mixed with clay or charcoal could retain about 50% survival after 6 months' storage.     

Comparative survival rates of human-derived probiotic Lactobacillus paracasei and L. salivarius strains during heat treatment and spray drying

Spray drying of skim milk was evaluated as a means of preserving Lactobacillus paracasei NFBC 338 and Lactobacillus salivarius UCC 118, which are human-derived strains with probiotic potential. Our initial experiments revealed that NFBC 338 is considerably more heat resistant in 20% (wt/vol) skim milk than UCC 118 is; the comparable decimal reduction times were 11.1 and 1.1 min, respectively, at 59 degrees C. An air outlet temperature of 80 to 85 degrees C was optimal for spray drying; these conditions resulted in powders with moisture contents of 4.1 to 4.2% and viable counts of 3.2 x 10(9) CFU/g for NFBC 338 and 5.2 x 10(7) CFU/g for UCC 118. Thus, L. paracasei NFBC 338 survived better than L. salivarius UCC 118 during spray drying; similar results were obtained when we used confocal scanning laser microscopy and LIVE/DEAD BacLight viability staining. In addition, confocal scanning laser microscopy revealed that the probiotic lactobacilli were located primarily in the powder particles. Although both spray-dried cultures appeared to be stressed, as shown by increased sensitivity to NaCl, bacteriocin production by UCC 118 was not affected by the process, nor was the activity of the bacteriocin peptide. The level of survival of NFBC 338 remained constant at approximately 1 x 10(9) CFU/g during 2 months of powder storage at 4 degrees C, while a decline in the level of survival of approximately 1 log (from 7.2 x 10(7) to 9.5 x 10(6) CFU/g) was observed for UCC 118 stored under the same conditions. However, survival of both Lactobacillus strains during powder storage was inversely related to the storage temperature. Our data demonstrate that spray drying may be a cost-effective way to produce large quantities of some probiotic cultures.     

Intrinsic tolerance of Bifidobacterium species to heat and oxygen and survival following spray drying and storage

AIMS: This study examined the tolerance of various species of the genus Bifidobacterium to heat and oxygen and evaluated the survival of selected strains following spray drying and during storage. METHODS AND RESULTS: Nine Bifidobacterium species were considered to be relatively tolerant to both heat and oxygen and mostly segregated into two clusters within the 16S rDNA phylogenetic tree. Four species were tolerant to oxygen and 12 species were considered sensitive to oxygen and heat. Using a skimmed milk-based carrier good survival following spray drying and storage at 4 degrees C correlated with tolerance to heat and oxygen. Viability was inversely related to storage temperature and at 15 degrees C and 25 degrees C, a significant decline was observed for all species. The inclusion of gum acacia had no significant affect on survival or viability. However, using a fluidized-bed spray dryer viability was greatly improved. CONCLUSIONS: A group of closely related species tolerant to heat and oxygen had high survival following spray drying and maintained viability during prolonged storage at 4 degrees C. SIGNIFICANCE AND IMPACT OF THE STUDY: Spray drying is a suitable method for the production of skimmed milk powder enriched with high numbers of viable bifidobacteria.     

Formulation development of the biocontrol agent Bacillus subtilis strain CPA-8 by spray-drying

Aims: To prepare commercially acceptable formulations of Bacillus subtilis CPA‐8 by spray‐drying with long storage life and retained efficacy to control peach and nectarine brown rot caused by Monilinia spp. Methods and Results: CPA‐8 24‐h‐ and 72‐h‐old cultures were spray dried using 10% skimmed milk, 10% skimmed milk plus 10% MgSO₄, 10% MgSO₄ and 20% MgSO₄ as carriers/protectants. All carriers/protectants gave good percentages of powder recovery (28–38%) and moisture content (7–13%). CPA‐8 survival varied considerably among spray‐dried 24‐h‐ and 72‐h‐old cultures. Seventy‐two hours culture spray dried formulations showed the highest survival (28–32%) with final concentration products of 1·6–3·3 × 10⁹ CFU g^?1, while viability of 24‐h‐old formulations was lower than 1%. Spray‐dried 72‐h‐old formulations were selected to subsequent evaluation. Rehydration of cells with water provided a good recovery of CPA‐8 dried cells, similar to other complex rehydration media tested. Spray‐dried formulations stored at 4 ± 1 and 20 ± 1°C showed good shelf life during 6 months, and viability was maintained or slightly decreased by 0·2–0·3‐log. CPA‐8 formulations after 4‐ and 6 months storage were effective in controlling brown rot caused by Monilinia spp. on nectarines and peaches resulting in a 90–100% reduction in disease incidence. Conclusions: Stable and effective formulations of biocontrol agent B. subtilis CPA‐8 could be obtained by spray‐drying. Significance and Impact of the Study: New shelf‐stable and effective formulations of a biocontrol agent have been obtained by spray‐drying to control brown rot on peach.     

香芋粉喷雾干燥工艺研究

我国香芋资源极为丰富,但开发程度远不及山药、甘薯、马铃薯等,所以对香芋加工工艺的研究对其潜在价值的开发具有重要意义。通过单因素试验考察了烫漂温度、烫漂时间、进风温度、进料速度及料液浓度对香芋粉喷雾干燥的影响。试验确定了香芋粉喷雾干燥的最佳工艺为:55℃烫漂10 min、进风温度为180℃、进料速度50 mL/min、料液浓度为0.1 g/mL。     

Spray drying of the dehalogenating bacterium Rhodococcus sp.

The bacteria Rhodococcus sp. and Xanthobacter autotrophicus have the ability to dehalogenate a broad range of halogenated hydrocarbons. The applicability of spray drying to the preservation of the microorganisms and the intracellular enzyme halidohydrolase (E.C.3.8.1.1) was examined. K₂SO₄, MgSO₄, glutamate and sucrose were added as stabilizers and carriers. Spray drying was carried out at inlet air temperatures of 100–120 °C and outlet air temperatures of 65–72 °C. Best results were obtained by the addition of 5% K₂SO₄ and at 107 °C air inlet temperature. Dried preparations of Rhodococcus sp. exhibited a crystalline consistency and a 95% recovery of cellular activity. After storage at 4 °C for six months the enzyme preparation showed no loss in activity. Spray dried preparations of Xanthobacter autotrophicus showed only a 4% recovery of cellular activity.List of Symbols MSG Monosodiumglutamate - RC % Recovery of stabilizer and biomass - RCA % Recovery of cellular activity ([U/g biomass after the spraydrying]/[U/g biomass of untreated cells]) 100 - RCB % Recovery of biomass - SR % Survival rate - T ₁ °C Inlet air temperature - T ₂ °C Exit air temperature - W % Water content - Y.Akt % Yield in enzyme activity This work was supported by the Jubiläumsfond der Oesterreichischen Nationalbank, Projekt No. 4499.     

Comparative Survival Rates of Human-Derived Probiotic Lactobacillus paracasei and L. salivarius Strains during Heat Treatment and Spray Drying

Spray drying of skim milk was evaluated as a means of preserving Lactobacillus paracasei NFBC 338 and Lactobacillus salivarius UCC 118, which are human-derived strains with probiotic potential. Our initial experiments revealed that NFBC 338 is considerably more heat resistant in 20% (wt/vol) skim milk than UCC 118 is; the comparable decimal reduction times were 11.1 and 1.1 min, respectively, at 59°C. An air outlet temperature of 80 to 85°C was optimal for spray drying; these conditions resulted in powders with moisture contents of 4.1 to 4.2% and viable counts of 3.2 × 10⁹ CFU/g for NFBC 338 and 5.2 × 10⁷ CFU/g for UCC 118. Thus, L. paracasei NFBC 338 survived better than L. salivarius UCC 118 during spray drying; similar results were obtained when we used confocal scanning laser microscopy and LIVE/DEAD BacLight viability staining. In addition, confocal scanning laser microscopy revealed that the probiotic lactobacilli were located primarily in the powder particles. Although both spray-dried cultures appeared to be stressed, as shown by increased sensitivity to NaCl, bacteriocin production by UCC 118 was not affected by the process, nor was the activity of the bacteriocin peptide. The level of survival of NFBC 338 remained constant at ~1 × 10⁹ CFU/g during 2 months of powder storage at 4°C, while a decline in the level of survival of approximately 1 log (from 7.2 × 10⁷ to 9.5 × 10⁶ CFU/g) was observed for UCC 118 stored under the same conditions. However, survival of both Lactobacillus strains during powder storage was inversely related to the storage temperature. Our data demonstrate that spray drying may be a cost-effective way to produce large quantities of some probiotic cultures.     

Species- and clone-specific responses to environmental stimuli in the cladocerans Bosmina cornuta and B. pellucida - a comparison with Daphnia

We studied the variation of small-scale swimming behaviour in eight Bosmina cornuta and ten B. pellucida clones in response to key environmental factors to test whether swimming behaviour and genotypes are linked in non-Daphnia cladocerans. We quantified (1) the short-term responses to changes in temperature, light intensity and pH, (2) the response to long-term temperature acclimation, and (3) the pH-related survival rates. Vertical swimming activity S was quantified in cuvette experiments as crossings of a line at 2 cm height per individual an hour. S differed significantly among species and conspecific clones. At any temperature, light intensity and pH tested, B. cornuta (clone variation: 40-58 crossings/ind.× h) showed a higher vertical swimming activity than B. pellucida (clone variation: 25-48 crossings/ind.× h). A short-term change of water temperature (range tested: 10-25°C) only affected S of B. cornuta, whereas that of B. pellucida remained unaltered. In contrast, S increased with rising temperature following long-term temperature acclimation (range tested: 10-20°C) in both species. Swimming activity was inversely related to the light intensity (range tested: 60-60,000 lux), but decrease of activity was stronger in B. pellucida (44 → 12 crossings/ind × h) than in B. cornuta (50 → 40 crossings/ind.× h). Short-term changes of pH (range tested: 4-6) did not influence swimming activity in any species, although a prolonged exposure (24 h) to pH 4 was lethal. Thus, Bosmina showed behavioural responses which permit to distinguish between the species and which are related to their seasonal succession and distribution pattern.     

Optimization of spray-drying conditions for the large-scale preparation of Bacillus thuringiensis var. israelensis after downstream processing

Reduction of water activity in the formulations of mosquito biocontrol agent, Bacillus thuringiensis var. israelensis is very important for long term and successful storage. A protocol for spray drying of B. thuringiensis var. israelensis was developed through optimizing parameters such as inlet temperature and atomization type. A indigenous isolate of B. thuringiensis var. israelensis (VCRC B-17) was dried by freeze and spray drying methods and the moisture content and mosquito larvicidal activity of materials produced by the two methods were compared. The larvicidal activity was checked against early fourth instars Aedes aegypti larvae. Results showed that the freeze-dried powders retained the larvicidal activity fairly well. The spray-dried powder moderately lost its larvicidal activity at different inlet temperatures. Between the two types of atomization, centrifugal atomization retained more activity than the nozzle type atomization. Optimum inlet temperature for both centrifugal and nozzle atomization was 160 degrees C. Keeping the outlet temperature constant at 70 degrees C the moisture contents for the spray-dried powders through centrifugal atomization and freeze-dried powders were 10.23% and 11.80%, respectively. The LC(50) values for the spray-dried and freeze-dried powders were 17.42 and 16.18 ng/mL, respectively. Spore count of materials before drying was 3 x 10(10) cfu/mL and after spray drying through nozzle and centrifugal atomization at inlet and outlet temperature of 160 degrees C/70 degrees C were 2.6 x 10(9) and 5.0 x 10(9) cfu/mL, respectively.     

Experimental determination of viability loss of Penicillium bilaiae conidia during convective air-drying

A study was conducted on the drying of Penicillium bilaiae, a fungal micro-organism used to promote soil-bound phosphorous uptake in several crop species, such as wheat, canola and pulse crops. A wet pellet formed from a mixture of the inoculant and a starch-based carrier was air-dried to the appropriate water activity to extend the shelf-life of the viable fungal conidia. Convective air-drying was examined as a low-energy alternative to the more expensive freeze-drying technology that is currently in use. Experiments were conducted to measure the loss of conidia viability during drying in a fixed-bed, thin-layer convective dryer. The dryer air inlet temperature and relative humidity were controlled in experiments to determine the effect of thermal and dessicative stresses on conidial viability. The measured survivor fraction was determined to be dependent on solids temperature, moisture content and drying rate. Thermal stresses became significant for process temperatures above 30°C, while the survivor fraction fell sharply below a dry basis moisture ratio of 30%. Slower drying kinetics associated with high inlet air relative humidity were found to significantly improve the recovery of viable conidia. By minimising environmental stresses, survivor fractions of up to 75% could be achieved, but this result fell dramatically with the introduction of more severe conditions. A general linear statistical model is used to quantify experimental error and the significance level of each factor.     

Statistical modeling of protein spray drying at the lab scale

The objective of this study was to examine the effects of formulation and process variables on particle size and other characteristics of a spray-dried model protein, bovine serum albumin (BSA), using a partial factorial design for experiments. Formulation variables tested include concentration and zinc:protein complexation ratio. Process variables explored were inlet temperature, liquid feed rate, drying air flow rate, and atomizing nitrogen pressure on a lab-scale spray dryer. Statistical data analysis was used to determine F ratios for each of the inputs, which provided a means of ranking the importance of variables relative to one another for each powder characteristic of interest. It was found that protein concentration and atomizing nitrogen pressure had the greatest effects on the particle size of the protein powder. For determining product yield, results showed that protein concentration was the critical variable. Finally, the outlet temperature was mostly influenced by inlet temperature and liquid feed rate. Mathematical models based on these input-output relationships were constructed; these models provide insight into some of the controllable variables of the spray-drying process. Published: March 20, 2002     

Spray-drying performance of a bench-top spray dryer for protein aerosol powder preparation

The objective of this work was to improve a bench-top spray dryer's efficiency in both production recovery and throughput for preparing protein aerosol powders. A Büchi mini-spray dryer was used to prepare the powders of recombinant humanized anti-IgE antibody. The resulting powder's physical properties such as particle size, residual moisture, and morphology, along with its recovery and production rate was the basis of this development work. Mass balance suggests that approximately 10-20% of powder was lost in the exhaust air, consisting primarily of particles less than 2 micrometer. Also, significant loss (20-30%) occurred in the cyclone. Attempts were made to improve product recovery in the receiving vessel using dual-cyclone configurations, different cyclone designs, cyclones with anti-static treatment, and different receiver designs. System modifications such as replacing the original bag-filter unit with a vacuum system effectively reduced drying air flow resistance, allowing the protein to be dried at a lower inlet air temperature and the production scale to be increased. We concluded that the modified spray-drying system is advantageous over the original bench-top spray dryer. This improvement will be beneficial to early-stage research and development involving high-valued protein powders.     

Spraydrying of yeast-lytic enzymes from Arthrobacter sp.

Summary Spray drying processes are widely used for large scale preservation of biological materials e.g milk, whey, yeast, egg white, etc. Nevertheless, there is an increasing tendency for drying of catalytic active proteins. The paper presents results from experiments which were carried out to test the applicability of spray drying for the preservation of several bacterial enzymes used for the lysis of yeast cell-walls (-1,3-glucanase, -mannanase, chitinase and amylase) and formed by an Arthrobacter species. Enzyme solutions were obtained by concentrating the cell free cultivation liquid by cross-flow ultrafiltration. Due to the low protein content of the liquid preparation, several substances were added as stabilizers or carriers (eg. sucrose, KCl, MgSO₄) and the effect was studied in further experiments. Spray drying was carried out at inlet air temperatures of 110 – 120°C and outlet air temperatures of 65 – 72°C. Best results were obtained by the addition of 10% KCl, the preparation having a crystalline consistency and a 60% yield in activity.     

Effect of spray dryer processing parameters on the insecticidal activity of two encapsulated formulations of baculovirus

The effect of spray dryer processing parameters on the product yield and insecticidal activity of baculovirus was evaluated. Spray-dried samples of a granulovirus (GV) from Pieris rapae (L.) and a multiple nucleopolyhedrovirus (MNPV) from Anagrapha falcifera (Kirby) were prepared using two dryer-atomiser configurations (rotary atomiser and two-fluid spray atomiser), four drying temperatures (50–100°C outlet temperatures) and two encapsulating formulations (lignin and methacrylic acid polymer). The samples were evaluated based on yield and insecticidal activity under laboratory conditions. The two atomising configurations produced similar outlet temperatures for dryer stock feed rates of 4.12 and 20 ml/min when processed using increasing inlet temperatures. The atomiser selection significantly affected the physical properties like the product yield; the microparticles produced with a two-fluid spray atomiser had lower product yields (57.8 ± 18.80% – 74.6 ± 4.26%) when compared with paired samples produced with a rotary-disc atomiser (58.1 ± 7.13% – 82.6 ± 3.12%). Spray drying reduced insecticidal activity of the GV but did not significantly reduce insecticidal activity of the MNPV when compared with samples that were not dried. Among dried samples, the spray dryer processing parameters (atomiser, drying temperatures and formulation) had minimal effect on the insecticidal activity of either baculovirus. The versatility of spray drying for processing baculoviruses was demonstrated by identifying parameters that improve process yield while having minimal impact on insecticidal activity.     

Synthesis of (R)-2-trimethylsilyl-2-hydroxyl-propionitrile catalyzed by (R)-hydroxynitrile lyase from Prunus japonica seed meal

Synthesis of (R)-2-trimethylsilyl-2-hydroxyl-propionitrile via asymmetric transcyanation of acetyltrimethylsilane with acetone cyanohydrin in an aqueous/organic biphasic system catalyzed by (R)-hydroxynitrile lyase from Prunus japonica seed meal was successfully carried out for the first time. The optimal volume ratio of aqueous to organic phase, buffer pH value and reaction temperature were 15% (v/v), 5.0 and 30°C, respectively, under which both substrate conversion and product enantiomeric excess (ee) were 99%. Silicon atom in the substrate showed great effect on the reaction. Acetyltrimethylsilane was a much better substrate for (R)-hydroxynitrile lyase from Prunus japonica than its carbon analogue.     

The detection of Cryptosporidium parvum and Escherichia coli O157 in UK bivalve shellfish

Optimised immunomagnetic separation methods to detect Cryptosporidium parvum and Escherichia coli O157 in UK shellfish are described. Whole tissue homogenates gave the best recoveries for C. parvum oocysts compared with gill or haemolymph extracts. The sensitivity of recovery from spiked samples was comparable to that achieved when processing water and varied from 12–34% in mussels, 48–69.5% in oysters and 30–65% in scallops. Maximum recovery of E. coli O157 was achieved by enriching in buffered peptone water supplemented with vancomycin at 42 °C. Increasing enrichment temperatures from 37 to 42 °C gave a significant increase in target number recovery. Implementation of these methods into monitoring programmes and end-product testing will enable shellfish producers to better assess product safety.     

Thermally reversible inactivation of Taq polymerase in an organic solvent for application in hot start PCR

In the past, Taq polymerase was reversibly inactivated by modification with a dicarboxylic acid anhydride in aqueous media, to enable ‘hot start PCR’. However, there are various constraints in using such a method including temperature and concentration. Here we describe an alternative method whereby Taq polymerase may be reversibly inactivated following incubation with an excess of citraconic anhydride at elevated temperatures, in an anhydrous non-protic organic solvent – tert-butyl methyl ether – by first drying the enzyme with a salt or carbohydrate excipient to form an amorphous powder. Reactivation of the enzyme is due to the instability of the chemical modification at low pH following a short incubation in a suitable buffer.     

Comparison of Spray Freeze Drying and the Solvent Evaporation Method for Preparing Solid Dispersions of Baicalein with Pluronic F68 to Improve Dissolution and Oral Bioavailability

The objective of this study was to prepare solid dispersions consisting of baicalein and a carrier with a low glass transition/melting point (Pluronic F68) by spray freeze drying (SFD). We compared these powders to those produced from the conventional solvent evaporation method. In the SFD process, a feeding solution was atomized above the surface of liquid nitrogen following lyophilization, which resulted in instantaneously frozen microparticles. However, solid dispersions prepared by the solvent evaporation method formed a sticky layer on the glass flask with crystalline baicalein separated out from the carrier. The powder samples were characterized by scanning electron microscopy (SEM), powder X-ray diffraction (PXRD), surface area measurement, differential scanning calorimetry, and Fourier transform infrared spectrometry. SEM and PXRD results suggested that the majority of baicalein in the SFD-processed solid dispersion was in the amorphous state, which has a higher specific surface area than pure baicalein. However, the majority of baicalein was recrystallized in the solid dispersion at the same composition prepared by the solvent evaporation method, which showed a similar dissolution rate to the physical mixture. SFD product was physically and chemically stable after being stored at 40°C with low humidity for 6 months. After enzyme hydrolysis, baicalein in the SFD product displayed a significantly shorter T _max and higher C _max than pure baicalein after oral dosing. The relative bioavailability of the SFD product versus pure baicalein determined by comparing the AUC_0–12 was 233%, which demonstrated the significantly improved oral bioavailability of baicalein produced by the SFD technique.     

Mathematical modeling and study of mass transfer parameters in supercritical fluid extraction of fatty acids from Trout powder

Mathematical modeling of fatty acids (FAs) extraction from Trout powder by supercritical carbon dioxide was performed in the present work. Trout powder with its low cost contains high amount of essential FAs and it is commonly available as a proper source of FAs. The effect of process parameters, such as pressure (25, 28, 31, 34 and 37 MPa) and temperature (310, 318 and 326 K) of extraction and void fraction of the bed (0.25, 0.35 and 0.45, v/v) on the yield of FAs extraction was examined in a series of experiments conducted in a laboratory scale apparatus. The results indicated a significant increase of extraction yield with an increase of pressure from 25 to 34 MPa, but working at the higher pressure (37 MPa) caused reduction of the extract. Increasing the temperature higher than 318 K revealed significant reduction of the FAs yield and increasing the bed void fraction from 0.25 to 0.45 showed enhancement of the extraction.

The mathematical model was developed considering diffusion-controlled regime in the particle and film mass transfer resistance around the particle with axial dispersion of the bulk phase at dynamic conditions. Henry law was used to describe the equilibrium state of solid and fluid phases. The proposed mass balance equations were numerically solved using implicit finite difference method and the model parameters were correlated using the experimental results of the outlet FAs concentration in the oil extracted at dynamic conditions. Well-known Nelder–Mead method was applied to estimate the four parameters of the model, namely, mass transfer coefficient (k_f), axial dispersion coefficient (D_ax) in the bulk phase, effective diffusivity (D_eff) into the pores and Henry coefficient (H). In the range of studied conditions, the higher extraction efficiency with higher pressure resulted lower correlated H, although the temperature increasing which showed a retrograde phenomena in the FAs yield, revealed H passing though a minimum.     

Itaconic acid production from xylose in repeated-batch and continuous bioreactors

Summary Itaconic acid production from xylose by immobilized Aspergillus terreus TKK 200-5-2 mycelia was optimized both in repeated shake-flask fermentations and in continuous column bioreactors using statistical experimental design and empirical modelling. Using continuous 9-1 scale air-lift bioreactors, a pH of 2.5, aeration rate of 0.6 v/v per minute and residence time of 160 h gave the highest itaconic acid concentration. In air-lift bioreactors a cubic carrier size of 0.5 cm gave a 3.3-fold higher product concentration than 1-cm cubes. Packed-bed column reactors had a higher production rate than air-lift reactors. Offprint requests to: H. Kautola