Freeze-dried Material Research Articles

Fruit Quality Changes of Salak “Pondoh” Fruits (Salacca zalacca (Gaertn.) Voss) during Maturation and Ripening

Salak is a very important fruit product in Indonesia that has been cultivated throughout Indonesia and has been exported to several countries. The study to determine biochemical and physical as well as physiological changes during fruit maturation and ripening was applied to two superior salak cultivars, “pondoh hitam” and “pondoh super” originally from Sleman, Yogyakarta of Indonesia. Fresh salak fruits of the cultivars “pondoh hitam” and “pondoh super” at three different ripening stages were used, i.e. stage 4, stage 5 and stage 5.5 (4, 5 and 5.5 months after pollination, respectively) for “pondoh hitam” and stage 4, stage 5 and stage 6 (4, 5 and 6 months after pollination, respectively) for “pondoh super”.<strong> </strong>Immediately after air transport from Indonesia to Germany, determination of fruit colour and texture as well as biochemical analyses were carried out in Berlin. Freeze-dried sample material was used for the determination of minerals, mono- and disaccharides, pectic substances and dietary fibre. Results of the study showed that increase in fruit size and weight as well as changes in peel and pulp colour occurred during maturation and ripening of salak fruits. Different patterns of peel and pulp colour changes were found in “pondoh super” and “pondoh hitam” during ripening. Physiological processes in “pondoh super” occurred to at a later stage but then accelerated faster than “pondoh hitam” in term of changes of mono- and disaccharides, resulting in a poorer marketability and shorter shelf life. In respect to the changes of sugar/acid ratio, there was a faster ripening process in “pondoh super” than in “pondoh hitam”. “Pondoh super” possessed higher content of polysaccharides and lignin, however, the ripening process accelerated earlier in comparison to “pondoh hitam”. Alterations in cell wall and middle lamella structure did not correlated with the physical non destructive texture measurement during ripening of salak.

Analysis of Flower Cuticular Waxes and Cutin Monomers

Here we describe procedures for the flower cuticular waxes extraction, modification and subsequent qualitative and quantitative analysis by gas-chromotography-mass spectrometry (GC-MS) and gas-chromotography with flame ionization detector (GC-FID), accordingly. To characterize flower cutin monomers two experimental setup are described: (i) analysis of enzymatically isolated cuticles in order to determine the relative proportions of cutin monomers; (ii) analysis of freeze-dried material for quantitative estimation of the cutin content. This report is an adaptation of the earlier published protocols developed for the chemical analysis of the cuticles in vegetative organs (Leide et al., 2007).

Biopolymer-based degradable nanofibres from renewable resources produced by freeze-drying

We describe a new biopolymer-based nanofibrous material possibly suitable for tissue engineering prepared by an environment-friendly organic solvent-free method. Glycogen, a biodegradable hyperbranched D-glucose polymer, comes from renewable resources and is normally present in man. It forms nanofibres by simple freeze-drying from aqueous solutions with concentration less than 0.5%. However, the architecture of the freeze-dried material depends on the starting biopolymer concentration within the tested range 0.1–5 wt%; in particular higher concentrations produce porous sponge-like structures with communicating pores. Because of the solubility of glycogen in water, nanofibres were modified by solvent-free grafting biodegradable poly(ethyl cyanoacrylate) from vapor phase. Exposing glycogen nanofibres to vapors of ethyl cyanoacrylate only slightly changed the material architecture while producing a water-insoluble biodegradable material with glycogen-to-poly(ethyl cyanoacrylate) ratio depending on the polymerization time. The material was proven to be hydrolytically degradable over the course of several months.

Swell Drying: Coupling Instant Controlled Pressure Drop DIC to Standard Convection Drying Processes to Intensify Transfer Phenomena and Improve Quality—An Overview

Increasing globalization is driving agricultural production in developing countries toward better recovery levels and high additional value. A great opportunity is thus offered to emerging economies. By adopting appropriate and improved methods that allow better preservation, transport, and storage and that conform to the rigorous requirements and standards of organoleptic and nutritional quality as well as stringent hygienic criteria, emerging economies can derive significant benefits. Drying of biological materials usually results in shrinkage. The impact of shrinkage is often highly prejudicial for both the function and use of these materials. On the one hand, shrinkage results in a sharp decrease in water diffusivity through the porous structure (for dehydration and rehydration processes). On the other hand, the compact structure of dried biological material is usually unsuitable in terms of organoleptic quality and is inadequate for grinding. Thus, it is becoming very important to include retexturing and structure-expanding processes in standard drying processes. To this end, the instant controlled pressure drop (DIC) technology has been proposed and successfully tested as a texturing process for partially dried materials, which should normally intensify the whole operation. The quality of the final product as well as the performance of the drying process can also be dramatically improved. Finally, for a large category of dried fruits and vegetables, and even freeze-dried materials, microbiological contamination and the presence of insects and larvae are serious problems. DIC technology can be used as an ultra-high-temperature treatment for a controlled decontamination. Such dried, expanded, and completely decontaminated products can be used as highly nutritional snacks. They can easily be ground to produce expanded granulated powders with excellent sensory, nutritional, functional, and textural properties.

Beef, chicken and lamb fatty acid analysis — a simplified direct bimethylation procedure using freeze-dried material

When fractionation of meat lipids is not required, procedures such as saponification can be used to extract total fatty acids, reducing reliance on toxic organic compounds. However, saponification of muscle fatty acids is laborious, and requires extended heating times, and a second methylation step to convert the extracted fatty acids to fatty acid methyl esters prior to gas chromatography. Therefore the development of a more rapid direct methylation procedure would be of merit. The use of freeze-dried material for analysis is common and allows for greater homogenisation of the sample. The present study investigated the potential of using freeze-dried muscle samples and a direct bimethylation to analyse total fatty acids of meat (beef, chicken and lamb) in comparison with a saponification procedure followed by bimethylation. Both methods compared favourably for all major fatty acids measured. There was a minor difference in relation to the C18:1 trans 10 isomer with a greater (P<0.05) recovery with saponification. However, numerically the difference was small and likely as a result of approaching the limits of isomer identification by single column gas chromatography. Differences (P<0.001) between species were found for all fatty acids measured with no interaction effects. The described technique offers a simplified, quick and reliable alternative to saponification to analyse total fatty acids from muscle samples.

Reinforced absorbent material: a cellulosic composite of TEMPO-oxidized MFC and CTMP fibres

This research aims to develop new materials based on renewable resources that can fulfill the functions necessary in the absorption core of a disposable diaper. Absorbent foam was recently produced from softwood kraft pulp by TEMPO oxidation, disintegration and freeze drying. In this study, the TEMPO-oxidized MFC was mixed with pulp fibres, thus forming a cellulosic composite, in an attempt to improve the mechanical stability of the freeze-dried absorbent material. The fibres were added in different amounts and the freeze-dried materials were evaluated for their absorption and retention properties. The results of this study suggest that the composite material has a better mechanical stability than the absorbent foam without fibres. It was shown that using spruce CTMP fibres in the composite resulted in better absorption and retention capacities than in a composite with softwood kraft pulp fibres. The higher stiffness of the CTMP fibres is a probable explanation for this difference. For the composite material with CTMP fibres, liquid porosimetry showed that pore size distribution was more or less retained when put under load. Furthermore, it was seen that the retention properties reached a maximum around 85 % CTMP fibres and 15 % TEMPO-oxidized MFC. In the centrifuge retention test, the retention of the TEMPO-oxidized MFC in the composite material reached about the same capacity as conventional superabsorbent polymers.

Certification of steroid carbon isotope ratios in a freeze‐dried human urine reference material

An accurate method for the measurement of carbon isotope ratios of steroids in human urine has been developed at the National Measurement Institute, Australia (NMIA) for the certification of a freeze-dried human urine reference material (CRM NMIA MX005). The method measures δ(13)C values by gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) analysis following hydrolysis, solvent extraction and high performance liquid chromatography (HPLC) purification. Reference δ(13)C values for testosterone metabolites etiocholanolone, androsterone, and endogenous reference compounds (ERCs) 11β-hydroxyandrosterone and pregnanediol were determined, as well as information δ(13)C values for testosterone, epitestosterone, 11-oxoetiocholanolone, and a range of differences (Δ(13)C) between testosterone metabolites and ERCs. The measurement uncertainty was rigorously evaluated with expanded uncertainties for the reference δ(13)C values between 1.1 and 1.6 ‰ at the 95% coverage level.

EFFECT OF LONG-TIME STORAGE ON THE CONTENT OF POLYPHENOLS AND ASCORBIC ACID IN OSMO-CONVECTIVELY DRIED AND OSMO-FREEZE-DRIED FRUITS

ABSTRACT The objective of the study was to investigate the stability of polyphenols and ascorbic acid in osmo-dried fruits: apples, sour cherries and blackcurrants, stored for 12 months. After 1 year of storage, the content of total polyphenols and anthocyanins in convectively dried blackcurrants was decreased by 45% and 90%, respectively. Flavonols remained highly stable. Convectively dried sour cherries contained particularly labile substances, including cyanidin derivatives, which content decreased by 98% after 6 months. Polyphenols in freeze-dried materials were more stable. After 1 year of storage, their content remained at 60–80% of the initial level. A high stability was in particular observed for delphinidin-3-rutinoside in blackcurrants, and cyanidin derivatives, chlorogenic acid, neochlorogenic acid and quercetin glucoside in sour cherries. The stability of ascorbic acid was also much higher in freeze-dried blackcurrants than in convectively dried fruit. After 1 year of storage, almost 70% of ascorbic acid was preserved in freeze-dried material. PRACTICAL APPLICATIONS Analyzed material was obtained in industrial conditions. Final products were used to produce bars; it can also be used in manufacturing of muesli and similar products. The results of performed analyses show a change in the content of polyphenols and ascorbic acid during product storage and could be useful to define the time in which a product is characterized by the highest content of health-promoting substances.

Sr isotope measurements in beef—analytical challenge and first results

The strontium isotope ratio ((87)Sr/(86)Sr) in beef, derived from 206 European cattle, has been measured. These cattle were located in 12 different European regions within France, Germany, Greece, Ireland, Italy, Spain and the UK. As animal protein is known to be a difficult material on which to conduct Sr isotope analysis, several investigations were undertaken to develop and improve the sample preparation procedure. For example, Sr isotope analysis was performed directly on freeze-dried meat and defatted dry mass from the same samples. It was found that enormous differences-sometimes exceeding the measurement uncertainty-could occur between the fractions and also within one sample even if treated in the same manner. These variations cannot be definitely allocated to one cause but are most likely due to inhomogeneities caused by physiological and biochemical processes in the animals as post mortem contamination during analytical processing could be excluded. For further Sr isotope measurements in meat, careful data handling is recommended, and for the authentic beef samples within this project, it was decided to use only freeze-dried material. It can be demonstrated, however, that Sr isotope measurements in beef proteins are a valuable tool for authentication of geographic origin. Although partly overlapping, some of the European sampling sites could be discriminated even by only using (87)Sr/(86)Sr.

Frozen polymerization for aligned porous structures with enhanced mechanical stability, conductivity, and as stationary phase for HPLC

A directional freezing and frozen polymerization method is developed to prepare crosslinked aligned porous polymers with improved mechanical stability. Monomer solutions are directionally frozen in liquid nitrogen to orientate the growth of solvent crystals. The frozen samples are polymerized by UV irradiation. The solvent is removed under vacuum at room temperature to produce aligned porous structure. The mechanical stability is improved by two orders of magnitude compared to the usually freeze-dried porous materials. The materials are modified with graphene and a conducting polymer to achieve conductivity at 1.9 × 10−4 S cm−1 and 5.2 × 10−6 S cm−1, with the stable aligned pore structures maintained. The aligned porous monolith is also assessed by high performance liquid chromatography (HPLC), showing fast separation of hydrocarbon compounds with low back pressure at 59 bar.

Feasibility study for producing a carrot/potato matrix reference material for 11 selected pesticides at EU MRL level: Material processing, homogeneity and stability assessment

The feasibility for producing a matrix reference material for selected pesticides in a carrot/potato matrix was investigated. A commercially available baby food (carrot/potato-based mash) was spiked with 11 pesticides at the respective EU maximum residue limits (MRLs), and further processed by either freezing or freeze-drying. Batches of some 150units were produced per material type. First, the materials were assessed for the relative amount of pesticide recovered after processing (ratio of pesticide concentration in the processed material to the initially spiked pesticide concentration). In addition, the materials’ homogeneity (bottle-to-bottle variation), and the short-term (1month) and mid-term (5months) stability at different temperatures were assessed. For this, an in-house validated GC–EI-MS method operated in the SIM mode with a sample preparation procedure based on the QuEChERS (“quick, easy, cheap, effective, rugged, and safe”) principle was applied. Measurements on the frozen material provided the most promising results (smallest analyte losses during production), and also freeze-drying proved to be a suitable alternative processing technique for most of the investigated pesticides. Both the frozen and the freeze-dried material showed to be sufficiently homogeneous for the intended use, and storage at −20°C for 5months did not reveal any detectable material degradation. The results constitute an important step towards the development of a pesticide matrix reference material.

Headspace Moisture Mapping and the Information That Can Be Gained about Freeze-Dried Materials and Processes

Regulatory authorities require proof that lyophilization (freeze drying) cycles have been developed logically and demonstrate uniformity. One measure of uniformity can be consistency of residual water content throughout a batch. In primary drying, heat transfer is effected by gaseous convection and conduction as well as the degree of shelf contact and evenness of heat applied; therefore residual water can be affected by container location, degree of container/tray/shelf contact, radiative heating, packing density, product formulation, and the cycle conditions themselves. In this study we have used frequency modulation spectroscopy (FMS) to create a map of headspace moisture (HSM) for 100% of vials within a number of freeze-dried batches. Karl Fischer (KF)/HSM correlations were investigated in parallel with the moisture mapping studies. A clear, linear relationship was observed between HSM and KF values for vials containing freeze-dried sucrose, implying a relatively straightforward interaction between water and the lyophilized cake for this material. Mannitol demonstrated a more complex correlation, with the interaction of different crystalline forms giving important information on the uniformity of the material produced. It was observed that annealing had a significant impact on the importance of heat transfer by conduction for vials in direct and non-direct contact with the shelf. Moisture mapping of all vials within the freeze dryer enabled further information to be obtained on the relationship of the formulation, process conditions, and equipment geometry on the intra-batch variability in HSM level. The ability of FMS to allow 100% inspection could mean that this method could play an important part in process validation and quality assurance. Lyophilization, also known as freeze drying, is a relatively old technique that has been used in its most basic form for thousands of years (e.g., preservation of fish and meat products). In its more advanced form it is used to preserve many medical products; for example, many vaccines are not stable in solution and therefore need to be dried to allow long-term storage. In order to produce a freeze-dried vaccine, a complex understanding of the processes and critical temperatures is required. Once these have been understood, the material is dried to give relatively low moisture content (e.g., 2% w/w.) This low moisture content is critical for the long-term stability of the product, allowing doctors/chemists to store these goods on site for use when required. This research paper provides further information on a technique called frequency modulation spectroscopy (FMS) that has been used to map the moisture variation across samples within a freeze dryer, enabling us to increase our understanding of the role various processing conditions play on the relationship between the product and water. It has demonstrated its potential application for 100% batch monitoring and the validation of a system or assessment of changes made. This method could assist in improving quality assurance and ultimately the final product that reaches the consumer.

Preparation of enhanced three-dimensional porous chitosan scaffolds by acetylation and aqueous extraction

We have devised a method to prepare a 3-dimensional (3D) porous acetylated chitosan scaffold for use as a cell adhesion matrix in tissue engineering applications. The scaffold was prepared by molding a mixture of chitosan and gelatin (as porogen), then removing the uncomplexed porogen by aqueous extraction from the freeze-dried material prior to acetylation. The extent of chitosan acetylation according to the reaction time was observed by X-ray diffraction (XRD) analysis. The differences between the aqueous-extracted and control phase-separated chitosan scaffolds in terms of pore morphology and interconnectivity were examined by scanning electron microscopy (SEM), enzymatic degradation, and surface roughness tests. The fibroblast cell line NIH-3T3 was used to test relative cell affinities for the acetylated versus untreated (control) chitosan scaffolds. The acetylated 3D porous scaffolds showed high interconnectivity and improved biocompatibility properties. Thus, these scaffolds may be very useful for a variety of tissue engineering applications.

Mutually reinforced multicomponent polysaccharide networks.

Networks made from chitosan and alginate have been utilized as prospective tissue engineering scaffolds due to material biocompatibility and degradability. Calcium (Ca(2+) ) is often added to these networks as a modifier for mechanical strength enhancement. In this work, we examined changes in the bulk material properties of different concentrations of chitosan/alginate mixtures (2, 3, or 5% w/w) upon adding another modifier, chondroitin. We further examined how material properties depend on the order the modifiers, Ca(2+) and chondroitin, were added. It was found that the addition of chondroitin significantly increased the mechanical strength of chitosan/alginate networks. Highest elastic moduli were obtained from samples made with mass fractions of 5% chitosan and alginate, modified by chondroitin first and then Ca(2+) . The elastic moduli in dry and hydrated states were (4.41 ± 0.52) MPa and (0.11 ± 0.01) MPa, respectively. Network porosity and density were slightly dependent on total polysaccharide concentration. Average pore size was slightly larger in samples modified by Ca(2+) first and then chondroitin and in samples made with 3% starting mass fractions. Here, small-angle neutron scattering (SANS) was utilized to examine mesh size of the fibrous networks, mass-fractal parameters and average dimensions of the fiber cross-sections prior to freeze-drying. These studies revealed that addition of Ca(2+) and chondroitin modifiers increased fiber compactness and thickness, respectively. Together these findings are consistent with improved network mechanical properties of the freeze-dried materials.

Determination of Soluble Immunoglobulin G in Bovine Colostrum Products by Protein G Affinity Chromatography–Turbidity Correction and Method Validation

Immunoglobulin-containing food products and nutraceuticals such as bovine colostrum are of interest to consumers as they may provide health benefits. Commercial scale colostrum products are valued for their immunoglobulin G (IgG) content and therefore require accurate analysis. One of the most commonly used methods for determining total soluble IgG in colostrum products is based on affinity chromatography using a Protein G column and UV detection. This paper documents improvements to the accuracy of the Protein G analysis of IgG in colostrum products, especially those containing aggregated forms of IgG. Capillary electrophoresis-sodium dodecyl sulfate (CE-SDS) analysis confirmed that aggregated IgG measured by Protein G does not contain significant amounts of casein or other milk proteins. Size exclusion chromatography identified the content of soluble IgG as mainly monomeric IgG and aggregated material MW > 450 kDa with small amounts of dimer and trimer. The turbidity of the eluting IgG, mainly associated with aggregated IgG, had a significant effect on the quantitative results. Practical techniques were developed to correct affinity LC data for turbidity on an accurate, consistent, and efficient basis. The method was validated in two laboratories using a variety of colostrum powders. Precision for IgG was 2-3% (RSD(r)) and 3-12% (RSD(R)). Recovery was 100.2 ± 2.4% (mean ± RSD, n = 10). Greater amounts of aggregated IgG were solubilized by a higher solution:sample ratio and extended times of mixing or sonication, especially for freeze-dried material. It is concluded that the method without acid precipitation and with turbidity correction provides accurate, precise, and robust data for total soluble IgG and is suitable for product specification and quality control of colostrum products.

Effect of preliminary processing, method of drying and storage temperature on the level of antioxidants in kale (Brassica oleracea L. var. acephala) leaves

The levels of vitamin C, polyphenol constituents and Trolox equivalent antioxidant activity (TEAC) were analyzed in raw kale leaves, blanched leaves and dried leaves obtained using air and freeze-drying methods. 100 g of raw kale leaves contained 683 mg vitamin C and 2236 mg polyphenols (identified using the HPLC method); the level of antioxidative activity was 71 μM Trolox/1 g dry matter. Compared with the raw material, blanching before drying brought about significant decreases of 15% in vitamin C, 32% in polyphenols and13% in TEAC. After 12-month storage, air-dried material retained 30–37% polyphenols; 43–57% vitamin C; and 41–50% of the initial TEAC level; the corresponding values for freeze-dried material were 40–47%; 50–65% and 54–66% depending on the type of sample. Freeze-dried kale leaves contained higher levels of antioxidants than air-dried material: polyphenols, vitamin C and TEAC were respectively 36%, 15%and 33% higher.

The influence of freeze drying conditions on microstructural changes of food products

The drying of food products can result in significant changes in the chemical composition, morphology and physical properties of foods and can lead to stabilized products with longer shelf-life and easier commercialization. Information on porous structure of foods is very important, characterizing the quality and texture of dehydrated food products. Therefore, structural properties, such as porosity, bulk density and true density of freeze-dried food products were investigated as affected by process conditions. Rice kernels were boiled for different time periods and agricultural products, including potato, mushroom and strawberry, were cut into cubes. The samples were frozen, tempered in liquid N2 and freeze-dried, under various vacuum conditions, using a laboratory freeze-dryer. True density of the products was measured using a helium stereo-pycnometer. Bulk density was obtained by measuring the dimensions of the samples with a Vernier caliper. Simple mathematical models were developed in order to correlate the structural properties with process conditions. The microstructure of food products was also analyzed by Scanning Electron Microscopy and image analysis. Bulk density of freeze-dried materials increased with the applied pressure during freeze-drying, while porosity decreased. In addition, bulk density of freeze-dried rice kernels decreased with the increment of boiling time, while porosity increased. The changes in bulk density and porosity were closely supported by microstructural observations, according to SEM images. The microstructural changes of products, freeze-dried under various vacuum conditions, can be predicted using the proposed models and image analysis.

Effect of drying and storage conditions on caffeic acid derivatives and total phenolics of Echinacea Purpurea grown in Taiwan

Fresh flowers, leaves, stems and roots of Echinacea purpurea were subjected to vacuum freeze-drying, cool wind-drying (30 °C), and hot air-drying (40, 55 or 70 °C), and then stored under different environmental conditions. The cichoric acid was the main phenolic compound detected in dried E. purpurea materials, followed by caftaric acid. The bioactive constituent contents in different plant parts were in the descending order: flowers > leaves > stems > roots. Both caffeic acid derivatives and total phenolics contents were affected by drying method and storage condition. Cool wind-dried materials retained more bioactive constituents content (>85%) compared to vacuum freeze-dried materials. The packing material also affected the storability of E. purpurea materials. The storability results indicated that the freeze-dried E. purpurea materials sealed in polyethylene terephthalate/aluminum foil/polyethylene or nylon/polyethylene bags and stored under 10–20 °C and 40–60% relative humidity without light conditions retained the highest content of bioactive compounds.

Determination of quercetin glycosides and free quercetin in buckwheat by capillary micellar electrokinetic chromatography

A simple and reliable method for determination of quercetin glycosides and free quercetin in buckwheat flower, leaves, stems and achenes was developed. The method consists of flavonoid extraction from freeze-dried homogenous material in 50% v/v methanol solution and in presence of an antioxidant, cleaning of extract and analyte isolation using SPE. Analytical step uses capillary micellar electrokinetic chromatography. The working ranges, LOD and LOQ, recovery, precision and measurement uncertainty were calculated. The method is suitable for samples from buckwheat. The highest content of rutin was found in flowers of both kinds of buckwheat (99,400 mg/kg in F. esculentum, 108,000 mg/kg in F. tataricum). The free quercetin occurs in flowers and achenes of F. esculentum, whereas flowers and achenes of F. tataricum contained quercitrin.

Rapid Method for Determination of Residual tert-Butanol in Liposomes Using Solid-Phase Microextraction and Gas Chromatography

A simple, rapid, and reliable method to detect residual levels of tert-butanol in liposomes using sec-butanol as an internal standard has been developed. Solid-phase microextraction (SPME) followed by gas chromatographic analysis was used to quantify the amount of residual tert-butanol in freeze-dried liposome material. Only 1 min was necessary for reproducible amounts of analyte to absorb onto the SPME fiber, and because this method requires very little sample preparation, a single analysis can be completed in less than 15 min. This method had a linear range of 10-600 microg/mL. Careful control of times of temperature equilibration and exposure to headspace was necessary to ensure reproducible results. This method can easily be applied to other applications in the food and pharmaceutical industries where detection of residual solvents, such as hexane and chloroform, is necessary.