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Biochemical and spectral characterization of micro- and macroalgae

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Abstract

Algae contain many compounds which are of interest to the pharmaceutical, agricultural and food industries, among others. Despite the potential for many applications, a quick, accurate and inexpensive method for characterising algae is lacking. The objective of this thesis was to investigate the ability to determine protein, carbohydrate and fatty acid contents in algae, using infrared spectroscopy. Algal samples were analysed using some of the most accurate chemical methods, by hydrolysing proteins and polysaccharides and quantifying the resulting amino acid and monosaccharide contents using chromatography techniques. Fatty acid contents were similarly quantified by extraction followed by gas chromatography. This data was subsequently used to calibrate spectral prediction models, using different infrared spectroscopies. The accuracy of protein prediction by diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS) and near-infrared spectroscopy (NIR) using partial least squares regression (PLSR) was compared to traditional methods like N-protein ratios and colorimetric assays. DRIFTS, attenuated total reflectance (ATR) Fourier transform infrared spectroscopy (ATR) and NIR were also comparatively used to characterise seaweed carbohydrates. Fatty acid contents were studied using DRIFTS in both microalgae and seaweeds. Infrared spectroscopy coupled with PLSR was shown to be highly accurate in characterising algal biomass, provided a sufficiently robust library of calibration samples. These methods require little or no chemicals and are rapid and easy to use, making them both environmentally and economically attractive.
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Comparison of data of protein content in algae is very difficult, primarily due to differences in the analytical methods employed. The different extraction procedures (exposure to water, grinding, etc.), protein precipitation using different amounts of 25% trichloroacetic acid and quantification of protein by two different methods and using two protein standards were evaluated. All procedures were tested using freeze-dried samples of three macroalgae: Porphyra acanthophora var. acanthophora, Sargassum vulgare and Ulva fasciata. Based on these results, a protocol for protein extraction was developed, involving the immersion of samples in 4.0 mL ultra-pure water for 12 h, followed by complete grinding of the samples with a Potter homogeniser. The precipitation of protein should be done with 2.5:1 25% TCA:homogenate (v/v). The protocol for extraction and precipitation of protein developed in this study was tested with other macroalgae (Aglaothamnion uruguayense, Caulerpa fastigiata, Chnoospora minima, Codium decorticatum, Dictyota menstrualis, Padina gymnospora and Pterocladiella capillacea) and microalgae (Amphidinium carterae, Dunaliella tertiolecta, Hillea sp., Isochrysis galbana and Skeletonema costatum). Comparison with the actual protein content determined from the sum of amino acid residues, suggests that Lowry's method should be used instead of Bradford's using bovine serum albumin (BSA) as protein standard instead of casein. This may be related to the reactivity of the protein standards and the greater similarity in the amino acid composition of BSA and algae. The current results should contribute to more accurate protein determinations in marine algae.
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The methods of Lowry and Bradford, commonly used for protein determination, were compared regarding the level of interference of some substances used for glucoamylase precipitation by ethanol. The method of Bradford suffers no interference while the method of Lowry showed protein concentration values 20% increased in the presence of ethanol and Tris. Despite these interferences, the Lowry method can evaluate more accurately the increase of purity during fractionation, due to its sensitivity to low molecular weight (below 6kDa) proteins and peptides.
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Algae hold particular promise as a feedstock for biomaterials, as they are capable of producing a wide variety of polymers with the properties required for 3D printing. However, the use of algal polymers has been limited to alginate, agar, carrageenan, and ulvan extracted from seaweeds. Diverse algal taxa beyond seaweeds have yet to be explored. In this comprehensive review, we discuss available algal biomaterials, their properties, and emerging applications in 3D printing techniques. We also identify elite algal strains to be used in 3D printing and comment on both advantages and limitations of algal biomass as a printing material. Global 3D printing market trends and material demands are also critically analyzed. Finally, the future prospects, opportunities, and challenges for using algal polymers in 3D printing market for a sustainable economy are discussed. We hope this review will provide a foundation for exploring the 3D printable biomaterials from algae.
Article
Two well characterised alginate samples with high and low M/G ratio were subjected to radiation decontamination dose at 15 kGy. Subsequently, a number of techniques were used to determine the effect of radiation on the gelling properties. These techniques include: (GPC-MALLS) gel permeation chromatography linked to a multi-angle laser light scattering and refractive index detector for the determination of molecular weight and distribution. To observe the mechanics, the deformation with flow of α-β alginates rheological measurements in the dynamic oscillation modes, and distinguish the functional group of the samples along with identifying the M/G ratio for quantitative determination of mannuronic and guluronic acid, the FTIR techniques were performed respectively. The results showed clear reduction in the molecular weight following irradiation which was also accompanied by a loss of the gelling properties as well as clear reduction intensity of FTIR peak at 1080 -1400 cm⁻¹ for carboxylic group, OH-bonding obtained at 1030 cm⁻¹ (1026 & 1028 cm⁻¹) while the weak signal and skeletal region is attributed at >2800 & <700 cm⁻¹ respectively. A sample (n = 1; H120L) is homopolyguluronic enriched (820 cm⁻¹ in fingerprint region) with small amount of mannuronic acid fraction. Sample (n = 2; LFR-5/60) is heteropolyguluronic acid enriched in presence of tiny amount guluronic acid (814 cm⁻¹ peak) i.e. comparatively high G to M in this fraction but % G is lower than n = 1. The rheological parameter storage modulus (G`) and loss modulus (G``) were noticed higher with the constant frequency mode, a solid-like nature gels was obtained with increasing the frequency in a certain time of relaxation. The gelling properties of the samples calculated is M/G < 1 which able to make hard and rigid gel. Thus, radiation effects for rheology and FTIR in case of functional group distribution was significant.
Article
Background With the increase in world population, decreased farmland, and global climate changes, ensuring adequate food supply to maintain food security is of immediate attention. As a key macronutrient for human health, the supply of sufficient dietary protein is undoubted of concern. Animal proteins are good dietary protein sources, but their production incurs a high carbon footprint; this has driven the effort to seek alternative protein sources. Scope and approach This review aimed to elaborate on the scientific research progress in red seaweed proteins, including the nutrition, functionalities, methods of extraction, and to explore their prospects as an alternative protein source. Applications of red seaweed protein in food and nutraceutical industries, environmental impact, affordability, and related safety concerns were also discussed. Key findings and conclusions Red seaweeds have a comparable essential amino acid profile to ovalbumin, representing a sustainable alternative to terrestrial proteins. Pre-treatment and extraction methods are pivotal in modulating protein digestibility and functionalities; enzymatic extraction approaches appear to improve nutritional value and food functionalities. Red seaweed proteins have a wide range of applications in food based on their physicochemical properties, while their bioactivities can be tailored for nutraceutical purposes. The use of red seaweed proteins as functional food ingredients is emerging, with good potential in bioactive microencapsulation. Efforts are required to improve the seaweed cultivation process to a commercial scale and gain consumer acceptance in the western world. More research is also necessary to enhance seaweed protein extraction and improve their functionalities for food and nutraceutical applications.
Article
Brown seaweeds are the most studied and exploited algae type for ruminant nutrition due to their biomass availability, ease of harvest and content of bioactive compounds. Infrared spectroscopy represents a rapid, non-invasive and chemical-free technique that is widely applied for the chemical characterization and digestible quality of many terrestrial forages. However, there is limited information regarding its application to seaweeds. This study compared the effectiveness of Near-Infrared (NIR: 9000-4000 cm⁻¹) and Mid-Infrared (MIR: 4000-400 cm⁻¹) spectroscopy to measure the nutritional value and in vitro dry matter rumen digestibility of brown seaweeds. Due to the small number of seaweed samples available, 40 samples were analysed in triplicate with a total dataset of 120 samples. For partial least-squares regression model development and evaluation purposes, the dataset (n = 120) was divided into two subsets, the first one for training and model development purposes (70% of data, n = 84), and the second one for model testing and evaluation (internal evaluation) purposes (30% of data, n = 36).Partial least-squares regression was employed to develop multivariate calibration models which were internally and externally validated. The samples were analysed using established wet chemistry methods which were regarded as the reference methods. NIR showed high accuracy for the quantitative prediction of crude protein (R²P = 0.99; RMSEP = 0.51; RER = 26.9; RPD = 6.9) and total polyphenolic content (R²P = 0.94; RMSEP = 0.20; RER= 10; RPD= 3.2), whereas MIR could only accurately predict crude protein (R²P = 0.96; RMSEP = 1.12; RER = 11.64; RPD = 3.14). Ash, neutral and acid detergent fibre, lignin (sa) and in vitro dry matter rumen digestibility models showed limited applicability for quantitative measurements (R²P < 0.85; RPD < 2). Overall, NIR and MIR could be used to rapidly evaluate the nutritional composition and digestibility of brown seaweeds in their dried form but further evaluation on an external database would be required to assess the robustness of these models on unrelated data. Furthermore, the use of these spectroscopic methods showed lower accuracy and precision compared to wet chemistry methods, which better qualifies them for screening rather than confirmatory analysis.
Article
3D bioprinting has emerged as a promising technology in the field of tissue engineering and regenerative medicine due to its ability to create anatomically complex tissue substitutes. However, it still remains challenging to develop bioactive bioinks that provide appropriate and permissive environments to instruct and guide the regenerative process in vitro and in vivo. In this study alginate sulfate, a sulfated glycosaminoglycan (sGAG) mimic, was used to functionalize an alginate-gelatin methacryloyl (GelMA) interpenetrating network (IPN) bioink to enable the bioprinting of cartilaginous tissues. The inclusion of alginate sulfate had a limited influence on the viscosity, shear-thinning and thixotropic properties of the IPN bioink, enabling high-fidelity bioprinting and supporting mesenchymal stem cell (MSC) viability post-printing. The stiffness of printed IPN constructs greatly exceeded that achieved by printing alginate or GelMA alone, while maintaining resilience and toughness. Furthermore, given the high affinity of alginate sulfate to heparin-binding growth factors, the sulfated IPN bioink supported the sustained release of transforming growth factor-β3 (TGF-β3), providing an environment that supported robust chondrogenesis in vitro, with little evidence of hypertrophy or mineralization over extended culture periods. Such bioprinted constructs also supported chondrogenesis in vivo, with the controlled release of TGF-β3 promoting significantly higher levels of cartilage-specific extracellular matrix deposition. Altogether, these results demonstrate the potential of bioprinting sulfated bioinks as part of a ‘single-stage’ or ‘point-of-care’ strategy for regenerating cartilaginous tissues. Statement of significance This study highlights the potential of using sulfated interpenetrating network (IPN) bioink to support the regeneration of phenotypically stable articular cartilage. Construction of interpenetrating networks in the bioink enables unique high-fidelity bioprinting and provides synergistic increases in mechanical properties. The presence of alginate sulfate enables the capacity of high affinity-binding of TGF-β3, which promoted robust chondrogenesis in vitro and in vivo.
Article
Seaweeds contain minerals, vitamins, soluble dietary fibers, and flavonoids, which are regarded as preventive agents against lifestyle-related diseases. Seaweeds are consumed commonly in East Asian countries including Japan. Thus, intake of seaweeds might contribute to Japanese longevity via prevention of lifestyle-related diseases. Recently, two large Japanese cohort studies have reported the association of seaweed intake with reduced risk of cardiovascular diseases. On the other hand, seaweeds also contain iodine and heavy metals such as arsenic species, which are considered to have adverse effects on health. We here reviewed studies of the association between seaweed intake and mortality from or incidence of cancer and cardiovascular diseases, and their risk factors such as blood pressure or serum lipids. We also summarized the adverse effects of iodine and arsenic species in seaweeds. Although seaweeds have not been widely consumed in Western countries, dietary diversification and an increased proportion of immigrants from East Asia may increase seaweed consumption in those countries. Further epidemiological studies including observational and interventional studies are necessary to clarify the effects of seaweeds on disease and health.
Article
The current hydrocolloid industry requires new techniques for biomass characterization, which can quickly and ecologically characterize contained sugars. This work proposes the use of Fourier Transform Infrared microspectroscopy in combination with multivariate methods, to localize and identify the main carbohydrates and other components present in fresh brown seaweeds, avoiding time-consuming samples pre-treatments. Infrared images of Macrocystis pyrifera samples were analyzed by Multivariate Curve Resolution-Alternating Least Squares (MCR-ALS) and Principal Component Analysis (PCA) as chemometrics techniques to identify the compounds. MCR-ALS was the best strategy, delivering pure spectra of chemical compound that PCA did not. The carbohydrates identified by this method were 1-3-β-glucans divided into endofibers and laminarin; two types of fucoidans (rich in fucose or mannuronic acid), alginate and mannitol, besides other compounds such as proteins. This technique represents an opportunity for the hydrocolloid industry for a modern, rapid and environmentally-friendly characterization of macroalgal biomass to enhance its use.
Article
The use of alginates as films in food applications has increased in the recent years due to their swelling capacity and overall functionality. This behaviour is a result of their capacity to crosslink with Ca²⁺ ion. Aiming to fully understand the effect of calcium chloride (CaCl2) crosslinking and the mannuronic (M) and guluronic (G) acid ratio (M/G) of alginate structure in the films' properties, alginate-based films with different (M/G) ratios were crosslinked at increasing CaCl2 concentrations. Films were produced by casting, and characterized in terms of mechanical properties (tensile strength and elongation-at-break), opacity, water sensitivity (moisture content, solubility and water vapour permeability) and morphology, evaluated by scanning electronic microscopy (SEM). Chemical interactions were studied by Fourier Transform Infrared Spectroscopy (FTIR) to assess possible chemical modifications of alginate-based films after crosslinking. Crosslinking significantly affected the alginate structure and properties, decreasing film thickness, moisture content, solubility and water vapour permeability of the alginate-based films. The mechanical properties were also influenced by the crosslinking and high CaCl2 concentrations lead to an increase of tensile strength. Results showed a relation between M/G ratios and CaCl2 concentrations and the resulting film's properties. Alginate and the respective crosslinker should be chosen taking into account M/G ratio, since high contents of M residues lead to fragile and flexible films and high content of G residues to stronger films, and these properties are highly dependent on the concentration of CaCl2. Overall, alginate-based films are a good candidate to obtain tailored made edible films for food applications. Further investigation should be done to fully understand the effect of the alginate chain composition and order (e.g. MM, GG, GM, MG) in alginate-based films properties.
Article
Increasing oil prices and depletion of existing fossil fuel reserves, combined with the continuous rise in greenhouse gas emissions, have fostered the need to explore and develop new renewable bioenergy feedstocks that do not require arable land and freshwater resources. In this regard, prolific biomass growth of invasive aquatic weeds in wastewater has gained much attention in recent years in utilizing them as a potential feedstock for bioenergy production. Aquatic weeds have an exceptionally higher reproduction rates and are rich in cellulose and hemicellulose with a very low lignin content that makes them an efficient next generation biofuel crop. Considering their potential as an effective phytoremediators, this review presents a model of integrated aquatic biomass production, phytoremediation and bioenergy generation to reduce the land, fresh water and fertilizer usage for sustainable and economical bioenergy.
Article
This study explored Himanthalia elongata brown seaweed as a potential source of dietary fucoxanthin which is a promising medicinal and nutritional ingredient. The seaweed was extracted with low polarity solvents (n-hexane, diethyl ether, and chloroform) and the crude extract was purified with preparative thin layer chromatography (P-TLC). Identification, quantification and structure elucidation of purified compounds was performed by LC-DAD-ESI-MS and NMR (1H and 13C). P-TLC led purification yielded 18.6 mg/g fucoxanthin with 97% of purity based on the calibration curve, in single-step purification. LC-ESI-MS (parent ion at m/z 641 [M + H-H2O]+) and NMR spectra confirmed that the purified band contained all-trans-fucoxanthin as the major compound. Purified fucoxanthin exhibited statistically similar (p > 0.05) DPPH scavenging capacity (EC50: 12.9 μg/mL) while the FRAP value (15.2 μg trolox equivalent) was recorded lower (p < 0.05) than the commercial fucoxanthin. The promising results of fucoxanthin purity, recovery and activity suggested that H. elongata seaweed has potential to be exploited as an alternate source for commercial fucoxanthin production.
Article
Importance: Defining what represents a macronutritionally balanced diet remains an open question and a high priority in nutrition research. Although the amount of protein may have specific effects, from a broader dietary perspective, the choice of protein sources will inevitably influence other components of diet and may be a critical determinant for the health outcome. Objective: To examine the associations of animal and plant protein intake with the risk for mortality. Design, setting, and participants: This prospective cohort study of US health care professionals included 131 342 participants from the Nurses' Health Study (1980 to end of follow-up on June 1, 2012) and Health Professionals Follow-up Study (1986 to end of follow-up on January 31, 2012). Animal and plant protein intake was assessed by regularly updated validated food frequency questionnaires. Data were analyzed from June 20, 2014, to January 18, 2016. Main outcomes and measures: Hazard ratios (HRs) for all-cause and cause-specific mortality. Results: Of the 131 342 participants, 85 013 were women (64.7%) and 46 329 were men (35.3%) (mean [SD] age, 49 [9] years). The median protein intake, as assessed by percentage of energy, was 14% for animal protein (5th-95th percentile, 9%-22%) and 4% for plant protein (5th-95th percentile, 2%-6%). After adjusting for major lifestyle and dietary risk factors, animal protein intake was weakly associated with higher mortality, particularly cardiovascular mortality (HR, 1.08 per 10% energy increment; 95% CI, 1.01-1.16; P for trend = .04), whereas plant protein was associated with lower mortality (HR, 0.90 per 3% energy increment; 95% CI, 0.86-0.95; P for trend < .001). These associations were confined to participants with at least 1 unhealthy lifestyle factor based on smoking, heavy alcohol intake, overweight or obesity, and physical inactivity, but not evident among those without any of these risk factors. Replacing animal protein of various origins with plant protein was associated with lower mortality. In particular, the HRs for all-cause mortality were 0.66 (95% CI, 0.59-0.75) when 3% of energy from plant protein was substituted for an equivalent amount of protein from processed red meat, 0.88 (95% CI, 0.84-0.92) from unprocessed red meat, and 0.81 (95% CI, 0.75-0.88) from egg. Conclusions and relevance: High animal protein intake was positively associated with mortality and high plant protein intake was inversely associated with mortality, especially among individuals with at least 1 lifestyle risk factor. Substitution of plant protein for animal protein, especially that from processed red meat, was associated with lower mortality, suggesting the importance of protein source.
Article
A novel method that uses the aqueous sodium alginate solution for direct gelation with metal ions is developed for effective removal and recovery of heavy metals from industrial wastewater. The experimental study was conducted on Pb2+, Cu2+, and Cd2+ as the model heavy metals. The results show that gels can be formed rapidly between the metals and alginate in less than 10 min and the gelation rates fit well with the pseudo second-order kinetic model. The optimum dosing ratio of alginate to the metal ions was found to be between 2:1 and 3:1 for removing Pb2+ and around 4:1 for removing Cu2+ and Cd2+ from wastewater, and the metal removal efficiency by gelation increased as the solution pH increased. Alginate exhibited a higher gelation affinity toward Pb2+ than Cu2+ and Cd2+, which allowed a selective removal of Pb2+ from the wastewater in the presence of Cu2+ and Cd2+ ions. Chemical analysis of the gels suggests that the gelation mainly occurred between the metal ions and the COO− and OH groups on alginate. By simple calcination of the metal-laden gels at 700 °C for 1 h, the heavy metals can be well recovered as valuable resources. The metals obtained after the thermal treatment are in the form of PbO, CuO, and CdO nanopowders with crystal sizes of around 150, 50, and 100 nm, respectively.
Article
Marine algal seaweed species are often regarded as an underutilized bioresource many have been used as a source of food, industrial raw materials, and in therapeutic and botanical applications for centuries. Seaweeds are best known for the natural polysaccharides. Moreover, seaweed and seaweed-derived products have been widely used as a key ingredient in many pharmaceutical preparations like tablets, as rate retardant in sustained and extended release dosage forms, as stabilizer in liquid orals, cosmetics, implants and as bioplastic in packaging industry. This article provides a comprehensive review of various seaweed species and seaweed products with an emphasis on the use of this renewable bioresource in pharmaceutical industry and medical field.
Chapter
Seaweeds, in particular red seaweeds, have been shown to possess significant levels of protein and in some cases contain higher quantities than some conventional protein-rich foods; however, seaweeds are rarely promoted for the nutritional value of their proteins, peptides, and amino acids. Seaweed-derived proteins, peptides, and amino acids are known to have a variety of health benefits against hypertension, including well-established antioxidative effects. Also, various studies indicate promising anticancer and antidiabetes effects. Seaweed-derived proteins, peptides, and amino acids have multiple health benefits; thus their consumption should be encouraged. In this chapter, identified bioactivities and potentialities of seaweed-derived proteins, peptides, and amino acids will be discussed for future nutraceutical and cosmeceutical applications.
Chapter
Seaweeds contain many molecules of interest including polysaccharides (~50% of the algae) as alginate, agar, carrageenans, fucoidan and laminaran. Many of these polysaccharides are used in foods as thickener, gelling agent and emulsion stabilizer while others are mostly known for their biological activities. This chapter highlights the recent knowledge on food grade and none-food polysaccharides found in algae. Their common source, structure, extraction method and food utilization (or potential utilization) are presented. In future years, food enriched in seaweed extracts or containing purified algal polysaccharide may be used to increase the functionality of the food available on the market.
Article
This chapter describes three different spectroscopic methods for structural characterization of the commercial important hydrocolloid alginate extracted from brown seaweed. The "golden" reference method for characterization of the alginate structure is (1)H liquid-state NMR of depolymerized alginate polymers using a stepwise hydrolysis. Having implemented this method, predictive and rapid non-destructive methods using vibrational spectroscopy and chemometrics can be developed. These methods can predict the M/G-ratio of the intact alginate powder with at least the same precision and accuracy as the reference method in a fraction of the time that is required to measure the alginate using the reference method. The chapter also demonstrates how solid-state (13)C CP/MAS NMR can be used to determine the M/G ratio on the intact sample by the use of multivariate chemometrics and how this method shares the characteristics of the solid-state non-destructive IR method rather than its liquid-state counterpart.
Article
Fourier transform infrared (FT-IR) spectroscopy is a fast, sensitive, inexpensive, and nondestructive technique for chemical profiling of plant materials. In this chapter we discuss the instrumental setup, the basic principles of analysis, and the possibilities for and limitations of obtaining qualitative and semiquantitative information by FT-IR spectroscopy. We provide detailed protocols for four fully customizable techniques: (1) Diffuse Reflectance Infrared Fourier Transform Spectroscopy (DRIFTS): a sensitive and high-throughput technique for powders; (2) attenuated total reflectance (ATR) spectroscopy: a technique that requires no sample preparation and can be used for solid samples as well as for cell cultures; (3) microspectroscopy using a single element (SE) detector: a technique used for analyzing sections at low spatial resolution; and (4) microspectroscopy using a focal plane array (FPA) detector: a technique for rapid chemical profiling of plant sections at cellular resolution. Sample preparation, measurement, and data analysis steps are listed for each of the techniques to help the user collect the best quality spectra and prepare them for subsequent multivariate analysis.
Article
Phycocolloids present in three brown (Himanthalia elongata, Bifurcaria bifurcata, Saccharina latissima) and five red edible seaweeds (Mastocarpus stellatus, Gigartina pistillata, Chondracanthus acicularis, Nemalion helminthoides and Dumontia contorta) were studied by FTIR-ATR spectroscopy. Infrared spectra of polysaccharide standards (alginate, agar, iota-, kappa- and lambda-carrageenan) were obtained for comparison. The main polysaccharide found in brown seaweeds was alginate, a linear copolymer of mannuronic (M) and guluronic acid (G). Alginate M/G ratio was tentatively estimated from specific absorption bands (808/787cm−1 and 1030/1080cm−1) in infrared spectra, suggesting higher values of mannuronic than guluronic acid blocks (M/G>1) for brown seaweeds. According to their infrared spectra, all the red seaweeds studied were mainly carrageenan producers. Thus, M. stellatus showed absorption bands at 929.0, 844.7 and 803.2cm−1 of a typical hybrid kappa/iota/mu/nu-carrageenan, meanwhile G. pistillata and C. acicularis, showed the characteristic broad band (830–820cm−1) of lambda-type carrageenan. Moreover, when the second-derivative was obtained to improve resolution of overlapped bands in the original FTIR spectra, this band was divided into several sharper signals, indicating the presence of lambda-/theta-/xi-carrageenans. Accordingly with their FTIR spectra, N. helminthoides contained sulphated polysaccharides, such as carrageenan or mannans, while D. contorta produced lambda- with lesser amounts of kappa-carrageenan. Therefore, FTIR-ATR spectroscopy is proposed as a useful tool for the food, pharmaceutical and cosmetics industry to check the phycocolloid quality of a raw seaweed material by a quick and non-destructive method.
Article
To characterize lipid content of microalgal cells rapidly and accurately, the gravimetric determination, FT-IR and Nile Red (NR) staining were investigated on six typical eukaryotic and prokaryotic algae species. FT-IR and Nile Red were relative quantification methods and a standard curve was required in contrast to the gravimetric method. The FT-IR method determined the lipid, carbohydrate and protein contents simultaneously assuming that the algal cells only consisted of those three components. The Nile Red method was a relatively rapid method for neutral lipid content characterization by spectrofluorometry and could locate lipid body of the algal cell by fluorescence microscopy. According to sample sources and processing purposes, the gravimetric determination was preferable for large-scale cultivation with low-frequency monitoring, while FT-IR and Nile Red were suitable for general laboratory cultivation with medium-frequency monitoring, in particularly Nile Red was appropriate for small samples when high-frequency screening was required.
Article
The separation and estimation of carbohydrates and related polyhydroxy compounds by gas-liquid chromatography of trimethylsilyl (TMS) derivatives is described. The formation of the TMS derivative, in pyridine containing hexamethyldisilazane and trimethylchlprosilane, occurs very rapidly at room temperature so that analyses can be made within a few minutes. Comparative studies of the reaction product of methyl a-glucopyranoside and authentic methyl (tetra-O-trimethylsilyl )-α-glucOpyranoside indicate that silylation of all free hydroxyl groups occurs and that the yield of TMS derivative is virtually quantitative. Conditions are described for chromatography of a wide variety of carbohydrates from C 2 (glycolaldehyde) to C 24 (stachyose) and related substances such as glycosides, deoxysugars, inositols, hexosamines, and N-acetylneuraminic acid. Most of the studies have been made with a silicone column (SE-52) and a polyester column (polyethylene glycolsuccinate) but separations of the TMS derivatives are possible on other polar and non-polar columns. Iso-thermal conditions are usually employed for separations within a narrow range of molecular weight; separations of more complex mixtures, with components of widely differing molecular weights, may be made by linear temperature-programmed analysis. Excellent separations are generally observed with anomeric pairs as well as configurational isomers within a given class such as pentoses, hexoses, disaccharides, etc. The identity of an unknown sugar may be determined by multiple analyses on a number of liquid phases or, alternatively, by analyses of the parent sugar and various derivatives such as methyl glycoside, alcohol, lactone, oxime, and acetal. In all such cases TMS derivatives are prepared prior to gas chromatography. Comparisons are reported for the compositions of aqueous equilibrium solutions of aldoses, by gas Chromatographic analysis, with those reported by measurements of optical rotation and bromine oxidation. Ih several cases unexpected retention times are interpreted in terms of conformational differences of the sugars.
Article
A modified method of preparing highly purified polymannuronic acid and polyguluronic acid was developed wherein acid hydrolysis was carried out using oxalic acid and separation was made using the difference of solubility with pH value. The simplified method using FTIR measurement to estimate the M/G ratio of alginate was investigated using Ca or Mn salts of alginate. The developed method appears to offer enhanced accuracy compared to that of previous techniques. © 2004 Wiley Periodicals, Inc. J Appl Polym Sci 93: 1372–1377, 2004
Article
There has been an increase in the number of colorimetric assay techniques for the determination of protein concentration over the past 20 years. This has resulted in a perceived increase in sensitivity and accuracy with the advent of new techniques. The present review considers these advances with emphasis on the potential use of such technologies in the assay of biopharmaceuticals. The techniques reviewed include Coomassie Blue G-250 dye binding (the Bradford assay), the Lowry assay, the bicinchoninic acid assay and the biuret assay. It is shown that each assay has advantages and disadvantages relative to sensitivity, ease of performance, acceptance in the literature, accuracy and reproducibility/coefficient of variation/laboratory-to-laboratory variation. A comparison of the use of several assays with the same sample population is presented. It is suggested that the most critical issue in the use of a chromogenic protein assay for the characterization of a biopharmaceutical is the selection of a standard for the calibration of the assay; it is crucial that the standard be representative of the sample. If it is not possible to match the standard with the sample from the perspective of protein composition, then it is preferable to use an assay that is not sensitive to the composition of the protein such as a micro-Kjeldahl technique, quantitative amino acid analysis or the biuret assay. In a complex mixture it might be inappropriate to focus on a general method of protein determination and much more informative to use specific methods relating to the protein(s) of particular interest, using either specific assays or antibody-based methods. The key point is that whatever method is adopted as the ‘gold standard’ for a given protein, this method needs to be used routinely for calibration.