Agar polysaccharides from Gracilaria species (Rhodophyta, Gracilariaceae)

Yield, physical and chemical properties of agar from three agarophytes species (Gracilaria gracilis, G. dura and G. bursa-pastoris) were determined. The agar yield from the three species varied significantly (P<0.01). The highest yields of agar (34.8%) and the lowest (30%) were obtained from G. bursa-pastoris and G. gracilis, respectively. Highest gel strength (630+/-15 g cm(-2)) was obtained from agar extracted from G. gracilis and lowest from G. bursa-pastoris (26+/-3.6 g cm(-2)). The values of 3,6-anhydrogalactose were similar for G. gracilis and G. dura and there were no significant differences among the species. The sulfate contents varied significantly (P<0.01) and the higher value was obtained from G. bursa-pastoris. Among the three species, G. gracilis showed superior agar quality than the other two species, hence it can be considered a good potential source for industrial use.     

Chemical composition of agars from a newly reported Japanese agarophyte,Gracilariopsis lemaneiformis

The chemical composition of agars from Gracilariopsis lemaneiformis, newly reported from Japan, was investigated. Native agars were isolated by a sequential extraction of plants in water at 22 °C and 100 °C, and in boiling 20, 40 and 60° ethanol. Agars in each extract were analyzed by chemical methods, ¹H, ¹³C NMR; and IR spectroscopy. The highest yield of agar (total carbohydrate) was obtained from the 40° ethanol extract (55°). Highest sulfate content was attained in non-alkali treated agars extracted with hot water (4.81°, DS 0.2). The 3,6-anhydrogalactose content was highest in the 40° ethanol extract (36.1° in non-alkali treatment, 40.3° in alkali treatment). The highest methoxyl content (6.51°, DS 0.66) was obtained in the 60° ethanol extract. The G. lemaneiformis agar is composed of the biological precursor to agarobiose repeating units and agarobiose containing 6-O-methyl agarobiose and a small amount of 2-O-methyl-α-l-galactopyranose residues. Alkali treatment improved the chemical quality of the agar fractions, which was comparable with Japanese commercial agar and agarose.     

Development of an eco-friendly agar extraction technique from the red seaweed Gracilaria lemaneiformis

The red seaweed, Gracilaria lemaneiformis growing as an aquaculture bioremediator along the coasts of Liaodong Peninsula, China, was investigated for the agar production. An eco-friendly method called agar photobleaching extraction process was developed for the benefit of workers' health and safety of the environment. The native agar (NA), alkali-modified agar (AA), chemical-bleached agar (CA) and photobleached agar (PA), which were extracted using different processes, were evaluated for their physical and chemical properties. The PA showed most desirable performances in terms of gel strength, gelling temperature, sulfate content and 3,6-anhydro-l-galactose content. Among the different processed agars, PA gel strength was 1913 g/cm2, the highest among the different processed agars, which increased 8.6% on the basis of the AA. Further we applied this new technique to extract agars from Gracilaria asiatica, and similar results were obtained with that of G. lemaneiformis. This indicates that the agar photobleaching extraction process is a feasible method for Gracilaria species and has a potential application. During the whole agar photobleaching extraction process the pigment content of G. lemaneiformis declined gradually and the TOC concentration in photobleaching solution increased along with the increase in the irradiation time. The mechanism of agar photobleaching could be elucidated by the photolysis theory.     

Agar quality of commercial agarophytes from different geographical origins: 1. Physical and rheological properties

Six economically important species ofGracilaria, from a number of commercial sources around the world, andGracilariopsis lemaneiformis, collected from two Japanese localities, were used as the sources of raw material for the evaluation of agar quality. Agar-agar was extracted by pretreatment with various concentratrions of NaOH (0%, 3%, 5%, 7%, 10%) incubated at 80 °C for 2 h. Agar yield, viscosity, dynamic gelling and melting temperature and gel texture were determined for 1.5% agar gels. The highest agar yield was obtained fromG. gracilis from Argentina (39.5%), while the lowest was from BrazilianG. gracilis (13.37%). Dynamic gelling temperature was highest in the agar fromG. gracilis from Turkey (59 °C) and lowest in the non-alkali treated agar isolated fromG. edulis from Indonesia (46 °C). Melting temperature ranged from 96 °C in the agars from the JapaneseGracilariopsis andG. chilensis from Chile to 69 °C in the non-alkali treated agar fromG. edulis from Indonesia. In general, all species produced an agar with high gel strength after treatment with 5% NaOH, except forG. chilensis and the twoGracilariopsis species, which produced an agar with high gel strength after treatment with 3, 7 and 10% NaOH. The highest gel strength (2056 ± 13.6 cm^–2) and hardest gel (261 ± 19.89 g mm^–2) were obtained fromG. lemaneiformis from Japan (Oita Prefecture) after treatment with 7 and 10% NaOH respectively. The lowest gel strength (351 ± 93 cm^–2) was obtained fromG. gracilis from Brazil after treatment with 3% NaOH. The softest gel (66.31 ± 9.63 g mm^–2) was isolated fromG. tenuistipitata from China, after treatment with 3% NaOH. The most flexible gel (11.62 ± 0.31 g mm^–2 × 10²) was obtained fromG. chilensis from Chile after treatment with 3% NaOH.Author for correspondence     

Influence of alkali treatment on agar from Gracilaria cornea from Yucatán, México

The effect of alkali treatments on the yield, rheological and chemical properties of agar from Gracilaria cornea growing along the Yucatáncoast were studied in order to evaluate its potential for industrial use inan attractive economic standpoint. Alkali treatment was carried out with NaOH concentrations of 0.5%, 1%, 3% and 5% in a water bath at 80, 85 and 90 °C. Agar yield, gel strength, gelling and melting temperatures, sulphate, 3,6-anhydro-galactose and ash content weredetermined. The different combinations of NaOH concentration and treatment temperature strongly influenced agar characteristics. There was a variation in the agar content for all NaOH treatments and temperature combinations, ranging between 14.5% to 22.1%. Although the yields obtained for 0.5% NaOH at all temperatures and 1% NaOH at 80 and 85°C were higher than those required by the industry, the physical and chemical characteristics of the agar were similar to those obtained fornative agar from the same species. The gel strengths, sulphate content and gelation hysteresis obtained with agar from the 1% NaOH treatment at 90 °C are in the range required by the food industry. Treatments with 3% and 5% NaOH at all temperatures improved significantly the agar quality giving higher gel strengths (974–1758 g cm ^-2) than those reported for other Gracilaria species. This revised version was published online in August 2006 with corrections to the Cover Date.     

Effects of sulfate starvation on agar polysaccharides of Gracilaria species (Gracilariaceae,Rhodophyta) from Morib,Malaysia

The effects of sulfate starvation on the agar characteristics of Gracilaria species was investigated by culturing two red algae from Morib, Malaysia, Gracilaria changii and Gracilaria salicornia in sulfate-free artificial seawater for 5 days. The seaweed samples were collected in October 2012 and March 2013, periods which have significant variation in the amount of rainfall. The agar yields were shown to be independent of sulfate availability, with only 0.60–1.20 % increment in treated G. changii and 0.31–1.40 % increment in treated G. salicornia while their gel strengths did not increase significantly (approximately 5–7 %) after sulfate starvation for both species. The gelling and melting temperatures did not vary between control and treated samples from both species, except for the treated G. changii collected in March 2013. The gel syneresis index of G. salicornia collected in March 2013 increased significantly after sulfate deprivation. Sulfate starvation introduced some variations in the content of 3, 6-anhydrogalactose and total sulfate esters, but the changes did not have a pronounced effect on the physical properties of agar.     

Chemical characteristics and gelling properties of agar from two Philippine Gracilaria spp. (Gracilariales, Rhodophyta)

The chemical structure of agars extracted from Philippine Gracilaria arcuata and G. tenuistipitata were determined by NMR and infrared spectroscopy. Agar with alternating 3-linked 6-O-methyl-β-D-galactopyranosyl and 4-linked 3,6-anhydro-2- O-methyl-α-L-galactopyranosyl units was isolated from G. arcuata, while the agar from G. tenuistipitata possesses the regular agarobiose repeating unit with partial methylation at the 6-position of the D-galactosyl residues. Both agars exhibit sulphate substitution at varying positions in the polymer. Chemical analyses reveal higher 3,6-anhydrogalactose and lower sulphate contents in alkali-modified than in native agar from both samples. Also, alkali modification enhanced agar gel strength and syneresis. Native G. arcuata agar produces a viscous solution (2000 cP at 75 °C) with a high gelling point (>60 °C) that forms a soft gel even after alkali modification (gel strength: <300 g cm⁻²). On the other hand, the agar from G. tenuistipitata exhibits gel qualities typical of most Gracilaria agars. This revised version was published online in June 2006 with corrections to the Cover Date.     

Agars from three species of Gracilaria (Rhodophyta) from Yucatán Peninsula

Gracilaria cervicornis, Gracilaria blodgettii and Gracilaria crassissima growing along the coasts of Yucatán were investigated for their agar content. The effect of different concentrations of NaOH in the alkali treatment was evaluated. The three species of Gracilaria produced agars, both native and alkali treated, with different properties confirming the heterogeneity of the agar polymers in this genera. G. cervicornis produces agar polymers with an occurrence of methoxylation and sulphation at the C-6 of the beta-D-galactose residues, and with an extra methylation due to the presence of the 4-O-methyl-alpha-L-galactose residue. The presences of these residues is responsible for the extremely poor gelling ability of its agarocolloids, whose commercial value seems to be quite low. Agar extracts from G. blodgettii showed the typical pattern of unsubstituted agar with a very low degree of methylation on both galactose residues. The discrepancy found between sulphate content and NMR data of agar from this species requires a more detailed structural investigation. G. crassissima produces a good quality agar after sulphate precursor sequences have been removed by alkaline treatment, and it may be considered for exploitation as a source of commercial grade agar. Alkali treatment was effective both in removing alkali-labile sulphate and increasing the gel strength in G. crassissima but not in G. cervicornis and G. blodgettii.     

Yield, chemical composition and gel strength of agarocolloids of Gracilaria gracilis, Gracilariopsis longissima and the newly reported Gracilaria cf. vermiculophylla from Roscoff (Brittany, France)

Agarocolloids were extracted from field samples of Gracilaria gracilis, Gracilariopsis longissima and the newly reported Gracilaria cf. vermiculophylla harvested at different periods of the year near Roscoff (France). Native and alkali modified extracts were characterized by GLC, HPLC and FT-IR spectroscopy. The main components of agarocolloids isolated by freeze-thawing method, were 3,6-anhydrogalactose and galactose. In addition, minor components (6-O-methyl-galactose, 4-O-methyl-galactose and sulfate groups ranging from 4.4 up to 6.6% [w/w]) were detected. The highest rate of 6-O-methylgalactose was observed in agarocolloids from vermiculophylla (14 mole%). Sulfates were mainly branched on C4 of the D-galactose in gracilis and Gs. longissima agarocolloids. G. vermiculophylla agaroids isolated by EtOH and NaCl precipitations from the syneresis water were characterized by a high sulfation on C6 of galactose and a low sulfation on C2 of 3,6-anhydrogalactose. Native agarocolloid gel strengths from Gracilaria species were clearly higher than those of Gracilariopsis. Alkali treatments reduced the sulfate levels but increased slightly the gel strengths. An approximation of the polymer sizes carried out with colorimetric assays indicated that the polymer sizes were higher in G. gracilis than observed in Gs. longissima. This revised version was published online in June 2006 with corrections to the Cover Date.     

Agar properties of two species of Gracilariaceae from the Gulf of California,Mexico

Agar properties of two potentially commercial important seaweeds from the Gulf of California were studied. Maximum yield in Gracilaria vermiculophylla (45.7%) occurred during the summer months, coinciding with high water temperatures (31°C) whereas minimum yields (11.6%) were obtained during the coldest months of the year when populations of this species diminish in the bay. Gracilariopsis longissima showed two yield peaks, one in spring and another in fall, before the maximum and minimum seawater temperatures. Gel strength in native agar from the two species was low (<22.5 g cm⁻²) for most of the year. G. vermiculophylla native agar showed a slight increase in gel strength from June to August, which were the hottest months. Maximum value was 85 g cm⁻¹ in August. Maximum gel strength in G. longissima was observed in October (91 g cm⁻¹), and an unusual native agar with no detectable gel strength was observed in March and April samples. Gelling temperatures range from 27.7 to 36.5°C in G. vermiculophyla and from 26.6 to 34.9°C in G. longissima, meanwhile melting points were 73.9 – 53.5°C and 75.5 – 56.6°C, respectively. Sulfate content was high, 6.3–13.9% in G. vermiculophylla and 1.9–11.9% in G. longissima, and on the other hand 3,6 anhydrogalactose content was low 12.1–26.7% and 9.1–23%, respectively compared to other species. Results obtained showed that mean native agar yields of Gracilaria vermiculophylla and Gracilariopsis longissima from the Gulf of California are comparable to other tropical Gracilaria. However, the low gel strength, high sulfate content and low 3,6 anhydrogalactose content observed in the native agar extracted from these species make this an agaroid, thus alternative methods of extraction should be used to evaluate the possibility of commercial utilization of both species.     

Agars fromGracilaria chilensis (Gracilariales)

The chemical composition of the agars extracted with water at 95 °C fromGracilaria chilensis of Estero de Lenga (43.2% of yield) and of Bahia de Coquimbo (43.4% of yield) were determined. The gel strength, gel melting and gelation temperature of the agars were also measured. Polysaccharides obtained by extraction with cold water and by extraction of the alkali pre-treatedG. chilensis of Bahia de Coquimbo were also analyzed. The physical constants and the contents of sulfate and 3,6-anhydrogalactose of the alkali-modified agar fromG. chilensis of Estero de Lenga were comparable to those of commercial agar. This algae represents an excellent potential commercial resource of agar.G. chilensis of Bahia de Coquimbo gave hydrocolloids with low gel strength values more suitable for use in the food industry. Anion exchange chromatography of the agars obtained by extraction with water at 95 °C indicated that the whole polysaccharides and all of the acidic fractions were not homogeneous. Neutral fractions comprised only about 25% of the agars.     

Superior quality agar from <Emphasis Type="Italic">Gracilaria</Emphasis> species (Gracilariales,Rhodophyta) collected from the Gulf of Mannar,India

Gracilaria edulis, G. crassa, G. foliifera, and G. corticata are naturally occurring agarophytes of Indian waters. These agarophytes were evaluated for their agar contents using an improved process recently reported by us (US Patent 2005/0267296A1). The effect of different concentrations of NaOH in the alkali treatment was studied for optimizing the extraction conditions. These Gracilaria species of Indian waters produced agars, both native and alkali treated, with different properties confirming the heterogeneity of the agar polymers in this genera, as one would expect. Among these, G. edulis and G. crassa produced agar polymers having high gel strengths of 490 ± 8.16 and 800 ± 15.4 g cm⁻², respectively, with 8% NaOH treatment as opposed the low gel strength agars that have been reported in the literature to date.     

Comparison of the photosynthetic efficiency,agar yield,and properties of <Emphasis Type="Italic">Gracilaria salicornia</Emphasis> (Gracilariales,Rhodophyta) with and without adelphoparasite

The red seaweed Gracilaria salicornia is widely distributed along the Gulf of Thailand and Andaman sea coast. In nature, the adelphoparasite Gracilaria babae is mostly found to grow on this alga, and it appears as a gall with dark green or yellowish orange color. This study was conducted to examine the photosynthetic efficiency, agar yield, and properties of G. salicornia in plants with (PGs) and without the adelphoparasite (Gs). The results showed that the photosynthetic efficiency of the PGs and Gs was influenced by environmental factors. The agar yield extracted from the Gs was mostly lower than that from the PGs. The gelling temperature of agar solution from the PGs was lower than in the Gs but was not significantly different (p > 0.05). The gel melting point and the viscosity of the agar solution of the PGs was significantly higher than those of the Gs (p < 0.05). The gel strength and total carbohydrate content of the PGs agar were higher than those obtained from the Gs agar, except for the sample collected in June. The content of 3,6-anhydrogalactose was higher in the PGs than in the Gs collected in February and August, when the PGs agar had greater gel strength. Similarly, the PGs agar had a higher sulfate content than did the Gs agar. This study showed that changes in the photosynthetic efficiency, yield, and properties of agar extracted from G. salicornia were influenced by the environmental factors in association with the presence of the adelphoparasite G. babae.     

Gracilaria species (Gracilariaceae, Rhodophyta) from southeastern Australia, including a new species, Gracilaria perplexa sp. nov.: Morphology, molecular relationships and agar content

Select species of the agarophyte Gracilaria were studied from southeastern Australia. The morphology and anatomy of species is described and molecular relations are inferred based on plastid and mitochon‐drial DNA sequence data. Agar yields and qualities are determined for each species. Gracilaria chilensis, found in Tasmania and Victoria, is morphologically and molecularly similar to G. chilensis from New Zealand and Chile and has low agar yields of 11–16%. Gracilaria cliftonii from Victoria, has high crude agar yield (52%) and is molecularly uniform. Gracilaria perplexa sp. nov., known only from Botany Bay, New South Wales, has an agar yield of 39%. The agar of G. perplexa is unusual in requiring the addition of 0.1 mol L^?1 NaCl for alcohol precipitation and is cold‐water (25°C) soluble because of the very high sulfate ester content. Molecular phylogeny shows that G. perplexa is closely related to Gracilaria preissiana from western Australia, but differs from the latter in its reduced branching and narrower more terete axes.     

Optimization of the yield and quality of agar from Gracilariopsis lemaneiformis (Gracilariales) from the Gulf of California using an alkaline treatment

The effect of several alkali treatments on the yield, gel strength, rheology, and chemical characteristics (quality) of the agar obtained from Gracilariopsis lemaneiformis from the Gulf of California was analyzed using different alkali concentrations, temperatures and treatment times. In the first stage of the experiment, all treatments lasted 60 min and the NaOH concentrations (2.5, 3.0, 4.0, 5.0, 6.0%) and temperature (80, 90, 100°C) varied. At constant time, temperature played the predominant role, promoting an increase in agar gel strength. Based on the best treatment conditions found (4% and 5% NaOH, and 90°C and 100°C temperature), in the second stage different treatment times (15, 30, 60, 90, 120 min) were used. Since agar yields were not significantly different among temperatures and times, the optimal conditions to obtain best quality agar were those providing the highest gel strength. Treatment time played an important role in increasing gel strength. Maximum gel strength (Nikan, 954 g cm⁻²) was obtained with 5% NaOH at 100°C after 90 min of treatment, though these conditions resulted in an agar yield reduction of 25.5% relative to native agar. This treatment proved to efficiently yield G. lemaneiformis agar that will meet the commercial quality requirements regarding gel strength, 3,6 anhydrogalactose and sulfate content, as well as rheology and hysteresis. Enrique Hernández-Garibay holds a CONACyT scholarship.     

Polysaccharides from the red seaweed Gracilaria dura (Gracilariales, Rhodophyta)

The yield and physical and chemical properties of agars from Gracilaria dura (C. Agardh) J. Agardh, harvested in Thau lagoon (Mediterranean sea, France), were investigated. The agar yield ranged from 32% to 35%. Gel strength of agar ranged from 263 to 600 g cm(-2), with the maximum observed in October. A positive correlation was found between agar yield and gel strength (r = 0.82; P < 0.01). The gelling temperature followed the same pattern of gel strength and also showed higher value in October (43 degrees C). The nitrogen content varied from 1.04+/-0.60% (June) to 4.70+/-0.01% (October). A positive correlation was noted between nitrogen content and gel strength (r = 0.77; P < 0.05). The 3,6-anhydrogalactose content ranged from 0.70 to 0.84 and showed monthly significant differences (P < 0.05). There was a positive correlation between 3,6-anhydrogalactose content and gel strength. The values of sulfate content were relatively constant during the studied period and no significant differences were observed. The relative high gel strength indicates that this species may be considered as source of agar for commercial use.     

Comparison of three gracilarioids: growth rate, agar content and quality

Three gracilarioid species, Gracilariopsis bailiniae and Gracilaria tenuistipitata from Vietnam and Gracilaria gracilis from Russia, were studied in order to determine whether Gracilaria gracilis might be a superior species for cultivation in brackish-water ponds for agar production compared with the Vietnamese species. The effects of different salinity levels on the growth rate and agar production as well as agar properties of three gracilarioid species were compared in controlled laboratory experiments. Gracilaria tenuistipitata and G. gracilis were tolerant to low salinity (∼10‰), whereas Gp. bailiniae died under these conditions. G. tenuistipitata showed superior growth among the three species examined. Gracilaria gracilis had the highest agar content [36.8–46.6% dry weight (dw)]. Agar yield from Vietnamese gracilarioids did not exceed 30% dw. Gel strength of native agar from Gracilaria gracilis was two-fold higher that from Vietnamese species (278 g cm⁻² vs 130 g cm⁻²). Alkali pretreatment increased gel strength significantly for Gracilaria gracilis (1.4-fold), and G. tenuistipitata and Gp. bailiniae (2.3-fold) compared with native agar. The results suggest that Gracilaria gracilis may be a suitable species for production of reasonably good quality agar.     

Isolation,chemical investigation and antiviral activity of polysaccharides from Gracilaria corticata (Gracilariaceae,Rhodophyta)

Polysaccharides were sequentially extracted from the agarophyte Gracilaria corticata. Chemical analysis combined with infrared spectroscopy showed that the cold water extracted material consists mainly of a high molecular weight sulfated galactan. Most of the sulfate groups are alkali labile and are located at C-4 of the 1,3-linked D-galactose units and C-6 of the 1,4-linked L-galactose residues. The autoclaved extracts contain agar type polysaccharide having a high pyruvate content and a variable degree of methylation, but were contaminated with floridean starch. 1H-NMR studies indicate that methoxyl groups, when present, occur at C-6 of the 1,3-linked D-galactose units and C-2 of the 1,4-linked L-galactose residues of agar polymer. Bioassays showed that a high molecular weight galactan sulfate, exhibited selective antiviral activity against herpes simplex virus types 1 and 2, likely due to an inhibition of the initial virus attachment to the host cell.     

Seasonal variation of the agar quality and chemical composition of Gracilaria veleroae and Gracilaria vermiculophylla (Rhodophyceae,Gracilariaceae) from Baja California Sur,Mexico

Yield and physico‐chemical properties of agar from Gracilaria veleroae E.Y. Dawson and Gracilaria vermiculophylla (Ohmi) Papenfuss were studied and the chemical composition of the two seaweeds was determined. Samples were collected seasonally from summer 2003 to spring 2005. The agar yield did not vary significantly between seasons for both species. The lowest agar gel strength was obtained from G. veleroae (207.5 g cm^–2) in summer 2003 and the highest from G. vermiculophylla (793.1 g cm^–2) in winter 2004. Melting temperatures and hysteresis were higher in G. vermiculophylla, whereas gelling temperatures and 3,6‐anhydrogalactose content were higher for G. veleroae. Moisture, ash, crude fiber, and ether extract showed no significant seasonal variation for G. veleroae. The chemical composition of G. vermiculophylla showed significant seasonal variation. G. vermiculophylla possesses a better agar quality than G. veleroae and is a species that could be considered as a source of agar for commercial use.     

Seasonal changes in the chemical and gelling characteristics of agar from Gracilaria verrucosa collected in Turkey

The chemical and gelling properties of agar from G. verrucosa collected from Izmit bay in Turkey at different months of the year were studied. Purification of agar was performed by using amylase treatment and isopropyl alcohol precipitation. The phycocolloid content was between 24.0–43.0% of the algal dry weight and was maximum in summer collected algae. Relative total sulfate and 3,6-anhydrogalactose content in the agar were determined from the ratios of infrared spectroscopy band intensities at 1250/2920 cm^–1 and 930/2920 cm^–1, respectively. 3,6-anhydrogalactose and sulfate contents were the highest in agar from algae collected from June until November and January until July, respectively. The gel strength of native agar were the highest from in autumn collected algae and increased to about 1250 N m^–2 after alkali treatment. Thus, this study demonstrated that G. verrucosa from Turkey produces an agar with optimal chemical and gelling properties after alkali-treatment in fall and winter collected algae and may be used for industrial agar production.