Brazilian native passion fruit (Passiflora tenuifila Killip) is a rich source of proanthocyanidins, carotenoids, and dietary fiber.

Passiflora tenuifila is a Brazilian native passion fruit consumed by the local population and is a dietary source of bioactive compounds with potential biological activity. The aim of this study is to evaluate the nutritional value of P. tenuifila fruit and its bioactive compounds at two ripening stages. Three batches of fruit were collected at mature-green and ripe stages, and phenolic compounds, carotenoids, and polyamines were analyzed by HPLC-DAD and LC-MS/MS. The fruit is a good source of dietary fiber. Proanthocyanidin dimers are the major phenolic compounds (up to 84%) at both stages, followed by the C-glycosylated luteolin. Lutein and ß-carotene are the major carotenoids, contributing up to 50% of total carotenoids. The OPLS-DA segregates the mature-green and ripe fruits, as carotenoids are responsible for this separation. In conclusion, passion fruit can be consumed at both stages of maturation without losses of bioactive compound contents or nutritional value.

Addition of either gastric lipase or cholesterol esterase to improve both ß-cryptoxanthin ester hydrolysis and micellarization during in vitro digestion of fruit pulps.

Using the INFOGEST in vitro digestion protocol adapted to carotenoids, the impact of additional rabbit gastric lipase (RGL) on the hydrolysis extent of ß-cryptoxanthin esters was evaluated for the first time, and compared with the addition of porcine cholesterol esterase (CEL). Both the modifications increased the hydrolysis of (all-E)-ß-cryptoxanthin esters from mandarin and peach pulps, although the outcomes were different. Addition of RGL consistently increased the average hydrolysis extent from 55.2% to 59.5% in mandarin pulp and from 22.7% to 48.8% in peach pulp (p < 0.05). The addition of CEL produced lower hydrolysis extents, i.e., 58.5% in mandarin (not statistically significant) and 28.4% in peach (p < 0.05), compared to those obtained with RGL. The hydrolysis extent positively correlated with the carotenoid ester concentration in both matrices. Bioaccessibility values were higher in mandarin pulp (range 32-34%) compared to those in peach pulp (range 16-21%), and were associated with the hydrolysis extent of the carotenoid esters during digestion. Addition of RGL and CEL produced no significant (p < 0.05) effect on the overall carotenoid bioaccessibility values of mandarin, while positively affected those in peach. Altogether these results corroborate that the hydrolysis extent of xanthophyll esters limits bioaccessibility. Additionally, hydrophobicity of the carotenoid inversely correlates with micellarization, as free (all-E)-xanthophylls micellarized in a higher extent compared to (all-E)-ß-carotene and xanthophyll esters. The new information of our results is that the addition of rabbit gastric lipase substantially contributes to the hydrolysis of ß-cryptoxanthin esters from fruit pulps, and consequently, to increase carotenoid bioaccessibility, being even more effective than CEL.

HPLC-PDA-MS/MS as a strategy to characterize and quantify natural pigments from microalgae.

Interest in pigment composition of microalgae species is growing as new natural pigments sources are being sought. However, we still have a limited number of species of microalgae exploited to obtain these compounds. Considering these facts, the detailed composition of carotenoids and chlorophylls of two species of green microalgae (Chlorella sorokiniana and Scenedesmus bijuga) were determined for the first time by high-performance liquid chromatography coupled to diode array and mass spectrometry detectors (HPLC-PDA-MS/MS). A total of 17 different carotenoids were separated in all the extracts. Most of the carotenoids present in the two microalgae species are xanthophylls. C. sorokiniana presented 11 carotenoids (1408.46 µg g-1), and S. bijuga showed 16 carotenoids (1195.75 µg g-1). The main carotenoids detected in the two microalgae were all-trans-lutein and all-trans-ß-carotene. All-trans-lutein was substantially higher in C. sorokiniana (59.01%), whereas all-trans-ß-carotene was detected in higher quantitative values in S. bijuga (13.88%). Seven chlorophyll compounds were identified in both strains with different proportions in each species. Concentrations of chlorophyll representing 7.6% and 10.2% of the composition of the compounds present in the biomass of C. sorokiniana and S. bijuga, respectively. Relevant chlorophyll compounds are reported for the first time in these strains. The data obtained provide significant insights for microalgae pigment composition databases.

Bile amount affects both the degree of micellarization and the hydrolysis extent of carotenoid esters during in vitro digestion.

Carotenoid esters are present in considerable amounts in most fruits, such as in citrus. Although the bioavailability of carotenoid esters is similar or even higher compared to that of free carotenoids, these molecules are generally detected only in the free form in human plasma, suggesting that hydrolysis of carotenoid esters occurs in vivo. However, the available in vitro digestion methods were not able to achieve satisfactory carotenoid ester hydrolysis so far. As bile salts play an essential role in the hydrolytic action of lipolytic enzymes from pancreatin, we evaluated the effect of increasing the bile extract/food ratio from 0.045 to 0.12 (g g-1) on the hydrolysis of ß-cryptoxanthin esters from mandarin pulp during in vitro digestion. Additionally, considering the positive effect of lipids on carotenoid bioavailability, the impact of soybean oil addition on carotenoid ester hydrolysis was studied. Finally, bioaccessibility and recovery of 33 carotenoids were assessed by LC-DAD-MS. The hydrolysis extent of ß-cryptoxanthin esters enhanced from 29% to 55% by increasing the bile extract/food ratio, but reduced respectively to 28% and 11% by the addition of 1% and 10% oil (p < 0.05). The bioaccessibility of overall carotenoids improved from 19% to 35% by increasing the bile extract/food ratio, along with that of (all-E)-ß-carotene (from 19 to 31%) and total (all-E)-ß-cryptoxanthin (17% to 49%). Soybean oil addition reduced carotenoid micellarization, regardless of the concentration (p < 0.05). Irrespective of the bile extract amount and oil addition, the bioaccessibility of carotenoids was inversely related to its hydrophobicity, with respect to the following ranking: free xanthophylls > carotenes ≥ xanthophyll esters. Altogether, these results indicate that increasing the bile extract amount is a simple and inexpensive option to improve carotenoid ester hydrolysis in in vitro digestion protocols. Additionally, the constant amounts of bile (and possibly enzymes) of static methods, such as INFOGEST, should be further optimized for experiments involving lipid addition in which carotenoid bioaccessibility is evaluated.

Carotenoid biosynthesis and quality characteristics of new hybrids between tangor (Citrus reticulata x C. sinensis) cv. 'Murcott' and sweet orange (C. sinensis) cv. 'Pêra'.

Phenotypic characteristics, as well as the relation between carotenoid accumulation and gene expression during ripening were determined in fruits of five new hybrids between tangor cv. 'Murcott' and sweet orange cv. 'Pêra'. The genotypes were classified into the orange-like group, showing mainly epoxycarotenoids, oval fruit shape and yellowish color, or in the mandarin-like group, showing mainly ß-cryptoxanthin, flattened shape and deep-orange coloration; although some hybrids presented intermediate characteristics. The diversity in carotenoid composition of hybrids and genitors were mostly explained by patterns of gene expression. High carotenoid (250-426 µg/g dry weight [dw]) and ß-cryptoxanthin (81-125 µg/g dw) contents, observed in the mandarin-like group, were generally associated with high expression of upstream genes (GGPPS1, PSY, PDS). On the other hand, low expression/repression of these genes and high expression of downstream genes (BCHX and ZEP) were associated with low carotenoid (~158 µg/g dw) and ß-cryptoxanthin (5-22 µg/g dw) contents and epoxycarotenoid accumulation, as occurred in the orange-like group. Breeding experiments resulted in hybrids with outstanding higher carotenoid contents than both genitors (up to 426 µg/g dw versus 158-250 µg/g dw in genitors), which was attributed to transgressive segregation. Differences among genotypes have great impact on commercial fruit quality and potential health benefits, such as the provitamin A content.

Impact of in vitro digestion phases on the stability and bioaccessibility of carotenoids and their esters in mandarin pulps.

The composition of carotenoids (carotenes and free and acylated xanthophylls) and their bioaccessibilities were determined for the first time in pulps of mandarins cultivated in Brazil. Two cultivars of mandarin, Citrus reticulata Blanco cv. 'Ponkan' and Citrus reticulata × C. sinensis cv. 'Murcott', showed higher contents of most carotenoids compared to those found in C. deliciosa Tenore cv. 'Rio'. The major carotenoids in mandarin cv. 'Ponkan' and 'Murcott' were (all-E)-ß-cryptoxanthin laurate (19-21%), (all-E)-ß-cryptoxanthin myristate (15-17%) and (Z)-ζ-carotene (7-12%), followed by (all-E)-ß-cryptoxanthin palmitate (4-7%), free (all-E)-ß-cryptoxanthin (5-6%) and (all-E)-ß-carotene (4-5%), while in mandarin cv. 'Rio' (all-E)-ß-cryptoxanthin myristate (22%) was the major compound, followed by (all-E)-ß-cryptoxanthin laurate (16%), (all-E)-ß-cryptoxanthin palmitate (11%), (all-E)-ß-cryptoxanthin (9%) and (all-E)-ß-carotene (6%). After in vitro digestion, the qualitative carotenoid profile of the supernatant containing the micellarized carotenoids was similar to that of fresh fruits, but the contents were significantly lower. Carotenoid and mandarin physico-chemical properties influenced the bioaccessibility of carotenoids. Free (all-E)-ß-cryptoxanthin showed the highest bioaccessibility in all mandarin cultivars (33-42%), while the bioaccessibilities of ß-carotene (16-36%) and the major carotenoid esters (18-33%) were lower. The overall recovery of carotenoids during in vitro digestion was around 98% after the oral phase, 79% after oral + gastric phases and 77% after oral + gastric + duodenal phases, with free (all-E)-ß-cryptoxanthin and (all-E)-ß-carotene being the most stable ones. Besides possible E-Z isomerization and ester hydrolysis, evident losses occurred in total carotenoid contents and also in the most individual carotenoids and they were not compensated for by the former reactions.

Carotenoid profile of three microalgae/cyanobacteria species with peroxyl radical scavenger capacity.

Carotenoids from cyanobacteria Aphanothece microscopica Nageli and green microalgae Chlorella vulgaris and Scenedesmus obliquus were identified. The total carotenoid content, based on dry weight of biomass, of A. microscopica Nägeli, C. vulgaris and S. obliquus were 1398.88µg/g, 1977.02µg/g and 2650.70µg/g, respectively. A total of 23 different carotenoids were separated in all the extracts, the major ones being all-trans-ß-carotene (29.3%) and all-trans-lutein (28.1%) in Scenedesmus; all-trans-echinenone (22.8%) and all-trans-ß-carotene (17.7%) in Chlorella; all-trans-echinenone (28.3%) and all-trans-ß-carotene (26.2%) in Aphanothece. The carotenoid extracts were shown to be a potent scavenger of peroxyl radical, with values of 31.1 (Chlorella), 14.0 (Scenedesmus) and 7.3 (Aphanothece) times more potent than α-tocopherol.

Carotenoid esters in foods - A review and practical directions on analysis and occurrence.

Carotenoids are naturally found in both free form and esterified with fatty acids in most fruits and some vegetables; however, up to now the great majority of studies presents data on carotenoid composition only after saponification. The reasons for this approach are that a single xanthophyll can be esterified with several different fatty acids, generating a great number of different compounds with similar chemical and structural characteristics, thus, increasing the complexity of analysis compared to the respective saponified extract. This means that since UV/Vis spectrum does not change due to esterification, differentiation between free and acylated xanthophylls is dependent at least on elution order and mass spectrometry (MS) features. The presence of interfering compounds, especially triacylglycerides (TAGs), in the non-saponified extract of carotenoids can also impair carotenoid ester analyses by MS due to high background noise and ionization suppression since TAGs can be present in much higher concentrations than the carotenoid esters. This leads to the need of development of new and effective clean-up procedures to remove the potential interferents. In addition, only few standards of xanthophyll esters are commercially available, making identification and quantification of such compounds even more difficult. Xanthophyll esterification may also alter some properties of these compounds, including solubility, thermostability and bioavailability. Considering that commonly consumed foods are dietary sources of xanthophyll esters and that it is the actual form of ingestion of such compounds, an increasing interest on the native carotenoid composition of foods is observed nowadays. This review presents a compilation of the current available information about xanthophyll ester analyses and occurrence and a practical guide for extraction, pre-chromatographic procedures, separation and identification by liquid chromatography-tandem mass spectrometry (LC-MS/MS).

Composition by LC-MS/MS of New Carotenoid Esters in Mango and Citrus.

Interest in the composition of carotenoid esters of fruits is growing because esterification may affect their bioavailability. Thus, the aim was to provide a detailed identification of carotenoid esters in citrus and mango. Orange cv. 'Valencia' and cv. 'Pera' presented 9 free carotenoids, 38 monoesters, and 60 diesters. Violaxanthin and luteoxanthin derivatives were the major ones, followed by antheraxanthin, lutein, zeaxanthin, ß-cryptoxanthin, and zeinoxanthin esters, many of them reported for the first time in orange pulp. The carotenoid ester composition of tangor cv. 'Murcott', reported for the first time, showed 8 free carotenoids, 34 monoesters, and 33 diesters, with ß-cryptoxanthin esters as major compounds, followed by violaxanthin and zeaxanthin esters. In citrus, carotenoids were acylated mainly with capric, lauric, myristic, myristoleic, palmitic, palmitoleic, and oleic acids. In mango, 5 free carotenoids, 2 monoesters, and 19 diesters were identified, from which many violaxanthin and neoxanthin esters were reported for the first time.

Extração de gelatina a partir das peles de cabeças de carpa comum / Gelatin extraction from skins of common carp heads

Neste estudo, foi avaliada a extração de gelatina a partir de cabeças de carpa comum (Cyprinus carpio) provenientes da industrialização. Para tanto, foram estudados os efeitos da concentração alcalina, o tempo de tratamento e troca de solução alcalina na etapa de tratamento das cabeças de carpa. Foi utilizado um delineamento experimental em estrutura fatorial completa com três fatores em dois níveis de variação. Os fatores de estudo foram concentração de solução de NaOH (3-4mol L-1), tempo de tratamento (45-105min) e troca de solução alcalina no tratamento, tendo como respostas rendimento em gelatina e força do gel. A condição mais adequada para se obter gelatina com bom rendimento (1,98 por cento) e força do gel (240,3g) foi utilizando solução alcalina 3mol L-1, tratamento de 105min e com troca de solução de NaOH.

The aim of the present research was to study the gelatin extraction from skin of common carp heads (Cyprinus carpio). The effects of alkaline concentration, pre-treatment time and change of alkaline solution in pre-treatment step of carp heads were evaluated. It was used a complete experimental design with three factors and two variation levels. Pre-treatment time (45-105min), concentration of alkaline solution (3-4mol L-1) and pre-treatment with change of alkaline solution were chosen as independent variable and gelatin yield and gel strength were the response variable. The best condition to obtain gelatin with good yield (1.98 percent) and gel strength (240.3g) was using alkaline solution 3mol L-1, pre-treatment time of 105min and with one change of NaOH solution.

Eggs fertility and biological aspects of the life cycle in natural populations of Simulium (Diptera, Simuliidae)

Eggs fertility rate and some biological aspects of the immature forms were studied on six Simulium species from ovipositions collected in natural breeding places located in rural area. The fertility was considerably high. For a total of 213 ovipositions containing 74,906 eggs, the average fertility rate was 99.13 percent with variation from 94 to 100 percent. The developmental cycle of the immature forms was observed from egg eclosion to adult emergence under temperature from 16.5 to 28°C. In S. orbitale the life cycle duration was 30 to 32 days, for S. perflavum 32 to 35 days, for S. subnigrum 33 to 39 days, for S. inaequale 41 to 47 days, for S. pertinax 42 to 48 days, and finally, the longest duration was observed for S. distinctum to range 48 to 49 days.

Foram estudados o índice de fertilidade de ovos de espécies de Simulium e o ciclo biológico das fases aquáticas em posturas trazidas de criadouro natural localizado na área rural. A fertilidade de ovos mostrou-se altíssima, das 213 posturas coletadas, contendo 74.906 ovos, o índice médio percentual geral foi de 99,13 por cento. Entre as diferentes posturas, a variação oscilou de 94 por cento a 100 por cento. O ciclo biológico a partir da incubação do ovo até a emergência de adulto em Simulium orbitale foi de 30 a 32 dias; Simulium perflavum de 32 a 35 dias; Simulium subnigrum de 33 a 39 dias, Simulium inaequale de 41 a 47 dias; Simulium pertinax de 42 a 48 dias e Simulium distinctum de 48 a 49 dias em temperaturas com variação de 16,5°C a 28°C.

Levantamento das espécies de Simulium (Diptera, Simuliidae), Riacho dos Padres, município de Almirante Tamandaré, Paraná, Brasil / Survey of the Simulium species (Diptera, Simuliidae), Riacho dos Padres, Almirante Tamandaré municipality, Paraná, Brazil

De janeiro a agosto de 2002, quinzenalmente, foi levada a efeito a coleta de simulídeos imaturos em substratos naturais e antrópicos no Riacho dos Padres. O trabalho visou proceder ao inventário das espécies e verificar as freqüências relativas. A área de propriedade particular localizada na Região Metropolitana de Curitiba constitui uma microempresa de agronegócio, com criação de peixes, coelhos, ovinos, bovinos, aves domésticas e, ainda, área de lazer com eqüinos para cavalgada de visitantes. A presença constante de animais hospedeiros e a ração fornecida aos peixes garantiram farta alimentação aos simulídeos, o que favoreceu a explosão desta praga. Em todos os substratos foram coletadas 24.021 pupas e um total de 197.812 larvas. A partir de pupas, foram identificadas com as seguintes freqüências: Simulium inaequale (55,24 por cento), Simulium perflavum (16,81 por cento), Simulium pertinax (13,93 por cento), Simulium orbitale (8,03 por cento), Simulium subnigrum (4,92 por cento), Simulium distinctum (1,03 por cento) e Simulium incrustatum (0,04 por cento). Verificou-se também a quantidade de larvas que colonizaram todos os substratos.

Controle integrado de espécies de Simulium (Diptera, Simuliidae) por Bacillus thuringiensis e manejos mecânicos no riacho e nos vertedouros de tanques de piscicultura, Almirante Tamandaré, Paraná, Brasil / Integrated pest control of Simulium (Diptera, Simuliidae) with Bacillus thuringiensis and mechanical handling in creek and pisciculture spillway, Almirante Tamandaré, Paraná, Brazil

No Riacho dos Padres na Chácara Evíssima, numa microempresa de agronegócio com criação de peixes e animais domésticos, área de lazer e salão de eventos, localizada na Região Metropolitana de Curitiba, os simulídeos em grande proliferação causaram sério impacto com as picadas a tal ponto que dificultaram o convívio habitual. De janeiro a agosto de 2002, coletou-se 24.021 pupas por meio das quais foram identificadas as espécies: Simulium inaequale, Simulium perflavum, Simulium pertinax com 85,98 por cento e Simulium orbitale, Simulium subnigrum, Simulium distinctum e Simulium incrustatum com somente 14,02 por cento de freqüência. O combate a estas populações iniciado em 28 de fevereiro terminou em 08 de agosto de 2002. O controle integrado consistiu na aplicação de biopesticida Bacillus thuringiensis var. israelensis (Bti) e em manejos mecânicos não só removendo os imaturos (ovos, larvas e pupas) por meio de escovação de vertedouros, mas também pela retirada de substratos naturais e antrópicos do leito. A densidade larvária média inicial, antes da aplicação de Bti no leito do riacho (75 m) e no último vertedouro, verificada em substratos antrópicos fitilhos (área 13.500 cm²), foi de 6,7 e 11,5 larvas/cm², respectivamente. Nestes habitats, o controle integrado com Bti de 1,77 a 2,09 mg/litro do ingrediente ativo, suplementado com manejos mecânicos, monitorado em substratos fitilhos (área 20.250 cm²), resultou em redução larvar de 72,61 por cento a 99,97 por cento e de 74,91 por cento a 99,45 por cento, respectivamente, nas vazões de 0,39 a 0,45 e de 0,38 a 0,43 m³/min. No primeiro vertedouro (área 5.110 cm²), porém, só com manejos mecânicos, atingiu-se a redução larvária de 53,81 por cento a 99,59 por cento. O controle efetuado reduziu as picadas hematófagas ao nível esperado, surtindo o efeito positivo almejado.

At Riacho dos Padres, Chácara Evíssima, a small agribusiness venture featuring fish, poultry and cattle farming; a recreation outdoor facility and Event Hall, ubicated in Curitiba's Metropolitan Area, simuliid populations boast high proliferation rates, delivering serious impact to local people with their noxious bites, to such an extent as to impair the daily meetings of people. From January to August 2002, 24,021 pupae were collected from which the following species and respective frequencies were identified: Simulium inaequale (55.24%), Simulium perflavum (16.81%), Simulium pertinax (13.93%), Simulium orbitale (8.03%), Simulium subnigrum (4.92%), Simulium distinctum (1.03%) and Simulium incrustatum (0.04%). Local pest control measures were carried out from the February 28 to August 8, 2002. Main integrated pest control measures taken consisted in the application of biopesticide Bacillus thuringiensis israelensis (Bti) and mechanical handling, not only by removing the immatures (eggs, larvae and pupae) by means of drain brushing, but also by removing natural and artificial substrates from the bed. Larval mean density, monitored at artificial polythene strips substrates (area 13,500 cm2), averaged 6.7 and 11.5 larvae/cm2 before the application of Bti at the creek bed extension 75 m and on the dam spillway, respectively. Larval mass reduction at creek bed and at the spillway with application of Bti from 1.77 to 2.09 mg/litre of active ingredient for one minute exposure time and means of mechanical handlings monitored at polythene strips substrates (area 20,250 cm2) varied, respectively from 72.61% to 99.97% and from 74.91% to 99.45 with outflows from 0.39 to 0.45 and from 0.38 to 0.43 m3/minute. At the one of the dam spillways with area of 5,110 cm2, average larval density in the first mechanic handling was 10.27 larvae/cm2. However, at this spillway, larval reduction by means of mechanical handlings was from 53.81% to 99.59%. Integrated pest ...