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LUDMILLA DE CARVALHO OLIVEIRA
ANTIOXIDANT PROPERTIES AND PHYSICAL, SENSORY
AND NUTRITIONAL CHARACTERISTICS OF EXTRUDED
CORN-BASED BREAKFAST CEREAL ELABORATED WITH WHOLE
GRAIN WHEAT FLOUR AND JABUTICABA (Myrciaria cauliflora)
PEEL POWDER
PROPRIEDADES ANTIOXIDANTES E CARACTERÍSTICAS
FÍSICAS, SENSORIAIS E NUTRICIONAIS DE CEREAL MATINAL
EXTRUSADO À BASE DE MILHO ELABORADO COM FARINHA DE
TRIGO DE GRÃO INTEIRO E CASCA DE JABUTICABA (Myrciaria
cauliflora) EM PÓ
CAMPINAS 2015
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UNIVERSIDADE ESTADUAL DE CAMPINAS Faculdade de Engenharia de Alimentos
LUDMILLA DE CARVALHO OLIVEIRA
ANTIOXIDANT PROPERTIES AND PHYSICAL, SENSORY AND NUTRITIONAL CHARACTERISTICS OF EXTRUDED CORN-BASED BREAKFAST CEREAL ELABORATED WITH WHOLE GRAIN WHEAT FLOUR AND JABUTICABA
(Myrciaria cauliflora) PEEL POWDER
PROPRIEDADES ANTIOXIDANTES E CARACTERÍSTICAS FÍSICAS, SENSORIAIS E NUTRICIONAIS DE CEREAL MATINAL EXTRUSADO À BASE
DE MILHO ELABORADO COM FARINHA DE TRIGO DE GRÃO INTEIRO E CASCA DE JABUTICABA (Myrciaria cauliflora) EM PÓ
Thesis presented to the School of Food Engineering of the University of Campinas in partial fulfillment of the requirements for the Ph.D. degree in Food Technology Tese apresentada à Faculdade de Engenharia de Alimentos da Universidade Estadual de Campinas como parte dos requisitos exigidos para a obtenção do título de Doutora em Tecnologia de Alimentos
Supervisor/Orientador: Profa. Dra. Caroline Joy Steel
Este exemplar corresponde à versão final da tese defendida pela aluna Ludmilla de Carvalho Oliveira e orientada pela Profa. Dra. Caroline Joy Steel
_______________________________________________________ ASSINATURA DO ORIENTADOR
CAMPINAS 2015
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Ficha catalográfica
Universidade Estadua de Campinas
Biblioteca da Faculdade de Engenharia de Alimentos Claudia Aparaecida Romano – CRB 8/5816
Informações para Biblioteca Digital
Título em outro idioma: Propriedades antioxidantes e características físicas, sensoriais e
nutricionais de cereal matinal extrusado à base de milho elaborado com farinha de trigo de grão inteiro e casca de jabuticaba (Myrciaria Cauliflora) em pó
Palavras-chave em inglês: Breakfast cereal Cooking extrusion Starch Dietary fiber Phenolic compounds
Área de concentração: Tecnologia de Alimentos Titulação: Doutora em Tecnologia de Alimentos Banca examinadora:
Caroline Joy Steel [Orientador] Maria Teresa Pedrosa Silva Clerici Mario Roberto Marostica Junior Reinaldo Eduardo Ferreira Ana Carolina Conti e Silva
Data de defesa: 26-06-2015
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BANCA EXAMINADORA
Profa. Dra. Caroline Joy Steel Orientadora
Profa. Dra. Maria Teresa Pedrosa Silva Clerici Membro Titular
Universidade Estadual de Campinas
Dr. Mario Roberto Marostica Junior Membro Titular
Universidade Estadual de Campinas
Dr. Reinaldo Eduardo Ferreira Membro Titular
Dedini S/A Indústrias de Base
Profa. Dra. Ana Carolina Conti e Silva Membro Titular
Uninversidade Estadual de São Paulo- IBILCE
Dr. Marcio Schmiele Membro Suplente
Universidade Estadual de Campinas
Dra. Rita de Cássia Salvucci Celeste Ormenese Membro Suplente
Instituto de Tecnologia de Alimentos-CCQA
Pfra. Dra. Thaís de Souza Rocha Membro Suplente
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ABSTRACT
Extrusion is the main technology used to obtain “ready-to-eat” (RTE) breakfast cereals, being starch their main structural component. The process submits the ingredients to
conditions of high temperature, high pressure, low moisture, and shear. Aware of the
growing demand of consumers concerned with their health, studies have shown that is
possible to introduce ingredients that enhance the nutritional and/or functional value of
extruded products and, in the case of breakfast cereals, whole-grain wheat flour and fruit
products are examples with the potential of associating functionality to the product. The
objective of this study was to evaluate the effect of the substitution of corn flour (CF) by
whole-grain wheat flour (WGWF), extrusion temperature and raw-material moisture
content on the technological, sensory and nutritional properties of RTE breakfast cereals.
The breakfast cereals were produced following a 23 central composite rotatabledesign (18 trials), being the independent variables: WGWF ratio, feed moisture and temperature of 2rd and 3th barrel zones. The extruded products were evaluated in terms of physical, chemical and nutritional aspects. The optimum point (OP) was selected, empirically, considering
intermediate sectional expansion, maximum dietary fiber (legislation value), minimum
compression force and maximum crispness, mainly after immersion in milk. In a second
stage, four breakfast cereal formulations were processed at temperature and feed moisture
corresponding to the OP (100°C and 16%, respectively) and with jabuticaba peel powder
(JPP) inclusion, being: (1) 100% CF; (2) 20% CF + 80% WGWF (3) 10% CF + 10% JPP +
80% WGWF e (4) 90% CF + 10% JPP. The breakfast cereals were evaluated in terms of
technological (expansion index, bulk density, dry texture, texture after soaking in milk,
water solubility index and water absorption index), nutritional (total dietary fiber) and
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antioxidant activity of cereals and raw materials were also evaluated by high performance
liquid chromatograph (HPLC) and oxygen radical absorbance capacity (ORAC assay),
respectively. The combination of wholemeal wheat flour with corn flour was a good
alternative for increasing the fibre content of extruded products. The textural properties of
the extrudates, hardness and crispness, were mainly influenced by WGWF and feed
moisture; with changing in properties after soaking in whole milk. In relation to color, the
extrudates elaborated with high amount of WGWF at high temperature or high feed
moisture content was darker. The starch nature as well the fiber presence governed the
starch transformations and interactions (gelatinization, amylose-lipid formation and
retrogradation). The low sugar breakfast cereals from WGWF and JPP showed acceptable
physical and sensory characteristics. The anthocyanins, cyanidin 3-glucoside (cyd-gluc)
and delphinidin 3-glucoside (dpd-gluc), and ferulic acid were the predominant phenolic
compounds in raw materials (JPP and WGWF, respectively), and were also identified in
extrudates, which showed in vitro antioxidant activity
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RESUMO
A extrusão é a principal tecnologia utilizada na obtenção de cereal matinal ready-to-eat
(RTE), sendo o amido o principal constituinte estrutural. O processo ocorre sob condições
de alta temperatura, alta pressão, baixa umidade e cisalhamento. Atentos à crescente
demanda de consumidores preocupados com a saúde, estudos têm mostrado que é possível
introduzir ingredientes na formulação de extrusados que incrementem o seu valor
nutricional e/ou funcional e, no caso de cereal matinal, a farinha de trigo de grão inteiro e
os produtos derivados de frutas são exemplos com potencial de associar funcionalidade ao
produto. O objetivo deste trabalho foi avaliar o efeito da substituição da farinha de milho
(FM) por farinha de trigo de grão inteiro (FTGI), da umidade de alimentação da
matéria-prima e da temperatura de extrusão sobre as propriedades tecnológicas, sensoriais e
nutricionais de cereal matinal RTE. Os cereais matinais foram elaborados seguindo um
delineamento composto central rotacional 23 (18 ensaios), sendo as variáveis independentes: proporção de FTGI, umidade de alimentação da matéria-prima e
temperatura das zonas 3 e 4 da extrusora. Os produtos extrusados foram avaliados quanto
aos aspectos físicos, químicos, nutricionais e sensoriais. O ponto ótimo foi selecionado de
forma empírica, tendo como critérios de definição expansão intermediária, máximo teor de
fibra alimentar (considerando os valores estabelecidos pela legislação), mínima dureza e
máxima crocância, principalmente após imersão em leite. Com base nisso, quatro
formulações de cereal matinal foram processadas nas condições de temperatura e umidade
do ponto ótimo (100°C e 16%, respectivamente) e com inclusão de casca de jabuticaba em
pó (CJP), sendo:(1) 100% de FM; (2) 20% FM + 80% FTGI; (3) 10% FM + 10% CJP +
80% FTGI e (4) 90% FM + 10% CJP. Os cereais resultantes foram igualmente avaliados
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textura em leite, índice de solubilidade em água, índice de absorção de água), nutricional
(fibra alimentar total, perfil de fenólicos e atividade antioxidante) e sensorial (testes de
aceitação e de intenção de compra). O perfil de fenólicos e a atividade antioxidante dos
cereais matinais e matérias-primas foram avaliados por cromatografia líquida de alta
eficiência (CLAE) e pela capacidade de absorbância do radical oxigênio (ORAC). A
combinação de FTGI com FM foi uma boa alternativa para o incremento do teor de fibra
alimentar nos produtos extrusados. As propriedades de textura dos extrusados, dureza e
crocância, foram principalmente influenciadas pela FTGI e umidade de alimentação; com
alterações nas propriedades após imersão em leite. Em relação à cor, os cereais elaborados
com alto teor de FTGI à alta temperatura ou alto teor de umidade foram mais escuros. A
natureza do amido, tão bem como a presença de fibra governou o processo de gelatinização,
a formação do complexo amilose-lipídeo e a retrogradação. Os cereais matinais contendo
FTGI e CJP apresentaram aceitáveis características físicas e sensoriais. Antocianinas,
cianidina 3-glicosídeo e delfinidina 3-glicosídeo, e ácido ferúlico foram os compostos
fenólicos predominantes na CJP e FTGI, respectivamente, e também presentes nos cereais
após extrusão, os quais apresentaram atividade antioxidante in vitro.
Palavras-chaves: cereal matinal, extrusão termoplástica, amido, fibra alimentar, compostos fenólicos.
xi Summary Abstract---vii Resumo---ix Introdução---1 Objetivos---5 Objetivo Geral---5 Objetivos Específicos---5
Paper 1. Whole grains and fruit as ingredients in functional breakfast cereals - Review Abstract---9
Overview of extrusion cooking---10
Extruded breakfast cereals---13
Definition and market---13
Raw materials---15
Extrusion process variables---18
Functional breakfast cereals---19
Dietary fiber---21
Phenolic compounds and antioxidant activity---23
Whole grain wheat flour (WGWF) ---26
Jabuticaba (Myrciaria cauliflora) ---29
Effect of extrusion process on bioactive compounds (fiber and phenolic compounds)31 Future trends---34
References---35
Paper 2. Effect of the addition of whole-grain wheat flour and of extrusion process parameters on dietary fibre content, starch transformation and mechanical properties of a ready-to-eat breakfast cereal Abstract---47
Introduction---48
Materials and Methods---51
Material---51
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Sample preparation and extrusion cooking---52
Particle size distribution of flours and proximate composition of flour and extruded products---53
Pasting properties---55
Cross section image analysis of extruded products---56
Statistical analysis---56
Results and Discussion---57
Dietary fibre (insoluble, soluble and total) ---58
Free sugar, digestible, resistant and total starch---60
Pasting Properties---63
Image analysis---65
Conclusion---71
References---72
Paper 3. Extrusion conditions and whole grain wheat flour effects on instrumental color and bowl-life textural properties of ready-to-eat breakfast cereals Abstract---87
Materials and Methods---91
Raw material---91
Sample preparation and extrusion cooking---91
Moisture and water activity---93
Sectional expansion (SE) ---94
Bulk density (BD) ---95
Instrumental texture of dry breakfast---95
Instrumental texture of bowl-life breakfast cereals---95
Instrumental color---96
Data analysis---97
Results and Discussion---97
Water activity---97
Sectional expansion and bulk density---102
Dry instrumental texture---106
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Instrumental color---114
Conclusions---114
Literature Cited---115
Paper 4. Starch transformations evaluated through thermal and pasting properties of extruded fiber-enriched corn-based breakfast cereals Abstract---125
1. Introduction---126
2. Materials and methods---130
2.1. Materials---130
2.2. Methods---130
2.2.1. Sample preparation---130
2.2.2. Specific mechanical energy---131
2.2.3. Water solubility index (WSI) and water absorption index (WAI) ---132
2.2.4. Pasting properties of unprocessed flours---132
2.2.5. Differential scanning calorimetry (DSC) ---133
2.2.6. X-ray diffraction (XRD) ---133
2.2.7. Statistical analysis---134
3. Results and Discussion---135
3.1. Specific mechanical energy (SME) ---135
3.2. Water solubility index (WSI) and water absorption index (WAI) ---138
3.3. Pasting properties of the raw material blends---143
3.4. Differencial scanning calorimetry---146
3.4.1. Gelatinization of unprocessed flours (UF) and blends---146
3.4.2. Thermal properties of extrudates---150
3.5. X-ray diffraction---152
4. Conclusions---155
References---156 Paper 5. Relationship of sensorial acceptability and physical properties of extruded breakfast cereals enriched with whole grain wheat flour and jabuticaba (Myrciaria cauliflora) peel powder
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Abstract---163
1. Introduction---164
2. Materials and methods---167
2.1. Materials---167
2.2. Methods---167
2.2.1. Jabuticaba peel powder (JPP) preparation---167
2.2.3. Sample preparation and extrusion cooking---168
2.2.4. Water solubility index (WSI) and water absorption index (WAI) ---168
2.2.5. Sectional expansion (SE) ---171
2.2.6. Bulk density (BD) ---170
2.2.7. Instrumental color---171
2.2.8. Mechanical properties---171
2.2.9. Bowl-life---172
2.2.10. Acceptability test and purchase intention---172
2.2.11. Statistical analysis---174
3. Results and Discussion---174
3.1. Proximate composition and particle size distribution of jabuticaba peel powder (JPP) ---174
3.2. Water solubility index (WSI) and water absorption index (WAI) ---175
3.3. Sectional expansion (SE) and bulk density (BD) ---176
3.4. Instrumental color---179
3.5. Mechanical properties---182
3.6. Principal component analysis (PCA) ---185
3.7. Acceptance test---186
4. Conclusion---191
References---192
Paper 6. Bound phenolic content, antioxidant activity and dietary fibre content of extruded breakfast cereals elaborated with whole grain wheat flour and jabuticaba (Myrciaria cauliflora) peel powder Abstract---199
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Materials and methods---203
Materials---203
Methods---203
Jabuticaba peel powder (JPP) preparation---203
Proximate composition of jabuticaba peel powder---204
Sample preparation and extrusion cooking---204
Determination of dietary fibre---205
Extraction of soluble free and insoluble bound phenolics acids from whole grain wheat flour and extrudates---206
Anthocyanin extraction from jabuticaba peel powder---207
Identification and quantification of phenolic compounds (phenolic acids and anthocyanins) in whole grain wheat flour, corn flour, jabuticaba peel powder and extruded breakfast cereals (HPLC-MS) ---207
Determination of total antioxidant activity by ORAC (oxygen radical absorbance capacity) assay---209
Statystical analysis---210
Results and discussion---211
Jabuticaba peel powder (JPP) characterization---211
Total dietary fibre (TDF) in extrudate cereals---212
Phenolic acids profile and quantification in unprocessed whole grain wheat flour, corn flour and extrudate breakfast cereals---214
Anthocyanin profile and quantification in unprocessed jabuticaba peel powder and extrudates breakfast cereals---215
Antioxidant activity---217
Conclusion---219
References---219
Conclusão Geral---225
Appendice and Attachment---229
Appendice 1: Experimental responses of central composite rotatable design (CCRD) ---231
Appendice 2: Optimum point definition---232
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Appendice 4:Ficha do teste de aceitação e intenção de compra de cereal matinal extrusado
---235
Appendice 5: Parecer consubstanciado do CEP---236
Appendice 6: Termo de Consentimento Livre e Esclarecido (TCLE) ---242
Appendice 7: Paper Publication---244
Attachment 1: Phenolic acid identification in whole grain wheat flour---245
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Dedico
Aos meus presentes e mais verdadeiros símbolos de doação e amor: papai (José Carlos), mamãe (Lúcia) e Ximena (irmã).
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"Diga-me, e eu me esqueço. Ensine-me, e eu me lembro. Envolva-me, e eu aprendo." Benjamin Franklin
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Agradecimentos
A minha família, em especial aos meus pais (José Carlos e Lúcia) e a minha irmã (Xi), meus amores, que sempre estiveram presentes na minha caminhada, pelo incentivo, crédito e suporte de todos os momentos, sempre com o imensurável zelo e amor. À vovó Tita, sempre acolhendo com carinho e entusiasmo.
Agradeço a Deus como esteio espiritual, por conduzir os meus caminhos, direcionar as oportunidades, a quem diversas vezes supliquei sabedoria e força (nas dificuldades) e também direcionei meu agradecimento por cada simbólica conquista.
Às especiais amigas-irmãs, assim como carinhosamente me refiro, Elisa Morais (Elisinha), Cinthia Rodrigues (Ci) e Ingrid de Moraes (Did), preciosidades que facilitaram os meus dias, amenizaram a adaptação, foram exemplos de companheirismo, de generosidade; quantos risos compartilhados na euforia dos momentos mais ternos e quantas palavras precisas e o conforto certo quando a dificuldade estava ali presente. Ainda, à amiga Adelaine (Adê) que também esteve presente com todo esse carinho, atenção e torcida.
À família do Laboratório de Cereais, Raízes e Tubérculos (LabCer) que foi o combustível diário e a família “científica”, reunida nas disciplinas, nos desafios experimentais (sem os quais seria impossível o sucesso), nos cafés, nos almoços, nas comemorações de aniversário, nas conversas, nos risos, pelo carinho e amizade. Foram momentos memoráveis ao lado dos colegas e amigos: Cinthia, André, Leandra, Simone, Eveline, Thaís, Georgia, Amanda, Fernanda, Lara, Adriana e Mária. À Alessandra (Alê) pelo apoio nas técnicas das atividades experimentais e pela amizade querida. Ao Marcio (Marcinho), pela imensurável ajuda, pelo ânimo e pela paciência em todos os momentos em que o processo de extrusão entrava em cena. Ao professor Yoon e à professora Maria Teresa Pedrosa, que foram também orientadores ao compartilhar um pouco do conhecimento e experiência.
Aos alunos Edna, Natássia e Lucas que participaram com muito carinho e dedicação em distintas etapas do trabalho. Vocês foram fundamentais. Aos amigos Milene, Janclei, Erick Almeida e Natália Manzatti pela generosidade, por compartilhar o que sabiam e assim contribuindo muito para a evolução do meu trabalho em momentos em que o assunto era desconhecido e até mesmo um desafio. Kazumi (Ka) pela doce parceria-amiga no
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momento em que os meus ânimos pareciam esgotar e a última etapa ainda não estava finalizada.
A minha orientadora Caroline Steel pela orientação, pelas oportunidades concedidas e pelos desafios impostos, explícitos e implícitos, os quais foram chaves para o meu crescimento e para uma visão mais madura da pesquisa.
Ao Instituto de Tecnologia de Alimentos (ITAL)/Centro de Ciência e Qualidade de Alimentos (CCQA) pela parceria e por possibilitar a realização de parte experimental, disponibilizando recursos físicos para a consolidação desta etapa (pesquisadora Rita Ormenese). À técnica Michele Reis pela graciosidade ao me receber, pela presteza e ajuda incondicionais.
Ao Instituto de Agroquímica y Tecnología de Alimentos (Valência, Espanha) que enquanto instituição foi a porta para a realização de um sonho, possibilitando o estágio no exterior (doutorado sanduíche). À supervisora Cristina Molina Rosell pela oportunidade, pelos ensinamentos e pela incrível e inesquecível oportunidade dada. Aos “novos” e especiais amigos de laboratório que durante o estágio no exterior foram uma extensão da família: Georgios Agg, Sebastián Pizarro, Meriem Ouazib, Federico Torturu e Crisy. Às técnicas Adelina e Rachel. À amável “irmanzinha” brasileira que a estadia na Espanha me proporcionou, Rafaela Peixoto (Rafinha), cujo suporte e carinho foram essenciais. Ao pesquisador e, então, amigo Quique pelo crédito no meu trabalho, pela orientação, pelo profissionalismo e incentivo.
À professora Célia Franco por ceder o Laboratório de Amido e Panificação do Ibilce/Unesp para a realização de algumas análises e ao Jefferson Barros pelo acolhimento e ajuda.
À banca examinadora pelas contribuições feitas, contribuindo assim para o enriquecimento da qualidade do trabalho.
À Faculdade de Engenharia de Alimentos da Unicamp pela equipe, comprometimento, qualidade e recursos capazes de influenciar positivamente o meu amadurecimento profissional.
Aos colegas que somaram ao longo dos quatro anos de Unicamp, que foram parceiros de disciplinas, de momentos singulares, que partilharam muitas alegrias, as ansiedades também, que foram ajudas essenciais e que conquistaram meu coração: Adriana
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Reis, Alexandre Azevedo, Ana Laura Tibério, Bianca Costa, Bruno Castro, Bruna Porto, Diana Nunes, Gustavo Pereira, Jéssika Gonçalves, Kazumi Kawazaki, Laura Botti, Leandro Oliveira, Meg Fernandes, Miguel Meireles, Marcelo C., Rita Garcia, Sebastián Rojas, Simone Melo, Talita Almeida, Thales Leandro e Thiago Leite. Muitos destes que por peculiares motivos tornaram-se muito especiais.
A todos que acompanharam e contribuíram.
À CAPES pela bolsa de doutorado concedida para a realização deste trabalho e à Fapesp pelo financiamento do projeto de pesquisa (2012/23281-2). Às empresas, pelas matérias-primas gentilmente doadas para a execução desta pesquisa: Milhão Alimentos (farinha de milho), Fagan propriedade agrícola (jabuticaba) e Domondo (Isomalte®).
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INTRODUÇÃO
Os cereais matinais extrusados ready-to-eat (RTE) são produtos de grande popularidade entre os consumidores e que vêm se adequando aos atuais requerimentos de saudabilidade e conveniência da modernidade, o que reflete a própria definição como “core food”, alimentos adequados ao consumo diário (AUSTRALIAN GUIDE TO HEALTHY EATING, 2010). No caso de cereais matinais expandidos, a matéria-prima base ainda é o milho, em função da tradição e das propriedades de expansão do amido. Assim sendo, a exploração de novas matérias-primas com o objetivo de diferenciação do produto, e de forma a atender os requerimentos do mercado e melhorar a qualidade nutricional, vem crescendo no ramo científico e industrial.
Os cereais integrais têm recebido grande atenção e aos poucos estão substituindo a farinha refinada nas formulações de produtos como pães, biscoitos e cereais matinais, como consequência da elucidação das propriedades funcionais a eles associados e da preocupação dos órgãos regulatórios de saúde em aumentar o consumo de produtos integrais, fontes de fibra alimentar (Food and Agriculture Organization of the United Nations, Food and Drug Administration and Whole Grains Council in the United States of America). A farinha de trigo de grão inteiro ainda é fonte de compostos fenólicos, que apresentam funções fisiológicas e propriedades antioxidantes e estão associados à redução do risco de doenças crônicas como doenças cardiovasculares, diabetes do tipo 2 e alguns tipos de câncer (Slavin, 2003; Liu et al., 1999).
Os cereais contêm uma diversidade de compostos fenólicos, com estruturas químicas diferenciadas, dos quais os ácidos fenólicos são os mais abundantes e de maior significância. Duas sequências de ácidos fenólicos têm sido estudadas: derivados do ácido cinâmico e do ácido benzoico. Em trigo, o ácido ferúlico aparece em maior quantidade, podendo ser encontrado nas formas solúvel livre, solúvel conjugada (ligações ésteres) ou em complexos ligados (Adom & Liu, 2002).
Do ponto de vista tecnológico, a incorporação de fibra alimentar em produtos extrusados expandidos ainda é limitada em função do aspecto negativo sobre as propriedades de expansão, densidade e textura, principalmente (Brennan et al., 2008; Robin
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et al., 2011). Então, os ajustes das condições do processo de extrusão que incluem a umidade de alimentação da matéria-prima, a vazão de alimentação, a temperatura e velocidade da rosca, em associação com a composição da matéria-prima, podem ser cruciais para a obtenção de um produto de qualidade. Além das propriedades físicas citadas, o índice de absorção de água, o índice de solubilidade em água e a energia mecânica do sistema são avaliações normalmente realizadas quando se trata de produtos extrusados. Além disso, a textura (dureza e crocância) é uma das principais características de cereais matinais, os quais são tradicionalmente consumidos imersos em leite. A avaliação de como o leite pode alterar as propriedades de textura é uma informação importante, principalmente quando novas matérias-primas são utilizadas (Pamies et al., 2000; Sacchetti et al. 2003; Sacchetti et al., 2005).
Sendo o amido o principal componente em formulações de cereais, na medida em que outros componentes como fibra alimentar são incluídos, as transformações do amido são alteradas por fatores que vão desde a diluição do amido até a interação amido-fibra-água, quando este é exposto a condições de calor e umidade. Como consequência, as propriedades físicas do produto final são alteradas, sendo a expansão diretamente afetada (Robin et al., 2011). As propriedades de pasta, as propriedades térmicas e o padrão de cristalinidade são normalmente avaliados para verificar as transformações físicas e estruturais do amido ocorridas durante a extrusão quando comparado com a farinha não processada.
Embora ainda pouco utilizadas, as frutas são também ingredientes que podem ser associados aos cereais na elaboração de cereal matinal extrusado, seja no aproveitamento de resíduos, do suco ou de alguma parte específica do fruto (polpa, casca ou semente). As frutas, assim como os cereais integrais, são fontes de fibra e contribuem com o aporte nutricional do produto final. Adicionalmente, algumas delas são fontes de pigmentos naturais, como as antocianinas, com propriedades antioxidantes e com a qualidade de melhorar o aspecto sensorial pela cor atribuída ao produto final (Camire et al., 2002). Camire et al., 2007; Khanal et al., 2009). Desta forma, um cereal matinal funcional contendo tanto cereal quanto fruta, tem um grande potencial de promover benefícios à
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saúde humana como consequência dos benefícios associados às matérias-primas. No entanto, o desafio está em minimizar os efeitos da extrusão, sobretudo da temperatura, sobre a integridade e propriedades dos compostos bioativos.
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OBJETIVOS Objetivo Geral
O objetivo deste trabalho foi avaliar o efeito da substituição da farinha de milho (FM) por farinha de trigo de grão inteiro (FTGI), da umidade de alimentação da matéria-prima e da temperatura de extrusão sobre as propriedades tecnológicas e nutricionais de cereal matinal extrusado RTE (ready-to-eat) de forma a otimizar a formulação e as condições de processamento para a obtenção de um produto final nutricionalmente enriquecido e com qualidade tecnológica. Além disso, foi incorporada casca de jabuticaba (Myrciaria cauliflora) em pó em combinação com farinha de trigo de grão inteiro e/ou farinha de milho.
Objetivos Específicos:
1) Avaliar o efeito da incorporação da FTGI e das variáveis de extrusão (umidade de alimentação da matéria-prima e temperatura) no aspecto nutricional (fibra alimentar, açúcares livres, amido digerível e amido resistente), na estrutura das células e nas propriedades de pasta dos cereais matinais à base de milho (artigo 2).
2) Avaliar o efeito da incorporação da FTGI e das variáveis de extrusão (umidade de alimentação da matéria-prima e temperatura) nos parâmetros de qualidade de cereal matinal extrusado: expansão, densidade, textura a seco e em leite, cor e atividade de água (artigo 3).
3) Avaliar o efeito da incorporação da FTGI e das variáveis de extrusão (umidade de alimentação da matéria-prima e temperatura) na drasticidade do processo de extrusão e no grau de transformação do amido avaliado pela energia mecânica específica do sistema, índice de absorção de água (IAA) e índice de solubilidade em água (ISA), propriedades térmicas e propriedades de pasta dos cereais matinais extrusados e das farinhas não processadas (artigo 4).
4) Estudar o efeito da adição da casca de jabuticaba (Myrciaria cauliflora) em pó (CJP) nas características físicas do cereal matinal (índice de expansão, densidade, cor,
6
propriedades de textura a seco e após imersão em leite) e relacionar com a aceitabilidade sensorial (artigo 5).
5) Avaliar o efeito do processo de extrusão no perfil e teor de compostos fenólicos e da capacidade de absorbância do radical oxigênio em cereal matinal extrusado à base de milho, contendo também FTGI e CJP (artigo 6).
7
PAPER 1
Whole grains and fruit as ingredients in functional
breakfast cereal – A review
Authors: Ludmilla C. Oliveira, Caroline J. Steel
9
Whole grains and fruit as ingredients in functional extruded breakfast cereals Review
Ludmilla C. Oliveira1, Caroline J. Steel1,2
Abstract
Ready-to-eat breakfast cereals are products with high popularity and their consumption
follows the new food trends, mainly concerned to healthy products. Extrusion technology,
more specifically twin-screw extruders, appeared to attend most of the requirements for
new breakfast cereals, as taste modification and texture improvement. In the context of
functional breakfast cereals, whole grains and fruits are an option of raw materials because
of their bioactive compounds. In this review, we have focused on the prospect of extrusion
process, breakfast cereals and alternative ingredients that can be applied in view of their
phytochemicals, with health benefits. We have also gathered current knowledge about the
extrusion effect on phenolic acids and anthocyanins from a variety of materials.
Highlights:
- Extrusion cooking is a promising technology to improve the nutritional quality of
breakfast cereals.
1
University of Campinas, School of Food Engineering, Department of Food Technology, P.O. Box 6121, Campinas, SP, Brazil, Zip Code 13083-862.
2
10
- Whole grains are rich phytochemical sources and should be applied in extruded
breakfast cereals.
- Fruit-based ingredients are a good alternative to improve the nutritional and sensorial
characteristics of breakfast cereals.
Overview of extrusion cooking
Extrusion cooking is considered to be a HTST (high-temperature, short-time)
processing method and is capable of preserving desirable food components and of
destroying microorganisms (White, Howard & Prior, 2009). The extruder provides many
advantages that minimize processing time, energy and cost, while introducing versatility
and flexibility not previously available by other cooking processes (Huber & Rokey, 1990).
One of the benefits of extrusion is to increase the variety of foods in the diet by producing a
range of products with different shapes, colors, flavors, and textures from basic ingredients.
The physical characteristics of extrudates are governed by the properties of feed materials
and extrusion-cooking parameters such as moisture content, die diameter, barrel
temperature, screw speed, compression ratio and feed rate (Ali, Hanna & Chinnaswamy,
1996; Harper, Linko & Mercier, 1989).
The twin-screw extruder got popularity in the food industry in the
mid-1980s-mid-1990s, originally developed for processing plastics. Very soon, twin-screw extruders
became popular with the food manufactures for specialized food items. They are more
complex and costly than single screw extruders, but at the same time provide much more
flexibility and better control. Efficient material transport due to the forward conveying
11
pressure and leakage flows by virtue of the direction of screw rotation, screw shape, screw
configuration and relative position of the screw sections (Noguchi, 1989). Twin-screw
extruders appeared to attend some requirements for new products as taste modification and
texture improvement. The improved mixing and kneading functions of the twin-screw
extruders provide the transport and heat exchange required to obtain a high number of gas
cells with lower volume, resulting in textural advantages that include expansion, crispness
and general mouthfeel. Furthermore, the development of extruders has evolved to yield
sophisticated products, new flavor generation, encapsulation and sterilization (Steel et al.,
2012). They are generally categorized according to the direction of screw rotation and to
the degree to which the screws intermesh in counter-rotating and co-rotating twin-screw
extruders (Guy, 2001).
Extruders are composed of five main parts: (i) the pre-conditioning system; (ii) the
feeding system; (iii) the screw or worm; (iv) the barrel; (v) the die and the cutting
12
Figure 1 - Schematic representation of an extruder including its main parts and zones (Steel
et al., 2012).
The twin-screw extrusion has become a popular food processing techniques
especially in the cereal and snack food industry. They are widely used in ready-to-eat
(RTE) breakfast cereal production and have revolutionized the cereal industry with high
quality extruded products, offered to consumers at competitive prices (Eastman, Orthoefer
& Solorio, 2001).
The two main factors that influence the characteristics of extruded products are: raw
material characteristics and operational conditions of the extruder. The operational
parameters that can be pointed out as important are: temperature, pressure, die diameter and
shear force, with the latter being influenced by the internal design of the extruder and by its
13
Extruded breakfast cereals
Definition and market
According to the Australian Guide to Healthy Eating definition, breakfast cereals
are foods of the group of “breads and cereals” and are a “core food”. Core foods are those
that are adequate for daily consumption with the quality of supplying nutrients of great
importance to health. In a technological aspect, breakfast cereals are products traditionally
consumed with milk, and starch is their main component (Souza & Menezes, 2006). The
breakfast cereal market may be segmented in RTE (ready-to-eat) cereals and hot cereals.
The first group comprises any cereals that are served without heating or cooking, such as
corn flakes or muesli, and include ready-to-serve snack packs. The hot cereal segment
consists of porridge, oatmeal and instant hot oat products (Research & Markets). Breakfast
cereals are generally high in carbohydrates, low in fat, some are high in fiber and many
contain appreciable amounts of certain vitamins and minerals, particularly those that have
been fortified (Galvin, Kiely & Flynn, 2003). These products have been in the market for
some decades and their consumption follows the new food trends, situated as energetic,
source of protein, nutritive, practical and healthy products.
Convenience, health and wellness are the main drivers of growth in breakfast
cereals. Breakfast cereals are traditionally associated with breakfast, increasing purchasing
power, combined with a more hectic lifestyle leading consumers to look for more
convenient foods and snack options; these issues are contributing to the increase of the
14
of some RTE cereals and especially their sodium content (Euromonitor, 2012; Takeuchi,
Sabadini & Cunha, 2005).
The cereals business is well-defined in the United States of America (USA),
Canada, United Kingdom and Australia, which together account for 54 % of global
consumption despite having just 6 % of the world population, according to
Switzerland-based Cereal Partners Worldwide. The global breakfast cereal market was evaluated at $
32.5 billions in 2012 and has an expected growth of 4 % for the next few years, with a
forecast of $ 43.2 billions in 2019. The Nielsen Company reported that in 2008 only 17.7 %
of the Brazilian population consumed breakfast cereals at home, which opened the
opportunity of increasing this market category. In 2011, breakfast cereals were amongst the
Brazilian products that gained consumption participation because of their health benefits.
According to Datamark, the corn flakes breakfast cereals consumption is estimated in
59.244 tons for 2016, with a forecast of US$ 574.65 millions. Nestlé and Kellogg’s are the
market leaders, having equally 39% of participation in the national market. São Paulo is the
state with the greatest breakfast cereal consumption, with 0.2 kg/person/year (2011).
In breakfast cereal processing, the dry ingredients and the processing parameters
should be carefully monitored to obtain high quality products (Eastman, Orthoefer &
Solorio, 2001). The control of feed rate, screw speed, barrel temperature and barrel
pressure, together with the above mentioned critical parameters, will determine the
crispness, hardness and various other characteristics that will influence the success of the
product (Harper, 1981). The success or failure of a new breakfast cereal is directly related
15
expected, texture is of major importance, with crispness being one of the most important
attributes (Pamies et al., 2000). Studies have shown that texture is one of the most
important quality characteristics in expanded extrudates (Ding et al., 2006; Mendonça,
Grossmann & Verhé, 2000; Peressini, Foschia, Tubaro & Sensidoni, 2015; Robin et al.,
2012; Yanniotis, Petraki & Soumpasi, 2007) and that it affects consumer acceptability. The
degree of expansion determines the extrudate structure and consequently its texture (Thymi,
Krokida, Pappa & Maroulis, 2005). Expansion index, bulk density and texture account for
the main technological characteristics evaluated in extruded products. Breakfast cereals are
traditionally consumed immersed in milk and during the immersion these products undergo
rehydration and moisture uptake leading to changes in texture that need to be controlled in
terms of raw material and process (Chassagne-Berces et al., 2011; Sacchetti et al., 2003).
Raw materials
As main characteristics of the raw materials for extrusion, the following can be
highlighted: type of material, moisture content, physical state, chemical composition
(quantity and type of starch, proteins, fats and sugars) and pH (Carvalho, Takeiti,
Onwulata & Pordesimo, 2010). Ingredients affect the characteristics of the finished product
in many ways. Different raw materials have been applied and the physical (bulk density,
expansion index, water absorption index, water solubility index and texture), starch pasting
properties, nutritional (total dietary fiber and glycemic index) and sensorial properties have
been studied (Brennan et al., 2008; Chassagne-Berces et al., 2011; Vernaza, Matsuura,
16
Although cereals constitute the basis of breakfast cereals formulations, fruit-based
ingredients (by-products, powder juice, peel) may be a good alternative to improve the
nutritional and sensorial characteristics of the final product. The fiber content, as well as
other bioactive components, i.e., phenolic compounds, with antioxidant activity, are the
target in applying these sources in food extrusion. Furthermore, Brazil, as a tropical
country, produces a large amount of native and exotic fruit species which are potentially
interesting for the food industry. Exotic fruits, consumed regionally, are gaining popularity
in the market place for their nutritional and functional value, due to the presence of
bioactive compounds, secondary metabolites, which have potential effects on human
health. Also, their pleasant flavors and variety of colors are attractive (Clerici &
Carvalho-Silva, 2011; Oliveira, Lopes, Cabral & Eberlin, 2006; Rufino et al., 2010).
Wheat, maize and rice are the most grown cereals and the most widely used among
cereal foods (FAO, 2011). In cereal grains, the endosperm comprises >80 % of the whole
grain, whereas the percentages accounted for the germ and bran components vary among
different grains. Generally, grains are high in dietary fiber, low in fat, have in the range of
10-15 % protein, and are concentrated sources of starch (about 70 %), are high in vitamins
(especially B-vitamins), and good sources of minerals, particularly the trace minerals (iron
and zinc) (Slavin, Martini, Jacobs & Marquart, 1999). Cereals can be applied as grits or
flours in whole or refined form.
Corn meal is a major ingredient for ready-to-eat breakfast cereal products because
of the unique starch characteristics on expansion (Jozinović et al., 2013). The effect of
17
behavior of corn grits and other flours have been extensively studied (Ali, Hanna &
Chinnaswamy, 1996; Altan, McCarthy & Maskan, 2009; Carvalho, Takeiti, Onwulata &
Pordesimo, 2010; Chuang & Yeh, 2004; Delgado-Licon et al., 2008; Jozinović et al., 2013;
Onwulata & Konstance, 2006).
Examples of whole grains include whole wheat, oats, brown rice, rye, barley, and
bulgur (Ye et al., 2012). Wheat accounts for one-third of the total worldwide grain
production (Slavin, Martini, Jacobs & Marquart, 1999) and has been a cereal widely used in
the extrusion industry (Ryu & Ng, 2001). The wheat grain is composed of three parts: the
endosperm containing mostly starch and proteins, the germ composed mostly of lipids and
proteins and the bran containing mainly dietary fiber (Marquart, Jones, Cohen & Poutanen,
2007).
Starch transformations during extrusion are the main occurrences in terms of
physical processing and determinant on product characteristics, especially the expansion
mechanism (Moraru & Kokini, 2003), and depend on process conditions. Maximum
expansion degree is closely related to starch content (Riaz, 2000). Inside the extruder,
starch passes through several stages. First, the initial moisture content is very important to
define the desired product type. Once inside the extruder, and at relatively high
temperatures, the starch granules melt, besides changing their structure that is compressed
to a flattened form (Guy, 2001). The application of heat, the action of shear on the starch
granule and water content destroy the organized molecular structure, also resulting in
molecular hydrolysis of the material, reducing viscosity and releasing amylose and
18
starch polymers are then dispersed and degraded to form a continuous fluid melt. At the
exit of the extruder, the fluid polymer continuum retains water steam bubbles and stretches
during extrudate expansion until the rupture of cell structure. The starch polymer cell walls
recoil and stiffen as they cool to stabilize the extrudate structure. Finally, the starch
polymer becomes glassy as moisture is removed, forming a hard brittle texture (Guy, 2001).
The final expanded product presents air cells that are formed due to superheated water
steam pressure. When the temperature of the extrudate is reduced below its glass transition
temperature (Tg), it solidifies and maintains its expanded form (Riaz, 2000).
Commercially available breakfast cereals are predominantly corn and wheat based
foods, though other grains such as rice, oat, sorghum, or a combination of grains, as in
multigrain breakfast cereals, are also produced through extrusion cooking. The specific
advantages of such combinations include the possibility of different textures, micronutrient
content, and lowering the cost of extrudate feed (Bhattacharya, 2011).
Extrusion process variables
The functional characteristics of the starch can be altered by the extrusion
conditions, operation parameters such as speed screw, screw configuration, extruder
temperature, feed moisture, die diameter; system parameters such as shear force, product
temperature, residence time and pressure; and structural parameters associated with the raw
material.
The effect of extrusion parameters has been reported (Ding et al., 2006; Ding,
19
dependent on the process conditions such as screw speed, barrel temperature, moisture
content, feed composition, and screw configuration. All of these affect the viscosity of the
material and the flow inside the screw channels, influencing the final product
characteristics such as water solubility index (WSI), extrudate density, expansion index and
hardness. SME values indicate the extent of molecular breakdown or degradation that the
material undergoes during the extrusion process (Harper, 1981; Mercier, Linko & Harper,
1989; Meuser, Pfaller & Lengerich, 1987; Van Lengerich & Meuser, 1989).
Feed moisture has been found to be the main factor affecting extrudate expansion
and density (Ding, Ainsworth, Tucker & Marson, 2005; Ilo, Liu & Berghofer, 1999) by
promoting changes in melt viscosity inside of barrel and consequently in specific
mechanical energy (SME).
Functional breakfast cereals
Consumers have developed a growing understanding of how the composition of
food products can impact on the nutritional quality of foods (Brennan, 2005), mainly due
the components naturally present in low concentration in fruits and vegetables (Kitts,
1994), the bioactive compounds. Bioactive compounds in food and food products play a
vital role in human health providing protection against many chronic and degenerative
diseases (Van Dokkum, Frølich, Saltmarsh & Gee, 2008). These compounds are naturally
present in cereals, fruits and vegetables and belong to diverse chemical classes, with
distinct structures and bioactive mechanisms. Some of these substances with physiological
20
bioactive peptides, polyunsaturated fatty acids and phenolic compounds (polyphenols,
anthocyanins, flavonoids) (Arvanitoyannis & Van Houwelingen-Koukaliaroglou, 2005).
Fruits and vegetables are considered as major sources of bioactive compounds however
grains are also potential source of bioactive compounds particularly phenolic acids
(Brennan, Brennan, Derbyshire & Tiwari, 2011).
Alternative ingredients can be incorporated in extruded products formulations to
improve the nutritional and sensory quality. These include whole grains, oat flour, pulse
grains and vegetables (fruits and others) by-products. More recently, colored cereals and
fruits (powder or juice) started to be explored in extruded products (Camire, Chaovanalikit,
Dougherty & Briggs, 2002; Camire, Dougherty & Briggs, 2007; Escalante-Aburto et al.,
2013), with the interest of improving the appearance and functional characteristics of the
final product (Liu et al., 1999).
Breakfast cereals made from whole grain flours containing a reasonable amount of
fiber are gaining popularity among consumers because of their health benefits. Enrichment
of extruded directly expanded starchy foods with dietary fibers is a major trend in the food
industry in response to the consumer demand for healthier products. When replacing
refined flours by whole grains in extruded products, several challenges need to be
addressed. They involve reduced expansion at the die exit, darker color of the product due
to the presence of bran and/or Maillard browning and lipid oxidation after extrusion that
may be compensated by changing the extrusion conditions (Camire, 2004; Oliveira, Rosell
21
The studies with whole grains are more widespread than those focused on the
incorporation of fruit and vegetables to obtain bioactive compound enriched breakfast
cereals products of acceptable quality (Dehghan-Shoar, Hardacre & Brennan, 2010). A
cereal based functional food that account with both fruit and whole grain, source of
polyphenols and fibers has a good potential to provide health human benefits.
Dietary fiber
The Codex Alimentarius and the European Food Safety Authority (EFSA) have
published definitions of dietary fiber which differ mainly concerning the inclusion of
oligosaccharides with DP 3-9 (included in EFSA but left for local regulatory authorities to
decide in the Codex definition) (Raninen, Lappi, Mykkänen & Poutanen, 2011). EFSA defines dietary fiber as “carbohydrate polymers with three or more monomeric units, which are neither digested nor absorbed in the human small intestine and belong to the following
categories: (i) edible carbohydrate polymers naturally occurring in the food as consumed;
(ii) edible carbohydrate polymers which have been obtained from food raw material by
physical, enzymatic or chemical means and which have a beneficial physiological effect
demonstrated by generally accepted scientific evidence; and (iii) edible synthetic
carbohydrate polymers which have a beneficial physiological effect demonstrated by generally accepted scientific evidence”. Lignin and other components such as phenolic compounds, waxes, saponins, phytates, cutin, and phytosterols are also considered as
dietary fiber when closely associated with carbohydrate polymers of plant origin and
22
addition to arabinoxylan, β-glucan and cellulose, lignin and other phenolics associated to carbohydrate polymers are part of the cereal dietary fiber complex. The most recent definition for “dietary fiber terminology” was adopted by Codex Alimentarius Comission in 2009. The Codex Alimentarius Commission’s Committee on Nutrition and Foods for Special Dietary Uses adopted a new definition of dietary fiber for inclusion in the
Guidelines on Nutrition and Health Claims. The committee defined dietary fiber as “carbohydrate polymers with 10 or more monomeric units, which are not hydrolyzed by the endogenous enzymes in the small intestine of humans” (Fuentes-Zaragoza, Riquelme-Navarrete, Sánchez-Zapata & Pérez-Álvarez, 2010). In 2003, the FAO/WHO expert
consultation recommended that whole grain cereals, fruits and vegetables are the preferred
sources of non-starch polysaccharides (NSP), the intake of which should be at least 20
g/day.
Whole grains, vegetables and fruits are important sources of many nutrients
including dietary fiber, consumption of which has been associated to health benefits
(Anderson et al., 2009). In general, fibers are constituents of plant cell walls, which consist
of polysaccharides (cellulose, hemicellulose, mucilage, pectin) (DeVries, Prosky, Li &
Cho, 1999). According to their solubility in water, total dietary fiber can be categorized into
two groups: soluble (pectin, glucan, soluble pentosans) and insoluble (cellulose, lignin,
insoluble pentosans, protopectin) dietary fiber.
Resistant starch (RS) was first recognized as a complicating factor in the
determination of total dietary fiber levels by the Prosky Method (Englyst, Trowell,
23
polysaccharides” (Asp et al., 1988), it is clear that any form of starch, which interferes with the assay, is not a traditional fiber. Subsequent research has shown that RS is not rapidly
digested like ordinary starch, and that this feature imparts biological benefits. Some of the
benefits are similar to traditional fiber, and others are unique to resistant starch (Haralampu,
2000).
RS has been defined as the fraction of starch which escapes digestion in the small
intestine, and may be partially or totally digested in the colon (Englyst, Kingman &
Cummings, 1992). A number of factors contribute to the resistance of starch to digestion,
which has led to five categories of RS, each with similar resistance properties. They are as
follows: (i) RS1 - physically inaccessible to digestion by entrapment in a non-digestible
matrix; (ii) RS2 - ungelatinized starch; (iii) RS3 - retrograded starch (Englyst, Kingman &
Cummings, 1992); (iv) RS4 - chemically modified starch (Eerlingen & Delcour, 1995); (v)
RS5 - amylose-lipid complex (Jiang et al., 2010). Extrusion cooking, in some cases, is
associated to resistant starch formation (Faraj, Vasanthan & Hoover, 2004; Gebhardt,
Dongowski, Huth & Rabe, 2001).
Phenolic compounds and antioxidant activity
Phenolic compounds are common dietary phytochemicals (bioactive compounds)
found in fruits, vegetables and grains. Phenolic acids (hydroxycinnamic acids and
hydroxybenzoic acids) and flavonoids (anthocyanins, chalcones, flavanols, flavanonols,
flavanones, flavonols, flavones, isoflavone and xanthones) are common phenolic in food
24
may have protective effects against degenerative diseases (Mazza, 2000). Most of the
beneficial characteristics of phenolic compounds have been ascribed to their antioxidant
activity, which is a fundamental property to life (Rice-Evans, Miller, & Paganga, 1997).
Antioxidants are substances that reduce, prevent or retard oxidative damage of a target
molecule (Gutteridge & Halliwell, 2010). They exert effect by removing of reactive
oxygen species (ROS) and reactive nitrogen species (ROS/RNS) or prevent their formation.
The increasing interest in natural antioxidants has contributed to the evaluation of the
antioxidant activity of many species of fruits, vegetables, herbs and cereals
(Liyana-Pathirana & Shahidi, 2005). These antioxidant compounds include flavonoids, phenolic
acids, carotenoids, and tocopherols that can inhibit Fe3+/ascorbic acid-induced oxidation,
scavenge free radicals, and act as reductants (Khanduja & Bhardwaj, 2003; Ozsoy, et al.,
2009).
The general analytical strategy to determination of phenols involves recovery of the
phenol from the sample matrix followed by separation, identification and measurement.
Separation is commonly achieved by HPLC although GC is used in some instances. The
most common mode of separation exploits reversed-phase systems typically with a C18
column and various mobile phases. Detection is routinely achieved by ultraviolet
absorption often involving a photodiode array detector although the versatility of the latter
often appears to have been neglected. Coupled techniques particularly various mass spectral
methods are being used increasingly for routine works although analyte collection using
25
samples. Otherwise, antioxidant activity of food extracts can be determined using a variety
of tests (stable free radical scavengers).
The most common way of estimating the potential free radical scavenging activity
of antioxidants is through antioxidant capacity, a parameter determined in aqueous-organic
food extracts by different procedures (ORAC, ABTS, FRAP, etc.) (Robards, 2003). The
oxygen radical absorbance capacity (ORAC) assay is still one of the few methods that
combines both inhibition percentage and inhibition time of the reactive species action by
antioxidants into a single quantity (Cao et al., 1995). The ORAC assay has been largely
applied to the assessment of free radical scavenging capacity of human plasma, proteins,
DNA, pure antioxidant compounds and antioxidant plant/food extracts (Prior & Cao, 1999).
Extraction procedures strongly influence the composition of the extracts and, therefore, also
influence the antioxidant activity results (Ju & Howard, 2003; Liyana-Pathirana & Shahidi,
2005; Wang et al., 2008). In addition, the effect of the antioxidant compound in a food
matrix may be significantly different than the activity of a purified extract (Brewer, 2011).
Anthocyanins (Figure 2), which belong to the flavonoids class, are an example of
bioactive compounds. They constitute the largest and probably the most important group of
water soluble natural pigments responsible for the vivid blue, purple and red color of many
vegetables and fruits (Kong et al., 2003). The majority of research has focused on the
antioxidant properties of these dietary polyphenols (He & Giusti, 2010; Konczak & Zhang,
2004). Many reports have shown that anthocyanin exhibits an array of pharmacological
properties, such as anti-inflammatory and anti-carcinogenic activities, as well as preventive
26
treatments (Nichenametla, Taruscio, Barney & Exon, 2006; Umar Lule & Xia, 2005) and
human nutrition (Stintzing & Carle, 2004). A linear correlation between the values of the
antioxidant capacity and the anthocyanins content in blackberries, red raspberries, black
raspberries and strawberries, which can be extended to jabuticaba (Myrciaria cauliflora)
peel, has also been reported (Wang & Lin, 2000).
Figure 2 - General structure of anthocyanins. Source: Castañeda-Ovando et al. (2009).
Whole grain wheat flour (WGWF)
Following reports of epidemiological studies showing the importance of whole
grain, numerous actions have been taken to promote the use of cereal foods containing
whole grain and dietary fiber. Whole grains are defined as intact, ground, cracked or flaked
caryopsis (grain), whose main morphological components – starchy endosperm, germ and
bran – are present in the same relative proportion that exist in the intact caryopsis (AACC,
1999). They are rich sources of fiber, vitamins, minerals, and phytochemicals including phenolics, carotenoids, vitamin E, lignans, β-glucan, inulin, resistant starch, sterols, and phytates (Liu, 2003, 2004; Slavin, 2000). Components in whole grains that may be
27
protective include compounds that affect the gut environment, such as dietary fiber,
resistant starch, and oligosaccharides. Whole grains are also rich in compounds that
function as antioxidants, such as trace minerals and phenolic compounds, and
phytoestrogens, with potential hormonal effects. The range of protective substances in
whole grains is impressive and advice to consume additional whole grains is justified
(Slavin, Martini, Jacobs & Marquart, 1999).
Epidemiological studies have associated the consumption of whole grain and whole
grain products with reduced incidence of chronic diseases such as cardiovascular diseases
(Jacobs, Meyer, Kushi & Folsom, 1998), diabetes (Meyer et al., 2000), and cancer (Jacobs,
Slavin & Marquart, 1995; Thompson, 1994). These health benefits have been attributed in
part to the unique phytochemical content of grains.
Whole grains are rich in fiber and phytochemicals and are among the healthiest
foods that individuals may consume and render a wide variety of health benefits
(Andreasen, Christensen, Meyer & Hansen, 2000a). Andreasen, Christensen, Meyer, and
Hansen (2000b) have reported positive effects of higher intake of whole grain foods in
lowering the risk of coronary heart disease. It has also been suggested that adults would
gain appreciable protection from coronary heart disease by consuming the recommended at
least three servings of whole grains daily (The Whole Grains Council).
Whole grain wheat flour contains phenolic compounds with antioxidant activity.
Two sequences of phenolic acids are of significant importance: derivatives of benzoic and
28
& Fornal, 2006). The chemical structures of benzoic and cinnamic acids, as well as their
derivatives, are presented in Figures 2 and 3.
Figure 3 - Cinnamic acid and its derivatives. Source: Borkowski (1993) in Klepacka &
Fornal (2006).
Figure 4 - Benzoic acid and its derivatives. Source: Borkowski (1993) in Klepacka &
29
Phenolic acids, particularly the hydroxy-cinnamic acids known as ferulic acid and
p-coumaric acid, are found in plant cell walls that generally link cellulose to other polysaccharide components. These compounds are thought to decrease the fermentability of
dietary fiber (Stephen, 1994). Ferulic, p-coumaric, and syringic acid are the main phenolic
compounds in WGWF (Klepacka & Fornal, 2006). A recent study of phenolic acids in
wheat flours found that ferulic acid was the most concentrated phenolic acid; its phenolic
acid content is lower in more refined flour. Free, soluble-conjugated, and bound ferulic acid
have been found in WGWF (Adom, Sorrells & Liu, 2003; Kim, Tsao, Yang & Cui, 2006;
Mateo Anson et al., 2008; Okarter, Liu, Sorrells & Liu, 2010).
The incorporation of wheat bran in extruded products is still limited due to its
negative effect on physical properties (Brennan, Monro & Brennan, 2008; Yanniotis,
Petraki & Soumpasi, 2007). Wheat bran contains a high amount of dietary fibers, reported
to bring nutritional and health benefits (Marlett, McBurney & Slavin, 2002).
Whole grain wheat flour is also an excellent fiber source, with approximately 13 %
(Andersson et al., 2013; Haskå, Nyman & Andersson, 2008), mainly in the insoluble form
(Adom, Sorrells & Liu, 2003), concentrated in the outer layer of the grain (Esposito et al.,
2005). The physiological effects of dietary fiber are dependent of their physical-chemical
properties and of the ratio of soluble to insoluble fiber (Plaami, 1997).
Jabuticaba (Myrciaria cauliflora)
The plant family Myrtaceae is pan-tropical in occurrence, with concentrations in
