Natural food colouring with dye sorghum leaf sheaths

Folachodé U.G. Akogou

Research output: Thesisinternal PhD, WU


Cereals are mainly used as staple foods as well as for animal feed and biofuel. In addition, certain cereals are of interest to the food industry for more specific reasons. This is the case for dye sorghum (Sorghum bicolor), due to its ability to produce significant amounts of pigment that potentially can be used as a food colorant. The red leaf sheaths formed by dye sorghum are used in Benin to produce a red biocolorant, which is also used in for foods. The red biocolorant is a rich source of 3-deoxyanthocyanidins (i.e. apigeninidin and luteolinidin). The 3-deoxyanthocyanidins are natural pigments with (a) a narrow distribution in nature, (b) a high antioxidant activity, (c) resistance to bleaching agents, (d) resistance to ring openings during thermal treatments, (e) a better stability to pH-induced colour changes as compared to common anthocyanins and (f) properties that inhibit the growth of cancer cells. These properties of 3-deoxyanthocyanidins contribute to colour stability during food processing. Currently, the red sorghum biocolorant is applied in Benin in cereal-, legume-, tuber- and milk-based foods. Dyed fermented cereal porridge (koko) and dyed soft cheese (wagashi) are the most popular food applications of this red biocolorant in Benin and illustrate the potential as a new stable natural colorant from sorghum that can be used in food industry. The growing interest of consumers for natural food ingredients leads to a growing demand of the food industry for natural food colorants. However, to date limited data are available on the optimal extraction conditions for sorghum biocolorant and its properties in a wider range of food products. This thesis studies the extraction conditions of biocolorant from dye sorghum leaf sheaths, evaluates its bioactive properties, as well as its nutritional and sensorial impact on food products.

The chapter 2 compares the colour and anthocyanin composition from various traditional extraction methods to determine options for improvement and use of the red biocolorant from dye sorghum in the food sector. After interviewing processors of dyed foods on the procedures of the biocolorant extraction, data were collected on the biocolorant yield and the pigment profile of the extraction methods. In addition, potential food applications were illustrated by a popular fermented porridge. Sorghum biocolorant was extracted from a batch of dye sorghum leaf sheaths with different extraction methods. The use of an alkaline rock salt (locally known as kanwu) and the temperature of the extraction water were found to be the two main parameters that differentiate the traditional extraction methods. Water, dye sorghum leaf sheaths and kanwu were mixed at the ratio 900:10.4:1.5 (w/w/w), 900:10.5:0.9 (w/w/w) and 900:11.3:0 (w/w/w) to obtain sorghum biocolorant by cool alkaline, hot alkaline and hot aqueous extraction. While the cool alkaline extraction was performed at room temperature, a gradual heating at the speed of 0.05 °C s-1 was applied to hot (alkaline and aqueous) extractions until 86 ºC was reached. Cool extraction was more efficient than hot alkaline and hot aqueous extractions in taking out anthocyanins. The total anthocyanin content of the cool alkaline extract was 228.5 µg mL-1. However, the alkaline extractions (hot and cool) at pH 8-9 were the most efficient methods because their apigeninidin content (131-152 µg mL-1) was three times higher than in the aqueous extract. Nevertheless, improvement of the extraction method is necessary since the residues still contained 82.6% of anthocyanins. In Benin, sorghum biocolorant is used to colour maize-based koko, a fermented and cooked porridge. This demonstrates the ability of sorghum biocolorant to colour foods produced by fermentation and elevated temperatures.

Sorghum biocolorant is perceived by local processors of a soft cheese (called wagashi) as a means to extend the shelf-life of wagashi. If such a claim could be substantiated, it adds a valuable functional property to this natural red colorant. Hence, Chapter 3 evaluates the antimicrobial properties of dye sorghum extracts using challenge tests in broth and wagashi as a model of a popular food application. The challenge tests (a) in dyed broth with Listeria monocytogenes and Escherichia coli O157:H7 and (b) on dyed wagashi with fungi (i.e. Penicillium chrysogenum, Cladosporium macrocarpum) and Escherichia coli O157:H7 were used to evaluate the antimicrobial activity of the sorghum biocolorant. Additional data on the physico-chemical parameters of the dyed wagashi (pH, dry matter and the acid value) were monitored as well. Data on the current practices of wagashi dyeing were collected as well from 90 processors. The application of sorghum biocolorant on wagashi had no inhibitory effect on the growth of fungi (Penicillium chrysogenum, Cladosporium macrocarpum) and Escherichia coli O157:H7 on wagashi. Furthermore, sorghum biocolorant in broth had no effect on growth of Listeria monocytogenes and Escherichia coli O157:H7. Consequently, the commonly used extracts for colouring soft West-African cheese did not show a preservative effect for the species tested. In addition, dyeing did not affect the physico-chemical properties of wagashi. Still, the red colour hampered visual detection of microbial growth, and this might clarify the preservative effect reported by users. Furthermore, the dyeing of wagashi is a means for the processors to maximise their revenues. Indeed, an increase of 11.5% of the revenues was recorded for dyed wagashi. The majority of the processors usually used a dyeing procedure with no heat treatment. This urges to raise the users’ attention on the absence of a preservative effect and to also apply a heat treatment to the colorant to minimise the risk of wagashi contamination when a conservation for several days is needed.

Chapter 4 evaluates the stability of apigeninidin, the main 3-deoxyanthocyanidin from sorghum leaf sheaths, to food processing conditions in watery extracts (alkaline and hot aqueous extracts) and in a maize porridge. The stability of apigeninidin in sorghum biocolorant extract was evaluated in (a) a pH range of 5-12, (b) heat treatments (65 °C, 95 °C and 121 °C) and (c) storage conditions (light exposure, protected from light exposure and temperature). The total anthocyanins before, after the heat treatments and during storage were used to evaluate the thermal stability of sorghum biocolorant. In addition, kinetic degradation of apigeninidin at pH 6 and 9 was compared. Apigeninidin was not soluble at pH 5.04 ± 0.02. At pH 5.04, the loss of net charge led to apigeninidin precipitation in the watery extract. This limits the solubility and the application of apigeninidin in acidic drinks (pH ≤ 5). At pH values higher than 6, apigeninidin was soluble with the quinoidal base as major form in solution. Apigeninidin was soluble and stable at pH 6–10 with increased colour density and resistance to bleaching at alkaline pH. At pH 11 and 12, 44.1% and 81.4% of the apigeninidin degraded. Apigeninidin has a remarkable resistance to high pH and can therefore be subjected to neutral and alkaline (e.g. alkaline fermented food condiments in West Africa) treatments, contrary to most anthocyanins, which are stable in acid and middle acid conditions. The stability of apigeninidin to the heat treatments (i.e. 65 °C / 30 min, 95 °C / 30 min and 121 °C / 30 min) was similar at pH 7 and 8.7 with a degradation of 17-18%, 59-66% and 60-61% of the anthocyanins at 65 °C / 30 min, 95 °C / 30 min and 121 °C/ 30 min, respectively. The comparable percentage of degraded anthocyanins at 95 °C and 121 °C suggests the good resistance of apigeninidin to severe heat treatments. However, the resistance of apigeninidin might be affected by the pH of the extract. At 65 °C, the degradation rate of apigeninidin was four times lower at pH 9.03 ± 0.04 than at 6.08 ± 0.02. Storage at room temperature promoted endothermic degradation of apigeninidin. Nevertheless, photodegradation of apigeninidin was not observed during storage. In the maize porridge, thermal stability of apigeninidin and redness were similar at pH 4–6 whereas they were higher at pH 9.03 ± 0.04.

Chapter 5 assesses the impact of the apigeninidin-rich sorghum biocolorant on the nutritional and sensorial quality in a fermented food. Maize dough was enriched with apigeninidin extract and fermented with Pichia kudriavzevii and Lactobacillus fermentum for 3 days at 30 °C. The colour, glucose content, pH, phytate dephosphorylation, oxalate degradation and formation of volatile organic compounds (VOCs) were monitored. The 3-day fermentation of the dyed maize dough (containing 327 μg/g DM of apigeninidin) led to a degradation of 69% of the apigeninidin content. The colour of the dyed dough evolved from red to orange red with increasing lightness. The antioxidant activity of the fermented dyed dough increased by 51% compared to the fermented non-dyed dough. New compounds (e.g. phenolic acids) with high antioxidant activity are apparently formed during fermentation. However, phytate dephosphorylation and volatile compound concentrations (alcohol, esters, aldehydes and alkene) were lower in dyed dough than in non-dyed dough. This suggests a lower mineral solubility and change in the sensory quality in fermented dyed dough.

Chapter 6 discusses the findings of the present study and presents the contribution of this study to (a) motivate the building of a better dye sorghum value chain, (b) the transition by the food industry to natural colorants, (c) the control of microorganisms with phenolic extracts and (d) a more accurate measurement of 3-deoxyanthocyanidins by spectrophotometry. Improvement of the sorghum value chain from traditional to traditional-to-modern or modern will allow farmers to work structured on a sustainable supply chain of dye sorghum leaf sheaths. A transition by the food industry to the use of natural colorants requires raw materials with a pigment content above 2% to be competitive. With an average pigment content of 3.1%, dye sorghum leaf sheaths are therefore a competitive natural biocolorant. However, successful substitution of artificial colorants by sorghum biocolorant does not solely depend on the stability of the natural colour regarding the processing condition but also on (a) regulations concerning the introduction of a new colorant on the market, (b) the colour difference due to the substitution, and (c) the consumer perception of the sensorial differences brought about by the substitution.

Original languageEnglish
QualificationDoctor of Philosophy
Awarding Institution
  • Wageningen University
  • Fogliano, Vincenzo, Promotor
  • Linnemann, Anita, Co-promotor
  • den Besten, Heidy, Co-promotor
  • Kayodé, A.P.P., Co-promotor, External person
Award date29 Aug 2018
Place of PublicationWageningen
Print ISBNs9789463434812
Publication statusPublished - 2018


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