Effect of Fermentation and Cooking on Soluble and Bound Phenolic

Subscriber access provided by Northern Illinois University

Article

Effect of fermentation and cooking on soluble and bound phenolic profiles of finger millet sour porridge Molly Gabaza, Habtu Shumoy, Maud Muchuweti, Peter Vandamme, and Katleen Raes J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.6b03090 • Publication Date (Web): 19 Sep 2016 Downloaded from http://pubs.acs.org on September 19, 2016

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

Journal of Agricultural and Food Chemistry is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 28

Journal of Agricultural and Food Chemistry

TOC 338x190mm (96 x 96 DPI)

ACS Paragon Plus Environment


1

Effect of fermentation and cooking on soluble and bound phenolic profiles of finger

2

millet sour porridge

3 4

Molly Gabaza1,2,3, Habtu Shumoy1, Maud Muchuweti2, Peter Vandamme3, Katleen

5

Raes1

6 7

1

8

University, Graaf Karel de Goedelaan 5, 8500 Kortrijk, Belgium

9

2

Department of Industrial Biological Sciences, Faculty of Bioscience Engineering, Ghent

Department of Biochemistry, Faculty of Science, University of Zimbabwe, P.O. Box MP

10

167, Mt Pleasant, Harare, Zimbabwe

11

3

12

Ledeganckstraat 35, 9000 Gent, Belgium

Department of Biochemistry and Microbiology, Faculty of Science, Ghent University,

13 14

Corresponding author: [email protected]

15 16 17 18 19 20 21

1 ACS Paragon Plus Environment

Page 2 of 28

Page 3 of 28


22

Abstract

23

The aim of this study was to evaluate the soluble and bound phenolic content of finger millet

24

and the impact of process induced changes on phenolic profiles of their sour porridge. Finger

25

millet porridge and intermediate products were collected from four groups of households in

26

Hwedza communal area, Zimbabwe, after which soluble and bound phenolic compounds (PC)

27

including condensed tannins (CT) were quantified. Bound PC and CT contributed 95 % of the

28

total PC and CT. The CT were only detected in the red varieties. Major individual PC

29

identified were catechin occurring in the soluble fraction only, while ferulic, sinapic and

30

salicylic acid were mainly present in the bound fraction. Fermentation and cooking caused a

31

more than two fold increase in soluble PC, CT and individual PC. Improved traditional

32

processing techniques optimized for improved bioavailability and health benefits of phenolics

33

are highly relevant for the low income populations.

34 35

Keywords: Phenolic compounds, condensed tannins, finger millet, fermentation, cooking

36 37 38 39 40



41

Introduction

42

Small grains such as sorghum and millets are drought resistant and thus are an important food

43

component to low income populations in many parts of Africa. Finger millet (Eleusine

44

coracana) is a small grain containing high amounts of phenolic compounds (PC), many of

45

which have therapeutic benefits such as heart disease prevention, anticancer, antihypertensive

46

and anti-inflammatory characteristics

47

and white varieties with the dark colored varieties possessing higher contents of PC, and also

48

condensed tannins (CT) than the light colored varieties

49

finger millet are mainly present in the black varieties compared to other varieties, similar to

50

the presence of anthocyanins in sorghum varieties 6, 7. According to Dykes and Rooney 3 and

51

Chandrasekara and Shahidi 4, CT have higher antioxidant activity than monomeric PC.

52

However, PC may behave like a double edged sword as on the one hand they exhibit positive

53

health effects but on the other hand certain PC with catechol and galloyl groups may

54

negatively affect mineral absorption

55

diets as they have adverse effects on protein digestibility, enzyme activity and mineral

56

absorption 10. Phenolic compounds in cereal grains exist as free, in a soluble conjugated or an

57

insoluble bound form 11, 12. PC in cereal grains exist mainly in the bound form, and values of

58

bound PC varying between 75 and 91 % have been reported for blue maize, brown rice, red

59

sorghum and wheat 11, 12.

60

In Africa, fermented thin porridge is quite common. It is consumed by both adults and

61

children and is also used as weaning and complementary food for young children.

62

Fermentation and cooking are two processing methods widely used in the African cuisine that

63

are able to induce important chemical changes, e.g. the destruction of anti-nutritional factors

64

such as phytates and CT. The effects of processing on PC differ among cereal grains and can

65

be related to the type of grains, the location of the different PC in the cereal matrix and the

1-4

. Finger millet consists of black, dark brown or red

3, 5

. In addition, the anthocyanins in

8, 9

. CT are even considered as anti-nutrients in many


Page 4 of 28

Page 5 of 28


13

66

intensity of the processing method

. There are contradictions in literature pertaining to the

67

effect of fermentation and/or cooking on the amount of PC and CT in cereals. Additionally

68

there is a paucity of information concerning the changes occurring to soluble and bound PC

69

after fermentation and cooking of cereals, as well as regarding different finger millet varieties.

70

Many studies report changes occurring to total contents of PC without making a distinction

71

between soluble and bound PC

72

the content and change in both soluble and bound PC fractions are not well known, thus their

73

possible related effects towards health or interactions with other food components are unclear.

74

Furthermore, many studies use model food products prepared under standardized laboratory

75

conditions and information about ‘real samples’ from the field is lacking.

76

The objective of this study is thus to compare the profile and content of soluble and bound PC

77

and CT of red and white varieties of finger millet, and to determine the effect of traditional

78

household processing, in particular fermentation and cooking, on the content of soluble and

79

bound PC and CT, and on changes in PC profile of finger millet porridge.

14-16

. This implies that the impact of processing conditions on

80 81

Materials and Methods

82

Materials

83

Methanol, sodium hydroxide (NaOH), sodium carbonate (NaCO3), hydrochloric acid (HCl)

84

were purchased from Merck (Darmstadt, Germany). Folin-Ciocalteu’s reagent was purchased

85

from ChemLab (Zedelgem, Belgium), chemical standards: gallic acid, catechin, p-coumaric

86

acid, ferulic acid, syringic acid, sinapic acid, vanillic acid, protocatechuic acid, p-

87

hydroxybenzoic acid, caffeic acid, cinnamic acid, were purchased from Sigma Aldrich

88

(Bornem, Belgium). HPLC-grade methanol (MeOH), HPLC-grade water, trifluoroacetic acid

89

(TFA) were purchased from VWR (Leuven, Belgium).

90



91

Methods

92

Sample preparation

93

Finger millet products were prepared by four groups of women (n=24) in Ushe communal

94

area (Hwedza district, Mashonaland East Province, Zimbabwe). The women were tasked to

95

prepare fermented slurries and fermented porridges according to their traditions and to record

96

the recipes accurately and with as much detail as possible. The products were prepared from

97

two red and two white varieties and each group will be identified according to the variety

98

used i.e. red variety 1 (RV1), red variety 2 (RV2), white variety 1 (WV1), white variety 2

99

(WV2). Finger millet grains were cleaned to remove glumes and foreign particles after which

100

they were finely milled to pass through a sieve of 1 mm size. The fermentation process was

101

done in a thatched kitchen with approximate temperature of 23-25oC. An aliquot of flour was

102

measured into a fermentation vessel which could be a plastic or metal container followed by

103

carefully adding a measured aliquot of water. The vessel was tightly closed and spontaneous

104

fermentation allowed to proceed for 24-36 hours. Successful fermentation was identified by a

105

characteristic frothing of the fermented slurry at the surface and a typical fermented aroma

106

(measured pH of 4 - 4.5). The fermented slurry was then cooked for 15-20 minutes, with or

107

without extra added water, to make the porridge. Table 1 shows the type of products and

108

amount of ingredients used by each group, as well as the used preparation conditions.

109

Sample collection

110

Fermented slurry and porridge samples were collected in aliquots of 50 mL in sterile falcon

111

tubes between July 1 and 12, 2014. The sampling was done twice within a time frame of two

112

weeks. Samples were always transported under cooled conditions and immediately frozen at -

113

80oC. Samples were transported to Belgium under dry ice and stored at -20oC. Flour of the

114

different varieties was also collected and stored at -20oC at the Laboratory of Food

115

Microbiology and Biotechnology, Ghent University, Campus Kortrijk, Belgium.


Page 6 of 28

Page 7 of 28


116

Extraction of Soluble PC

117

Extraction of soluble PC was based on Gonzales, et al. 17. Briefly, 5 g of porridge samples and

118

fermented slurries and 2 g of flour samples were added with 15 ml methanol in 50 ml falcon

119

tubes. The mixture was homogenized using an ultra turrax (IKA T18 Basic) at 10000 rpm for

120

45 s and placed on ice for 15 minutes thereafter. To separate the soluble extract from the

121

residue, the mixture was centrifuged (Z 300 K, Hermle Labortechnik, GmbH, Germany) at

122

4000 rpm, 15 minutes at 4oC. Re-extraction was done using 80 % methanol following the

123

same procedure. The collected supernatants were pooled and filled to 25 ml with 80 %

124

methanol. The extract was transferred to storage bottles covered with aluminum foil to protect

125

from sunlight and stored at -20oC until further analysis. The residue was dried overnight at

126

room temperature and bound PC were extracted from it the next day.

127

Extraction of Bound PC

128

Bound PC were extracted according to method described by Gonzales, et al.

129

residue, obtained after extraction of the soluble PC (0.1 g) was placed in 25 ml screw-capped

130

falcon tubes previously flushed with nitrogen and hydrolyzed with 2 ml NaOH in a sonicated

131

water bath (UP 400 S, dr.Hielscher, GmbH, Germany) at 60oC for 30 minutes. The mixture

132

was neutralized with 4 ml HCl and extracted twice with 0.1 % formic acid in methanol for 2

133

minutes under vortex. Supernatants were always separated by centrifugation at 4000 g for 15

134

minutes and 4oC. The supernatants were pooled and filled to 25 ml using 80 % methanol and

135

stored in storage bottles at -20oC covered in aluminum foil.

136

Determination of total PC

137

The total PC were determined on both the soluble and bound fractions according to the Folin-

138

Ciocalteu method as described by Singleton and Rossi 18. Briefly, extracts from soluble and

139

bound fraction (1 ml) were added to test tubes with 1 ml deionized water following by 0.5 ml

140

ten times diluted Folin-Ciocalteu’s reagent. After 5 minutes, 1.5 ml of 20 % NaCO3 and 1 ml


17

. The dried


141

deionized water was added and the mixture was allowed to stand for 2 hours in the dark after

142

which the absorbance was read at 760 nm. The phenolic contents for each fraction was

143

calculated based on a standard curve generated from gallic acid and expressed as mg Gallic

144

Acid Equivalents (GAE)/100g dm.

145

Determination of Condensed Tannins (CT)

146

Proanthocyanidins or CT were analyzed according to the method described by Price, et al. 19.

147

Bound and soluble extracts and standards (1 ml) were added to test tubes followed by 5 ml of

148

vanillin reagent (50:50 mixture of 5 % vanillin and 24 % HCl w/v) and incubated for exactly

149

20 minutes in a water bath at 30oC. A second set of control tubes was prepared by adding 1 ml

150

of sample or standards followed by 12 % HCl, and incubated under the same conditions after

151

which the absorbance was read at 500 nm. The final absorbance was determined by

152

subtracting the absorbance of the sample control from the corresponding vanillin-containing

153

sample. The CT for each fraction was calculated based on a standard curve generated from

154

catechin and expressed as mg catechin equivalents (CE)/100 g dm.

155

Identification of PC by HPLC

156

Identification of PC was done following the method described by Wen, et al. 20. Soluble and

157

bound sample extracts were dried under nitrogen, and redissolved in 500 µL of

158

MeOH:water:trifluoroacetic acid (TFA) (50: 50: 0.1) and filtered. An internal standard of o-

159

coumaric acid was added to the samples and the samples were injected directly into the HPLC

160

system. The separation of phenolic acids was carried out with Agilent HPLC equipped with

161

column (AlltimaTM –Column 18 5u (4.6 mm × 150 mm), GRACE, Deerfield, USA). The

162

mobile phase consisted of solvent mixture A (0.02 % TFA in water) and solvent mixture B

163

(0.02 % TFA in methanol). The following program was used for gradient elution: 0-5 mins

164

(25 % B), 5-10 mins (25-30 % B), 10-16 mins (30-45 % B), 16-18 mins (45 % B), 18-25 mins

165

(45-80 % B), 25-30 mins (80 % B), 30-40 mins (80-25 % B). Samples were injected at a


Page 8 of 28

Page 9 of 28


166

volume of 10 µL. The DAD detection was set at four wavelengths namely 254 nm (for

167

identification of protocatechuic and vanillic acid), 275 nm (gallic, syringic, cinnamic, catechin

168

and internal standard), 305 nm (p-coumaric and salicylic acid) and 320 nm (caffeic, ferulic,

169

sinapic and rosmarinic acid). Quantification of identified phenolic acids was done using

170

corresponding regression equation and values are expressed as µg/g dm.

171 172

Statistical Analysis

173

All statistical analysis were done using IBM SPSS software version 20. The differences of

174

mean values among the varieties were determined using one-way analysis of variance

175

(ANOVA) followed by Tukey’s Honestly Significant Differences (HSD) multiple rank test at

176

P < 0.05 significance level.

177 178

Results

179

Soluble and bound PC and CT of finger millet flour

180

Figure 1 shows the soluble and bound PC found in red and white finger millet samples from

181

Ushe communal area, Zimbabwe. The total content of PC of the red and white finger millet

182

ranged from 1108-1330 and 334-359 mg GAE/100g dm, respectively. The bound fraction

183

accounted for approximately 95 % of the total PC for both red and white millet samples.

184

Figure 2 shows the content of CT in the red finger millet. The CT in the white varieties could

185

not be detected. Higher amounts of CT in the bound fraction (89-94 % of the total CT

186

content) than in the soluble fraction was also observed, due to the association of CT with cell

187

walls. The white varieties of finger millet contain lower amounts of PC than the red varieties.



188

PC profile of finger millet flour

189

Seven phenolic acids i.e. cinnamic acid, syringic acid, p-coumaric acid, salicylic acid, sinapic

190

acid, ferulic acid, protocatechuic acid, caffeic acid and one flavonoid namely catechin were

191

identified in the different millet samples (Table 2). Caffeic acid was only found in the bound

192

fraction. Catechin was the only flavonoid identified in the soluble fraction where its peak

193

occupied more than 50 % of the total area in soluble extracts. The major phenolic acids

194

identified were ferulic acid, sinapic acid and salicylic acid. Ferulic acid was the most

195

abundant phenolic acid in the bound fractions of both the white and red varieties with values

196

ranging from 107-193 µg/g. Sinapic acid was higher in the white varieties (241-330 µg/g)

197

than in the red varieties (49.1-110 µg/g) and 37-59 % was found in the bound fraction.

198

Sinapic acid appeared to be the major phenolic acid in white variety and also appeared to be

199

the dominant soluble phenolic acid in all the varieties. Salicylic acid was also found in high

200

amounts with up to 195 µg/g in the bound fraction of WV2 and 75-98 % existing in the bound

201

fraction.

202

Effect of fermentation and cooking on soluble and bound PC and CT of finger millet

203

porridge

204

The soluble, bound and total PC and CT of fermented and finger millet porridges from red

205

and white varieties are shown in Tables 1 and 2. After fermentation, there was a general

206

reduction in the content of bound and total PC and CT while there was a significant increase

207

in the soluble PC in all the varieties (p < 0.05). This shows that despite different household

208

processing conditions, in general, fermentation reduced the content of bound and total PC and

209

increased the content of soluble PC. Total PC were reduced by 6-31 % while bound PC were

210

reduced by 17-37 % with the lowest reductions observed for WV1. Soluble PC were increased

211

by 16-167 % and the highest increase was observed for WV1 and lowest increase for RV2. In

212

terms of CT, a decrease of 32-52 % and 43-47 % was observed for total and bound CT,


Page 10 of 28

Page 11 of 28


213

respectively while an increase of 34-55 % was seen for the soluble CT. The level of reduction

214

of PC and CT in all varieties was different which could be explained by the different

215

fermentation conditions used as this influences the microbial diversity of the fermented

216

slurries.

217

After fermentation, cooking of fermented slurries follows to make the porridge. In

218

comparison with fermented slurries, soluble PC significantly increased in porridges by 33-186

219

% with the highest increase observed for RV2 and WV2 while soluble CT increased by 76

220

and 92 % in RV1 and RV2, respectively (p < 0.05). The bound PC increased by 22 and 34 %

221

in RV1 and RV2 while they decreased by 26 and 44 % in WV2 and WV1. In general, cooking

222

after fermentation caused an increase in total PC for the red varieties while there was a

223

decrease for the white varieties. In terms of the CT in the porridges, no significant change was

224

observed for the bound and total CT after cooking compared to fermented slurries. The

225

overall effect of cooking combined with fermentation caused more than two times increase in

226

both the soluble PC and CT. Bound PC decreased by 4-53 % while total PC decreased by 14-

227

36 % for all varieties except for RV2 where no change in total PC was observed. For CT,

228

bound CT decreased by up to 63 % and total CT was reduced by up to 35 %.

229 230

Effect of fermentation and cooking on soluble and bound individual PC of finger millet

231

In all varieties, an increase of all individual soluble phenolic acids was observed after

232

fermentation and cooking while an inconsistent trend was seen for bound phenolic acids

233

(Table 2). Catechin increased more than four times after fermentation and cooking, with the

234

highest increase in the red varieties. There was an increase of bound phenolic acids after

235

fermentation for RV1 and WV1, while a decrease of bound phenolic acids was observed for

236

WV2 and RV2. After cooking, levels of phenolic acids increased in bound fraction except for

237

samples from WV1 where all bound phenolic acids decreased after cooking.



Page 12 of 28

238 239 240

Discussion

241

The PC in finger millet are known to be mainly concentrated in the seed coat of the grain

242

and are present both in soluble and bound form and also occurring more abundantly in the red

243

varieties than in the white varieties. A major proportion of PC found in cereal grains are in the

244

bound form and can be up to 91 % for some maize varieties

245

approximately 95 % of the total PC were in the bound form and, to our knowledge, such

246

associations have never been reported in finger millet. A high amount of phenolic acids in

247

both red and white varieties was also found in the bound fraction. The chief hydroxycinnamic

248

acids, ferulic, p-coumaric, caffeic and sinapic acid are known to be ester linked to various cell

249

wall polymers such as arabinoxylans thus they exist in high amounts in the bound fraction

250

21

251

also in the present study the most abundant phenolic acid in the bound fraction of both the

252

white and red varieties. Subba Rao and Muralikrishna

253

µg/g in finger millet and this is in agreement with our findings of 107-193 µg/g. The phenolic

254

acids identified in the present study have also been reported by others 3, 4, 24 but their amounts

255

varied largely showing that the occurrence of different PC in grains may depend on several

256

factors such as environmental, edaphic agricultural and varietal differences.

257

Fermentation caused an increase in the soluble PC, CT and individual PC and a decrease in

258

the bound ones. Different mechanisms have been suggested to explain the changes occurring

259

to PC and CT after fermentation. The low pH attained during fermentation can cause the

260

removal of hydride ions and reorganization of PC 9, some PC may self-polymerize and also

261

interact with other macromolecules such as proteins thus decreasing their extraction,

262

polymerized PC may be degraded into oligomeric PC thus increasing the content of the

5

11, 12

. In the present study,

7,

. Ferulic acid was reported as the major bound phenolic acid in cereal grains 12, 21-23 and was

22

reported ferulic acid content of 186


Page 13 of 28


10, 25

263

soluble PC

. Also the synthesis or enzymatic transformations of PC may occur during

264

fermentation

265

microflora may result in structural breakdown of cell walls thereby increasing the

266

accessibility of bound PC to enzymatic attack. Tannin-protein and tannin-iron complexes are

267

hydrolyzed during fermentation therefore the reduction of PC and CT after fermentation could

268

improve the digestibility of proteins and the bioaccessibility of trace minerals such as iron and

269

zinc

270

for degradation of PC and CT could result in improved protein and mineral nutrition. The

271

increase of phenolic acids in the soluble fraction suggest that esterase and glucosidase

272

activities may have been rampant during fermentation as their activities are known to release

273

phenolic acids bound to cell walls in particular protocatechuic and p-hydroxybenzoic acids 27.

274

The effect of thermal processes on PC is dependent on the type of thermal process employed,

275

i.e. wet cooking, steam cooking, extrusion cooking, baking and roasting15. In our case, wet

276

cooking was employed during the preparation of porridge. The further increase of soluble PC,

277

soluble CT and individual soluble PC after cooking was consistent in all millet samples as

278

shown in Table 2-4. In contrast to soluble PC, there was a variable response to bound PC and

279

CT after cooking. According to Duodu 25, PC in grains can increase, decrease or not change

280

after processing depending with the mechanism of action of the processing method employed.

281

The decrease of bound PC, CT and phenolic acids could be a result of leaching of PC into the

282

cooking water with subsequent increase of the soluble PC. Increases of soluble PC observed

283

in this study are caused by processes that release bound PC making them more extractable and

284

assayable. Such processes may involve depolymerization reactions. The increase of the

285

soluble PC and CT in our study could be mainly attributed to the increase of catechin,

286

particularly in the red varieties. This is because finger millet contains proanthocyanidins,

287

large polymers of catechin, which were depolymerized during fermentation and cooking. The

10, 13

. During fermentation, enzymes from both the cereal matrix and the

26

. It also implies that a controlled fermentation with the use of starter cultures suitable



Page 14 of 28

288

catechin data shows that it is the main flavonoid occurring in the soluble fraction. Catechin

289

accounts for 84 % of the soluble fraction in finger millet

290

measure CT is a crude method that unfortunately, also measures catechin, explaining the

291

observed increase of CT in the soluble fraction. The CT in finger millet have not been

292

characterized in detail like CT in sorghum but they are likely to be oligomers of catechin as

293

catechin, epicatechin, gallocatechin, epigallocatechin, trimers and tetramers of catechin and

294

procyanidin dimers B1 and B3 have been identified in finger millet

295

bound PC increased in the red varieties but decreased in the white varieties after cooking. The

296

decrease of bound PC and phenolic acids in WV1 may be as a result of the cooking process

297

employed. After fermentation, the slurry was decanted and the coarse particles containing the

298

seed coat with the highest PC was discarded. The red varieties of finger millet contain CT

299

whose extractability could have been increased after thermal treatment. Several authors have

300

found a decrease of total PC after cooking of sorghum, fonio, pearl, kodo, finger, foxtail,

301

proso and little millet 7, 15, 29, 30 while others reported increases 31. Varying effects of thermal

302

treatment on bound PC and CT of different type of grains have also been reported by other

303

workers 15, 27, 32.

304

Soluble phenolics are stored in the intracellular space of cells while bound phenolics are

305

located in proximity and in association with cell walls

306

coupled with general decrease of bound phenolics after fermentation and cooking alludes to

307

the fact that there is depolymerisation of bound phenolics and their transportation into the

308

intracellular space. As the decrease of the bound phenolics does not add up with the increase

309

of the soluble phenolics, it suggests that some other chemical and physical interactions come

310

into play. The changes occurring to soluble and bound phenolics can thus allow for prediction

311

of localization, transport mechanism and chemistry of phenolics during processing 33.

24

. The vanillin method used to

24, 28

. The content of

33

. The increase of soluble phenolics


Page 15 of 28


312

Several studies have shown improved mineral bioaccessibility after reduction of PC and CT 9,

313

34

314

astringency, improved color, acceptance, protein digestibility and mineral bioavailability

315

On the other hand, fermentation and/or cooking has also been shown to effect changes on PC

316

and CT which subsequently increase the antioxidant capacity of the products 16, 31, 36. Kumari

317

et al

318

finger millet, the antioxidant activity of the soluble extract measured by various antioxidant

319

tests i.e. DPPH radical scavenging activity, trolox equivalent antioxidant activity, β-carotene-

320

linoleate model system and ferrous ion-chelating activity, was consistently at least twice that

321

of the bound extracts. Yeo and Shahidi

322

phenolics as an estimate to predict the localization, transportation mechanisms and

323

antioxidant activity of phenolics in germinating lentils. The ratio of bound to soluble

324

phenolics of our samples decreased from 16.7-20.8 in the flour to 2.18-5.67 after fermentation

325

and cooking. Using this ratio of bound to soluble phenolics, we can postulate that the

326

antioxidant activity after fermentation and cooking will increase by a large magnitude.

327

However, the benefits of finger millet phenolics go beyond their antioxidant activity. Future

328

research should thus focus on determining whether the increase in soluble PC also improves

329

the absorption of phenolics via in vivo and in vitro studies. Increased absorption is a

330

prerequisite for the phenolics to demonstrate their purpoted health effects.

331

The improvement of processing techniques is highly relevant for low income populations

332

subsisting on finger millet. Household processing techniques such as fermentation and

333

cooking can induce important changes to phenolic phytochemicals which could influence

334

their function both as anti-nutrients and functional ingredients. The increase of soluble PC

335

after household processing is beneficial as simpler and soluble PC are more bioaccessible and

336

may lead to improved bioaccessibility of minerals. The adequacy of household processing in

. The reduction of CT in cereals is encouraged as it leads to products with reduced

37

35

.

revealed that even though bound phenolics were higher than soluble phenolics in

33

proposed to use the ratio of bound to soluble



337

terms of improving mineral bioaccessibility and improved utilization of PC as functional

338

foods should be tested by in vitro and in vivo assays.

339 340

Abbreviations

341

CT: condensed tannins; PC: phenolic compounds; TFA: trifluoroacetic acid

342 343

Acknowledgements

344

The authors are very grateful to the women from Hwedza communal area and to the

345

agricultural extension workers for provision of the raw materials and preparation of the

346

products. Further gratitude goes to SOFECSA (Soil Fertility Consortium of Southern Africa)

347

team led by Prof. Paul Mapfumo at the University of Zimbabwe for being advisors in the

348

study and for assisting in field work.

349 350

Funding information

351

We are thankful to VLIR-UOS for PhD funding of Molly Gabaza.

352 353

References

354

(1)

355

millets and pseudocereals: Developments in the science of their phenolic phytochemicals,

356

biofortification and protein functionality. J. Cereal Sci. 2014, 59, 257-275.

357

(2)

358

Food Sci. Nutr. 2014, 56, 25-35.

359

(3)

360

2006, 44, 236-251.

Taylor, J.; Belton, P. S.; Beta, T.;Duodu, K. G. Increasing the utilisation of sorghum,

Van Hung, P. Phenolic compounds of cereals and their antioxidant capacity. Crit. Rev.

Dykes, L.;Rooney, L. W. Sorghum and millet phenols and antioxidants. J. Cereal Sci.


Page 16 of 28

Page 17 of 28


361

(4)

Chandrasekara, A.;Shahidi, F. Content of insoluble bound phenolics in millets and

362

their contribution to antioxidant capacity. J. Agric. Food Chem. 2010, 58, 6706-6714.

363

(5)

364

effect of pH on their stability. Food Chem. 2007, 105, 862-870.

365

(6)

366

sorghum. J. Agric. Food Chem. 2004, 52, 4388-4394.

367

(7)

368

cooking on the total antioxidant capacity and phenolic profile of some whole‐meal African

369

cereals. J. Sci. Food Agric. 2013, 93, 29-36.

370

(8)

371

bioaccessibility of minerals: Influence of localization of minerals and anti-nutritional factors

372

in the plant. Trends Food Sci. Technol. 2014, 37, 32-41.

373

(9)

374

sorghum gruels: effects on phenolic compounds, phytate and in vitro accessible iron. Food

375

Chem. 2006, 94, 369-376.

376

(10)

377

phytochemicals and the implications of this to the health‐enhancing properties of sorghum

378

and millet food and beverage products. J. Sci. Food Agric. 2015, 95, 225-237.

379

(11)

380

foods, a review. Food Chem. 2014, 152, 46-55.

381

(12)

382

6182-6187.

383

(13)

384

compounds in cereal grains through processing technologies: A concise review. J. Funct.

385

Foods. 2014, 7, 101-111.

Chethan, S.;Malleshi, N. Finger millet polyphenols: Optimization of extraction and the

Awika, J. M.; Rooney, L. W.;Waniska, R. D. Properties of 3-deoxyanthocyanins from

N'Dri, D.; Mazzeo, T.; Zaupa, M.; Ferracane, R.; Fogliano, V.;Pellegrini, N. Effect of

Raes, K.; Knockaert, D.; Struijs, K.;Van Camp, J. Role of processing on

Towo, E.; Matuschek, E.;Svanberg, U. Fermentation and enzyme treatment of tannin

Taylor, J.;Duodu, K. G. Effects of processing sorghum and millets on their phenolic

Acosta-Estrada, B. A.; Gutiérrez-Uribe, J. A.;Serna-Saldívar, S. O. Bound phenolics in

Adom, K. K.;Liu, R. H. Antioxidant activity of grains. J. Agric. Food Chem. 2002, 50,

Wang, T.; He, F.;Chen, G. Improving bioaccessibility and bioavailability of phenolic



386

(14)

Chandrasekara, A.;Shahidi, F. Bioaccessibility and antioxidant potential of millet

387

grain phenolics as affected by simulated in vitro digestion and microbial fermentation. J.

388

Funct. Foods. 2012, 4, 226-237.

389

(15)

390

and bioaccessibility in finger millet (Eleusine coracana) and pearl millet (Pennisetum

391

glaucum). Food Chem. 2014, 164, 55-62.

392

(16)

393

phytochemicals during malting and fermentation of type III tannin sorghum and impact on

394

product biofunctionality. J. Agric. Food Chem. 2013, 61, 1935-1942.

395

(17)

396

and ultrasound-assisted extraction for the release of nonextractable phenolics from

397

cauliflower (Brassica oleracea var. botrytis) waste. J. Agric. Food Chem. 2014, 62, 3371-

398

3376.

399

(18)

400

phosphotungstic acid reagents. Am. J. Enol. Vitic. 1965, 16, 144-158.

401

(19)

402

as an assay for tannin in sorghum grain. J. Agric. Food Chem. 1978, 26, 1214-1218.

403

(20)

404

determination of phenolic acids in compound herbal medicines. J. Agric. Food Chem. 2005,

405

53, 6624-6629.

406

(21)

407

and bound phenolic acids from native and malted finger millet (Ragi, Eleusine coracana

408

Indaf-15). J. Agric. Food Chem. 2002, 50, 889-892.

Hithamani, G.;Srinivasan, K. Effect of domestic processing on the polyphenol content

Kayodé, A. P.; Mertz, C.; Guyot, J.-P.; Brat, P.;Mouquet-Rivier, C. Fate of

Gonzales, G. B.; Smagghe, G.; Raes, K.;Van Camp, J. Combined alkaline hydrolysis

Singleton, V. L.;Rossi, J. A. Colorimetry of total phenolics with phosphomolybdic-

Price, M. L.; Van Scoyoc, S.;Butler, L. G. A critical evaluation of the vanillin reaction

Wen, D.; Li, C.; Di, H.; Liao, Y.;Liu, H. A universal HPLC method for the

Subba Rao, M. V.;Muralikrishna, G. Evaluation of the antioxidant properties of free


Page 18 of 28

Page 19 of 28


409

(22)

Mattila, P.; Pihlava, J.-m.;Hellström, J. Contents of phenolic acids, alkyl-and

410

alkenylresorcinols, and avenanthramides in commercial grain products. J. Agric. Food Chem.

411

2005, 53, 8290-8295.

412

(23)

413

acids from native and malted finger millet (Ragi, Eleusine coracana, Indaf-15). Food Chem.

414

2001, 72, 187-192.

415

(24)

416

hydrolyzed fractions of millet grains and characterization of their phenolic profiles by HPLC-

417

DAD-ESI-MS n. J. Funct. Foods. 2011, 3, 144-158.

418

(25)

419

Cereal Food World. 2014, 59, 64-70.

420

(26)

421

production: Effects of composition and selected functional properties. Pak. J. Nutr 2009, 8,

422

737-744.

423

(27)

424

acids and flavonoids in nonfermented and fermented red sorghum (Sorghum bicolor (L)

425

Moench). J. Agric. Food Chem. 2010, 58, 9214-9220.

426

(28)

427

of finger millet (Eleusine coracana L.) seed coat phenolics: Mode of inhibition of α-

428

glucosidase and pancreatic amylase. Food Chem. 2009, 115, 1268-1273.

429

(29)

430

activity of millet grains. Food Chem. 2012, 133, 1-9.

431

(30)

432

extractable phenolic groups in cereals and legumes commonly consumed in Tanzania. J. Sci.

433

Food Agric. 2003, 83, 980-986.

Subba Rao, M. V.;Muralikrishna, G. Non-starch polysaccharides and bound phenolic

Chandrasekara, A.;Shahidi, F. Determination of antioxidant activity in free and

Duodu, K. Effects of processing on phenolic phytochemicals in cereals and legumes.

Onweluzo, J.;Nwabugwu, C. Fermentation of millet and pigeon pea seeds for flour

Svensson, L.; Sekwati-Monang, B.; Lutz, D. L.; Schieber, A.;Ganzle, M. Phenolic

Shobana, S.; Sreerama, Y.;Malleshi, N. Composition and enzyme inhibitory properties

Chandrasekara, A.; Naczk, M.;Shahidi, F. Effect of processing on the antioxidant

Towo, E. E.; Svanberg, U.;Ndossi, G. D. Effect of grain pre‐treatment on different



434

(31)

Pradeep, S.;Guha, M. Effect of processing methods on the nutraceutical and

435

antioxidant properties of little millet (Panicum sumatrense) extracts. Food Chem. 2011, 126,

436

1643-1647.

437

(32)

438

and biological activities for production of sorghum tea. Food Res. Int. 2013, 53, 678-685.

439

(33)

440

soluble phenolics on antioxidant activity of lentils during germination. J. Agric. Food Chem.

441

2015, 63, 379-381.

442

(34)

443

zinc in vitro availability in pearl millet flours (Pennisetum glaucum) with varying phytate,

444

tannin and fibre contents. J. Agric. Food Chem. 2005, 53, 3240-3247.

445

(35)

446

processing on the antioxidant properties of African sorghum-based foods. Food Chem. 2007,

447

105, 1412-1419.

448

(36)

449

antioxidant capacity and polyphenol distribution in selected edible legumes. Int. J. Food Sci.

450

Tech. 2016, 875-884.

451

(37)

452

antioxidant activities of millet varieties grown in different locations in Sri Lanka. Food

453

Sci.Nutr. 2016, 1-13.

Wu, L.; Huang, Z.; Qin, P.;Ren, G. Effects of processing on phytochemical profiles

Yeo, J.;Shahidi, F. Critical evaluation of changes in the ratio of insoluble bound to

Lestienne, I.; Besancon, P.; Caporiccio, B.; Lullien-Pellerin, V.;Treche, S. Iron and

Dlamini, N. R.; Taylor, J. R.;Rooney, L. W. The effect of sorghum type and

Gan, R. Y.; Shah, N. P.; Wang, M. F.; Lui, W. Y.;Corke, H. Fermentation alters

Kumari, D.; Madhujith, T.;Chandrasekara, A. Comparison of phenolic content and

454 455


Page 20 of 28

Page 21 of 28


456

Figure Captions

457

Figure 1. Soluble and bound PC (mg GAE/100g dm) of red and white finger millet samples.

458

Figure 2. CT (mg CE/100g dm) of soluble and bound extracts of red finger millet samples



Page 22 of 28

Table 1: Levels of ingredients and processing parameters used in preparation of finger millet fermented slurries and porridges, as noted by the women preparing the porridges Variety

RV1

RV2

WV1

WV2

100 g flour

100 g flour

100 g flour

100 g flour

300 mL water

200 mL water

300 mL water

400 mL water

24 h fermentation

24-30 h fermentation

24 h fermentation

36 h fermentation

Extra 200 mL water

Extra 300 mL water

No extra water added. Extra 400 mL water

Products Fermented slurry

Cooked porridge (~20 minutes cooking for

Fermented slurry was

all porridges)

decanted

and

coarse

particles discarded.

21

ACS Paragon Plus Environment

Page 23 of 28


1600

mg GAE/100g dm

1400

b

1200

a

b

a

1000 800 600 c

400 200

b

a

c

c

c

c

c

0 RV1

RV2 soluble PC

WV1 Bound PC

WV2

Total PC

Figure 1: Soluble and bound PC (mg GAE/100 g dm) of red and white finger millet samples. Bars with the same color with different letters are significantly different (p ≤ 0.05), n=2.



b

900 b

mg CE/100g dm

800 700 600

a

a

500 400 300 200 100

b a

0 RV1

RV2

CT in soluble extract

CT in bound extract

Total CT

Figure 2: CT (mg CE/100 g dm) of soluble and bound extracts of red finger millet samples. Bars with the same color with different letters are significantly different (p ≤ 0.05), n=2.


Page 24 of 28

Page 25 of 28


Table 2: PC in finger millet flour, fermented slurries and porridges of finger millet RV1 Soluble Flour

RV2 Bound

Total

WV1

Soluble

Bound

55.1±0.1a 1053±5.2a 1108±4.2a 75.1±4.7a 1255±89a

Fermented 107±1.9b

WV2

Total

Soluble

Bound

1330±84a

20.1±0.6a 339±12a

982±98b

53.7±4b

Total

Soluble

Bound

Total

359±11a 15.3±0.9a 318±42a 334±41a

662±70.5b 769±70b

87.2±6.2a 895±100b

281±24a,b 336±21a 21.7±2.1a 215±13b 237±12b

142±24c

805±21c

947±49c

249±4.9b

1200±4.6a 1450±0.3a 71.6±10c

156±85b

255±91a 51.4±8.6b 159±14b 213±21b

Effect of Fermentation and Cooking on Soluble and Bound Phenolic

Recommend Documents