Investigations on the Maillard reaction in sesame (Sesamum

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Food and Beverage Chemistry/Biochemistry

Investigations on the Maillard reaction in sesame (Sesamum indicum L.) seeds induced by roasting Ecem Berk, Aytül Hamzal#o#lu, and Vural Gökmen J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.9b01413 • Publication Date (Web): 10 Apr 2019 Downloaded from http://pubs.acs.org on April 12, 2019

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Journal of Agricultural and Food Chemistry

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Investigations on the Maillard reaction in sesame (Sesamum indicum L.) seeds

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induced by roasting

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Ecem Berk, Aytül Hamzalıoğlu, Vural Gökmen*

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Department of Food Engineering, Hacettepe University, 06800 Beytepe, Ankara,

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Turkey

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E-mail: [email protected]

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* Corresponding Author

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Prof. Dr. Vural Gökmen, Tel: 90-312-297-7108, Fax: 90-312-299-2123

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E-mail: [email protected]

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ACS Paragon Plus Environment

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ABSTRACT

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This study investigated the formation of Maillard reaction products in sesame seeds

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under different roasting conditions. Sesame seeds were roasted at 150, 180, 200 and

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220

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hydroxymethylfurfural,

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deoxyglucosone, 3-deoxyglucosone, methylglyoxal and diacetyl) together with

18

glycation markers namely N-ε-fructosyllysine, N-ε-carboxymethyllysine and N-ε-

19

carboxyethyllysine, were monitored. Roasting induced the formation of 5-

20

hydroxymethylfurfural, acrylamide and dicarbonyl compounds, except furan,

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significantly (p < 0.05). 5-hydroxymethylfurfural and acrylamide content of roasted

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sesame seeds were found varying as 3-40 mg/kg and 135-633 µg/kg, respectively.

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Dicarbonyl compounds were in the following order; methylglyoxal > 3-deoxyglucosone

24

> 1-deoxyglucosone > diacetyl. On the other hand, N-ε-fructosyllysine concentration

25

decreased

26

carboxymethyllysine, N-ε-carboxyethyllysine formation was induced under those

27

conditions. This is the first study reporting the formation of thermal process

°C

for

10

while

min

the

and

thermal

acrylamide,

roasting

process

furan

and

temperature


contaminants dicarbonyl

increased,

including

compounds

however,

5(1-

N-ε-

2


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contaminants and glycation markers in sesame seeds through Maillard reaction

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during roasting.

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Keywords: Sesamum indicum L., roasting, Maillard reaction, acrylamide, furan, 5-

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hydroxymethylfurfural, N-ε-fructosyllysine, N-ε-carboxymethyllysine, α-dicarbonyl

32

compounds


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INTRODUCTION

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Sesame (Sesamum indicum L.) is an important source of edible seed for humans,

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whose origin dates back to about 6000 years ago, and is widely cultivated in Asia and

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Africa. India, China, Sudan, Nigeria, Myanmar are leading countries in the production

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of sesame seed.1–3 The total world production is 6.1 million tons in 2016, about 65%

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of which has been processed as a sesame oil.4 Sesame seed is mostly used in bakery

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goods and confectionery products such as decorating on breads and cookies in Asian

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countries.1 Moreover, its oil is utilized for cooking oil, salad dressing, producing tahini

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and margarine. Cosmetics, chemicals and pharmaceuticals are the other fields using

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sesame seeds and its oil, apart from food applications.1,5

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There are many reasons that makes sesame attractive to people; the main ones are

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being good source of energy (up to 50% oil) and high nutritional value owing to its

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20% protein content.1,6 In addition, sesame seed shows resistance to oxidative

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deterioration because of being rich in bioactive components.7 Although sesame oil

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contains high levels of unsaturated fatty acids (80-85%), it is much more stable than


4

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other vegetable oils.2 This stability is attributed to lignans, tocopherols and, also

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browning reaction products generated as a result of roasting of sesame.2,7

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Roasting is the key step for revealing distinctive flavor, enhancing color and texture

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of sesame seeds.8 Sesame roasting is generally performed at 150-200 °C for 10-20

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min.1 Expectedly, heat treatment provides a pleasant aroma and appreciated taste for

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consumers. In addition to that, roasting process not only provides desirable

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consequences to foods, but also leads to some reactions such as Maillard reaction,

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sugar degradation, protein denaturation, lipid oxidation and vitamin degradation, as

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well.9 As a result of these reactions, formation of new compounds having antioxidant

57

activity, a desirable outcome, is possible, however, undesirable changes may also

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occur, leading to nutritional losses and formation of potentially toxic compounds.9

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Maillard reaction is initiated by condensation of an amine with a carbonyl compound

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inducing the formation of Schiff base. In the case of an aldose sugar as a carbonyl

61

compound, formation of Schiff base is followed by rearrangement to a 1-amino-1-

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deoxy-2-ketose, which is known as Amadori product.10 Roasting gives rise to the

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progress of Maillard reaction resulting in reduction in the concentration of amino acids,


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especially lysine that is an essential amino acid, and also arginine.11,12 Loss of lysine

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is monitored with the formation of N-ε-fructosyllysine, Amadori product of lysine,

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known as the early stage product of Maillard reaction.13 Furosine, an acid derivative

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of N-ε-fructosyllysine, is used for evaluation of heat load in processed foods such as

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cookies, jams and infant foods.14,15 During roasting, N-ε-fructosyllysine could

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transform into the advanced stage of Maillard reaction products such as N-ε-

70

carboxymethyllysine (CML) and N-ε-carboxyethyllysine (CEL), pyrraline depending

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upon temperature and the extent of heating.15,16 In the course of advanced stage,

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degradation of Amadori product leads to the formation of brown nitrogenous

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compounds, named as melanoidins.17 Melanoidins are responsible for changes in

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color and taste of foods. As reactive intermediates, degradation of Amadori product

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results in the formation of α-dicarbonyl compounds such as methylglyoxal (MG),

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glyoxal (GO), glucosone (G), diacetyl (DA), 3-deoxyglucosone (3-DG) and 1-

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deoxyglucosone (1-DG).18,19 They are involved in carbonyl stress in vivo20 and then,

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in the advanced stage, they induce the modification of protein bound amino acids,

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subsequently forming advanced glycation end products (AGEs). Presence of AGEs

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in foods possess a potential risk on individuals because of its link to various chronic


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diseases like diabetes, diabetic nephropathy and Alzheimer’s disease.21,22 However,

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there is a debate on whether or not dietary AGEs cause for concern about human

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health recently.23

84

Another important consequence of the Maillard reaction is the formation of toxic

85

compounds such as 5-hydroxymethylfurfural (HMF), acrylamide and furan, named

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thermal process contaminants. HMF is a common product formed through

87

caramelization (by dehydration of hexose sugars) or Maillard reaction as a result of

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heating.24,25 The content of HMF is marker of thermal load applied to many foods rich

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in carbohydrates and indicates unsuitable storage conditions, as well.25 In addition to

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this, presence of HMF in foods might bring about potential health risks. There is still

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controversial data about the effects of HMF on humans. In vitro data showed that HMF

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has caused mutagenic effects in DNA26 and cytotoxic effects on eyes, skin, upper

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respiratory tract and mucus membranes25, on the other hand, it was also concluded

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that toxic potential of HMF is low and no adverse effect level for HMF was found

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comparably higher than estimated daily intake. However, since it is the indicator of

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thermal process, HMF levels might give the information about the thermal load, and

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accordingly about the other thermal process contaminants.


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Acrylamide is categorized as probably carcinogenic to humans by International

99

Agency for Research on Cancer (IARC). Due to genotoxic and carcinogenic features,

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also neurotoxic effects in high doses27, acrylamide formation leads to a great concern

101

in foods, thus, the determination of its level is an important issue for human health.

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Furan is one of the thermal process contaminants that could arise from degradation

103

of reducing sugars like glucose, fructose and lactose together with amines or alone.

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It is also considered that oxidation of polyunsaturated fatty acid (PUFA) is one of the

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possible pathways in the formation of furan.28

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Sesame seeds have been an attractive foodstuff in human life for many years and

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utilized in various way throughout the world. As mentioned above, the nature of

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sesame seeds is convenient for the reactions induced by heat treatment. Among the

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chemical reactions occurring in foods during heating, Maillard reaction is important

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that should be investigated due to its both positive and negative impacts. However,

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the literature is lacking in the investigation of the roasting effect on the formation of

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Maillard reaction products in sesame seeds. Therefore, the objective of this study was

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to offer an insight into formation of Maillard reaction products including HMF,


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acrylamide, furosine, CML, CEL, furan and α-dicarbonyl compounds (3-DG, 1-DG,

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MG, DA) and their alteration under different roasting conditions.

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MATERIALS AND METHODS

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Chemicals and consumables

118

3-Deoxyglucosone

119

methylglyoxal (40%), 2,3-dimethylquinoxaline (97%), 5-methylquinoxaline (98%), o-

120

phenylenediamine (98%), diethylenetriaminepentaacetic acid (DETAPAC) (98%),

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ammonium formate (97%), L-theanine and sodium borohydride powder (≥98%) were

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purchased from Sigma-Aldrich (Steinheim, Germany), whereas furosine standard was

123

purchased

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hydroxymethylfurfural (98%) was purchased from Acros (Geel, Begium). Formic acid

125

(98%), methanol and acetonitrile were obtained from JT Baker (Deventer, Holland).

126

Potassium hexacyanoferrate, zinc sulfate, disodium hydrogen phosphate anhydrous,

127

sodium dihydrogen phosphate dihydrate, sodium hydroxide, boric acid, hydrochloric

128

acid (37%) were purchased from Merck (Darmstadt, Germany). Syringe filters (nylon,

129

0.45 μm), Oasis HLB and Oasis MCX cartridges were supplied by Waters (Millford,

from

(75%),

quinoxaline

Neosystem

(99%),

Laboratoire


2-methylquinoxaline

(Strasbourg,

(97%),

France).

5-

9


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MA). Ultrapure water was used throughout the experiments (Mili Q-System, Milipore,

131

Milford, MA).

132

Roasting of Sesame Seeds

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Sesame (Sesamum indicum L.), originated from Nigeria, was purchased from a local

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market (Ankara, Turkey). Ten grams of sesame seeds were placed in a petri dish as

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a homogeneous thin layer (3 mm thickness) and roasted in an oven (Memmert UN

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55, Germany) at 150, 180, 200, 220 °C for 10 min. After cooling to room temperature,

137

the samples were finely ground and kept frozen at -18 °C prior to analysis. All samples

138

were roasted in triplicate.

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Extraction of roasted sesame samples

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One gram of ground sample was triple extracted with 20 mL of water (10, 5, 5 mL),

141

and vortexed for 3 min in each step. Supernatants were centrifuged at 5000 g for 3

142

min in each step and transferred to in a test tube. Then, the tube was centrifuged at

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5000 g for 3 min and a clear extract was obtained.

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Analysis of 5-Hydroxymethylfurfural

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For the analysis of HMF in roasted sesame samples, extracts were obtained


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according to the procedure given in Extraction of roasted sesame samples section.

147

After the extract centrifuged at 10000 g for 3 min, the mixture was filtered through

148

0.45 μm nylon filter and collected into a vial. The analysis of HMF was performed with

149

respect to a previously described method.29 The filtered extract was injected into

150

Agilent 1200 HPLC system (Waldbronn, Germany) coupled with a diode array

151

detector (DAD), a quaternary pump, a temperature-controlled oven and an

152

autosampler. Separation was performed on an Atlantis dC18 column (250 mm x 4.6

153

mm, 5 μm) at 25 °C and isocratic mixture of 10 mM of formic acid solution and

154

acetonitrile (90:10, v/v) used as the mobile phase at a flow rate of 1 mL/min. The

155

detection was carried out at 285 nm. The quantification of HMF was based on the

156

external calibration curve built ranged between 0.5 and 10 μg/mL.

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Analysis of Acrylamide

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For the analysis of acrylamide in roasted sesame samples, extracts were obtained

159


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The extract was passed through a preconditioned Oasis MCX cartridge for clean-up

161

following Carrez clarification. After the first eight drops of the eluent were discarded,


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the remaining drops were taken into a vial prior to analysis. Acrylamide were analyzed

163

by a Waters Acquity H Class UPLC system operated in positive mode.

164

Chromatographic separation was performed in Hypercarb (100  2.1 mm, 3 μm)

165

column at 50 °C and 0.1% of formic acid solution was used as the mobile phase at a

166

flow rate of 0.2 mL/min. Chromatographic run was completed in 7 min and acrylamide

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was eluted in 3.80 min. The electrospray source had the following settings: capillary

168

voltage of 0.80 kV; cone voltage of 21 V; extractor voltage of 4 V; source temperature

169

at 120 °C; desolvation temperature at 450 °C; desolvation gas (nitrogen) flow of 900

170

L/h. The flow rate of the collision gas (Argon) was set to 0.25 mL/min. Acrylamide was

171

identified by multiple reaction monitoring (MRM) of two channels. The precursor ion

172

[M + H]+ 72 was fragmented and product ions 55 (collision energy of 9 V) and 44

173

(collision energy of 12 V) were monitored. The dwell time was 0.2 s for all MRM

174

transitions. Acrylamide concentration in samples was quantified by virtue of calibration

175

curve built in the range between 1.0 and 100 ng/mL (1, 2, 5, 10, 20, 50 and 100

176

ng/mL).

177

Analysis of Asparagine


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For the analysis of asparagine in roasted sesame samples, extracts were obtained

179


180

A part of combined extract was blended with an equal volume of acetonitrile:water

181

(80:20, v/v) and the mixture was centrifuged at 10000 g for 3 min. After passing

182

through a 0.45-μm syringe filter, 10 μL was injected into LC–MS/MS system.

183

Chromatographic separation was performed on Synchronis-HILIC column (100 × 2.1

184

mm, 3 μm) at 40 °C using a gradient mixture of 0.5% formic acid and 5 mM ammonium

185

formate in water (A) and 0.5% formic acid and 5 mM ammonium formate in 90%

186

acetonitrile (B) at a flow rate of 0.7 mL/min. The eluent composition starting with 100

187

% of B retained for 3 min and linearly decreased to 70% in 4 min. Then, it was linearly

188

decreased to 20% in 10 sec and retained until the end of 10th min. It was linearly

189

increased to its initial conditions (100 % of B) in 1 min and retained under this

190

conditions for 4 min. Doing so, the total chromatographic run was completed in 15

191

min. Theanine was used as an internal standard at concentration of 0.5 mg/L. Waters

192

TQD LC–MS/MS system was operated in positive ionization mode using the following

193

interface parameters: source temperature of 120 °C, desolvation temperature of 350

194

°C, collision energy 12 V, desolvation gas flow of 900 L/h, capillary voltage of 3.5 kV,

195

cone voltage of 20 V, and extractor voltage of 3 V. Data acquisition was performed by

196

monitoring m/z ratio of 133.0 for asparagine and 87.0 and 74.0 for its product ions.

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Quantification was performed by means of external calibration curves built for

198

asparagine in a range between 0.25 and 1 mg/L.

199

Analysis of Furan


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200

For the analysis of furan, an array of experiments was performed. After roasting and

201

grinding steps, 1 g of sesame samples were weighed in headspace vials and sealed

202

immediately. The content of furan was determined as described previously.30

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Analysis of Furosine

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Analysis of furosine was carried out as described previously elsewhere.31 For the

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hydrolysis process, 100 mg of ground sesame samples were weighed into a glass

206

tube and 5 mL of 8 N HCl was added onto it. After the headspace of the tubes were

207

flushed with nitrogen gas, screw caps firmly closed. Hydrolysis was carried out in an

208

oven at 110 °C for 23 h and the hydrolysates were subsequently filtered through filter

209

paper. 200 μL of the acid hydrolysates were put into a glass tube and purged with

210

nitrogen gas. Then the residue dissolved in 1 mL of deionized water, passed through

211

an Oasis HLB cartridge preconditioned with 1 mL methanol and 1 mL water. Drops,

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except for the first eight were taken into a vial. Analysis of furosine was carried out an

213

Agilent 1200 HPLC (Agilent Tech., Waldbronn, Germany) consisting of a diode array

214

detector, a quaternary pump, a temperature-controlled column oven and automatic

215

sample injection system. For the chromatographic separation, Atlantis HILIC column


14

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216

(250  4.6 mm, 5 μm) was used and conditioned to 40 °C. The mobile phase was 1%

217

formic acid in water and the flow rate was 1 mL/min. Data acquisition was performed

218

at the detector set to at 280 nm. Standard solution of furosine with concentrations

219

ranging between 1 and 10 mg/L were prepared and by means of creating a calibration

220

curve, furosine concentrations of sesame samples were calculated.

221

Analysis of N-ε-Carboxymethyllysine and N-ε-Carboxyethyllysine

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The hydrolysis procedure according to the method described by Charissou et al32 was

223

performed with some modifications. 20 mg of ground sesame sample was weighed in

224

a glass tube and mixed with 100 μL deionized water. 450 μL of sodium borate buffer

225

(0.2 M prepared and adjusted the pH of 9.2 by mixing properly 0.2 M boric acid and

226

NaOH) and 500 μL of sodium borohydride (1 M prepared in 0.1 M NaOH) were put

227

into it. Then, the tubes were kept at room temperature for 4 hours to transform

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fructosyllysine to hexitol lysine. Thereafter, 2 ml of 8 N HCl was added and the caps

229

were closed under nitrogen gas. The tubes were hydrolyzed at 110 °C for 24 hours.

230

After cooling to room temperature, 20 μL hydrolysate was transferred into a glass tube

231

and evaporated with nitrogen. By adding 1 mL of deionized water to the residue and


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passed through a preconditioned Oasis HLB cartridge. After the first eight drops, the

233

rest eluent was collected in an autosampler vial for LC-MS/MS analysis. Sample was

234

injected into Waters TQD LC–MS/MS system and the chromatographic separation

235

was performed in Atlantis T3 column (4.6  150 mm, 3 μm) at 40 °C. MS system was

236

operated in positive ionization mode using the following interface parameters: Source

237

temperature of 120 °C, desolvation temperature of 450 °C, collision gas flow of 0.20

238

mL/ min, desolvation gas flow of 900 L/min, capillary voltage of 3 kV, cone voltage of

239

25 V, and extractor voltage of 2 V Mobile phase consisting of 0.1% formic acid in

240

water (A): 0.1% formic acid in acetonitrile (B) was used at a flow rate of 0.20 mL/min.

241

The gradient mixture was started from 100% A remaining for 5 min and decreased to

242

70% in 1 min. After remaining for 1 min, it was increased to 100% in 1 min and 100%

243

A remained for 8 min. The chromatographic run was completed in 15 min. Instrument

244

was run in Multiple Reaction Monitoring (MRM) mode and acquisition was performed

245

by monitoring m/z ratio of 205.10 for CML and 219 for CEL. The precursor ions were

246

fragmented and two product ions having m/z ratio of 84 and 130 for both CML and

247

CEL were monitored. Quantification was performed by means of a matrix-matched

248

calibration curve. Sesame seeds roasted at 150 °C was used as blank matrix.


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249

Calibration solutions of CML and CEL were prepared in the blank matrix at

250

concentrations of 0, 1, 2.5, 5.0, 10, and 20 μg/mL. Then, they were subjected to the

251

lengthy extraction procedure that was used for actual samples and analyzed as

252

described above.

253

Analysis of α-Dicarbonyl Compounds

254

Extraction was performed as described above. The coextracted colloids were

255

precipitated by mixing with acetonitrile. Two hundred μL of the combined extract was

256

diluted with 800 μL of the mixture of acetonitrile:water (5:3, v/v), and centrifuged at

257

15000 g for 5 min. For derivatization; five hundred μL of supernatant was mixed with

258

150

259

diethylenetriaminepentaacetic acid and 150 μL of 0.5 M sodium phosphate buffer (pH

260

7). The mixture was immediately filtered through 0.45 μm syringe filter into an

261

autosampler vial. It was kept at room temperature, at dark for 2 h prior to LC-MS/MS

262

analysis. Waters TQD LC–MS/MS system operated in positive ionization mode was

263

used. Chromatographic separation was performed in Waters Acquity UPLC BEH C18

264

column (2.1  100 mm, 1.7 μm) at 60 °C using mobile phase consisting of 0.1% formic

μL

of

0.2%

o-phenylenediamine

solution


containing

11

mM

17


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265

acid in water: 0.1% formic acid in acetonitrile (90:10, v/v) at a flow rate of 0.40 mL/min.

266

MS system was using the following inter face parameters: Source temperature of 130

267

°C, desolvation temperature of 400 °C, collision gas flow of 0.20 mL/ min, desolvation

268

gas flow of 800 L/min, capillary voltage of 3 kV, cone voltage of 20 V, and extractor

269

voltage of 3 V. Acquisition was performed by monitoring m/z ratios for quinoxaline

270

derivatives as following: 235.2 for 3-DG and 1-DG, 145 for MG, 159.2 for DA. 5-

271

methylquinoxaline was used as an internal standard. The MRM ions of the quinoxaline

272

derivatives of α-dicarbonyl compounds were used for quantitation. The concentrations

273

of quinoxaline derivatives were calculated by means of external calibration curves in

274

the range between 0.02 and 2 mg/L. Working solutions of glucosone and 3-

275

deoxyglucosone in the concentration range between 0.01 and 2.0 mg/L were

276

derivatized and analyzed as described above to build its external calibration curve. All

277

working solutions were prepared in acetonitrile−water (50:50, v/v).

278

Analysis of Color

279

Color measurements of roasted sesames were performed by using computer-vision

280

based image analysis technique as described previously.33

281

Statistical Analysis

282

Experimental data was reported as mean ± standard deviation. All analytical

283

determinations were carried out in triplicates. Significance of differences among


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roasted samples were statistically analyzed by using one-way ANOVA Duncan’s test

285

(p < 0.05) by using SPSS Version 16.0.

286

RESULTS AND DISCUSSION

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The Maillard reaction plays a key role in browning development due to formation of

288

melanoidins. As browning is simultaneously developed together with formation of

289

thermal process contaminants, determination of the color changes during roasting

290

under different conditions is of importance. For this purpose, computer-based image

291

analysis was performed in order to obtain L*, a*, b* values for the samples roasted at

292

150, 180, 200 and 220 °C for 10 min. As shown in Figure 1, color of roasted sesame

293

seeds indicated the degree of roasting resulting in yellowish, brownish, brown and

294

dark brown color at 150, 180, 200 and 220 °C, respectively. However, sesame roasted

295

at 220 °C for 10 min had burnt appearance with dark brown color.

296

Table 1 gives L*, a*, b* values obtained for the samples roasted at different

297

temperatures. As represented, L* (lightness) values of sesame seeds tended to

298

decrease as the thermal load increased indicating darker color. However, a* values

299

inclined to increase within the progress of roasting owing to formation of brown


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300

pigments via Maillard reaction. a* values (redness) increased significantly (p < 0.05)

301

for sesame seeds roasted at 150, 180 and 200 °C, whereas a significant (p < 0.05)

302

decrease was observed in a* value for roasted sesame at 220 °C. On the other hand,

303

the values of b* (yellowness) decreased by roasting at temperatures higher than 180

304

°C, indicating the loss of yellowness and production of brownness at those

305

temperatures.

306

Kahyaoğlu and Kaya34 roasted sesame seeds at 120, 150 and 180 °C up to 120 min,

307

and they reported that L* value of sesame seeds constantly increased at 120 °C

308

during roasting, however, a decline in L* values was observed for the seeds roasted

309

at higher roasting temperatures (150 and 180 °C). Additionally, they pointed out that

310

increasing a* values well correlated with decreasing L* values according to roasting

311

conditions. On the contrary to our results, they concluded that b* values increased as

312

a result of increase in roasting temperatures.

313

Figure 2 illustrates HMF formation in sesame seeds roasted at 150, 180, 200 and 220

314

°C for 10 min. Formation of HMF showed an increasing trend during roasting at all

315

temperatures. HMF formation at lower temperatures such as 150 and 180 °C was


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316

relatively less, however, when the temperature was raised to 200 °C, HMF content in

317

sesame seeds significantly (p < 0.05) increased to 30.2 ± 7.7 mg/kg reaching the

318

highest level at 220 °C (39.8 ± 1.1 mg/kg). This level was almost 13 times higher than

319

that of at 150 °C. An increase in temperature promoted the formation of HMF during

320

roasting, as expected. As indicated before, HMF is generally used as a heat load

321

indicator in a wide range of foods such as bakery products, breakfast cereals, coffee

322

and honey.35 Thereby, HMF formation in sesame seeds could also be monitored to

323

investigate the heating intensity applied during roasting. According to the results

324

provided by Agila and Barringer36, increasing both the temperature and time during

325

roasting of almonds showed an increase in HMF concentration, reaching to maximum

326

level, 905 μg/L, in almonds roasted at 177 °C for 20 min. Likewise, it was reported by

327

Fallico37 that HMF content of hazelnuts rapidly increased during progress of roasting

328

at a constant temperature. In a similar study, HMF formation in roasted hazelnuts was

329

highly affected by the increase in roasting time and temperature.38 Since sesame

330

seeds are mostly used in various bakery products, that are subjected to elevated

331

temperatures, formation of HMF in sesame seeds under these conditions should be

332

considered. Considering the serving size of sesame seeds as 10 g (with consumption


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of baked buns, bagels or breads), calculated HMF intake is 0.3 mg, which is almost

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equal to HMF intake by consumption of 1 glass of fruit juice or 20 g of jams.39

335

Effect of roasting on acrylamide levels, which is recognized a probable human

336

carcinogen, were also determined in sesame seeds. As shown in Figure 3, acrylamide

337

formation in sesame seeds was facilitated as a result of increasing the roasting

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temperature from 150 to 200 °C. Acrylamide concentration in sesame seeds roasted

339

at 200 °C for 10 min reached to 633.0 ± 49.4 μg/kg as the highest level. However,

340

acrylamide content in sesame seeds drastically (p < 0.05) decreased to 191.9 ± 22.4

341

μg/kg when roasting temperature was raised to 220 °C. This substantial reduction in

342

acrylamide is the net result of acrylamide formation/elimination kinetics. As it is known,

343

both formation and elimination of acrylamide progress simultaneously during heating

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of foods and observed levels of acrylamide in foods is the net amount. As the amounts

345

of precursors of acrylamide play a key role for the determination of dominant reaction,

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acrylamide formation is dominant in the presence of asparagine or carbonyl

347

compounds, however, its elimination becomes dominant in the lack of any precursor.

348

According to this, during roasting of sesame seeds, acrylamide formation was

349

predominant up to 200 °C, whereas at higher temperatures, acrylamide elimination


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350

became predominant. It was noticed from these results that this type of kinetic pattern

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was similar to roasted coffee.29,40 Kocadağlı et al29 reported that maximum acrylamide

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concentration was 468 μg/kg in coffee beans roasted at 220 °C for 5 min. They

353

concluded that free asparagine was the limiting reactant in green coffee beans, thus,

354

owing to the lack of free asparagine, a rapid decrease in acrylamide content was

355

observed after 5 min roasting. It was also reported that acrylamide was formed in

356

other seeds roasted at elevated temperatures. A study revealed that acrylamide

357

formed in almonds ranging from no detectable levels to 236 μg/kg, when almonds

358

were roasted at 130 °C for 2.5-40 min. However, acrylamide concentration changed

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between 715-1044 μg/kg in almond roasted at 150 °C for 15-30 min.41 For roasted

360

hazelnuts, this level was found ranging between 14-22 μg/kg.42

361

Hereby, acrylamide content in roasted sesame seeds was found as comparably

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higher than common acrylamide rich foods such as potato crisp, snacks and French

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fries (332-550 μg/kg) depending on heating conditions and levels of reaction

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precursors.43

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It is known that free asparagine is the main precursor for occurrence of acrylamide in


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heated foods. For understanding the effect of asparagine on acrylamide formation in

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sesame seeds, free asparagine contents of roasted samples were monitored. As

368

shown in Figure 3, free asparagine content was found to be 207.9 ± 5.6 mg/kg in

369

sesame seeds roasted at 150 °C for 10 min. A dramatic (p < 0.05) decline was

370

observed when the roasting temperature was higher, and, asparagine completely run

371

out in samples roasted at 220 °C. It can be clearly seen from these findings that the

372

changes in asparagine and acrylamide concentration well correlated. Thus, it could

373

be stated that the amount of free asparagine is the limiting precursor for the

374

acrylamide formation during roasting of sesame seeds. Namely, a rapid decrease in

375

acrylamide level of roasted sesame seeds was due to insufficient level of asparagine

376

as the roasting temperature increased.

377

As stated before, acrylamide content of roasted sesame seeds was found almost

378

equal to or higher than mostly consumed acrylamide-rich foods. Even though the

379

amount of sesame seeds consumed with bakery products is comparably low, they

380

also contribute to total intake of dietary acrylamide. In addition, considering that

381

acrylamide is carcinogenic and its metabolite glycidamide is genotoxic, it is reported

382

that a tolerable daily intake level could not be set for acrylamide in food.43 Accordingly,


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contribution of sesame seeds to daily acrylamide intake is a concern.

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Furosine is an early glycation marker, indicating the progress of Maillard reaction.

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Furosine, the compound formed from N-ε-fructosyllysine during acid hydrolysis, was

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analyzed in roasted sesame seeds in order to monitor the progress of Maillard

387

reaction during roasting. Furosine could not be detected in unroasted sesame seeds

388

(data not shown), but its concentration reached to a maximum value of 128.8 ± 3.7

389

mg/kg in sesame seeds roasted at 150 °C for 10 min and it was followed by a

390

decreasing trend when the thermal load increased (Figure 4). The reason of the

391

decline in furosine levels as a result of increasing temperature might be its possible

392

conversion to other compounds. Furosine might be degraded or oxidized to advanced

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glycation end products like CML, during overheating. Similar to our findings, furosine

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concentrations were analyzed in both raw and roasted hazelnuts, and a drastic (p

Investigations on the Maillard reaction in sesame (Sesamum

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