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Production of Maillard reaction flavour precursors by extrusion processing
Food Research Internationul25
(1992) 1755180
Production of Maillard reaction flavour precursors by extrusion processing V. A. Yaylayan, J. Fichtali” & F. R. van de Voort Department qf Food Science und Agricultural Chemistry, McGill University, Mucdonuld Cumpus, St0 Anne de Brllrvuc. Quebec, Canada, H9X I CO
In-situ generation of Maillard reaction flavours by extrusion was investigated using a model system consisting of 15% (w/w) glucose, 5% (w/w) tryptophan and 80% (w/w) inert microcrystalline cellulose under preselected conditions of temperature, moisture, screw speed and feed rate. Samples were analyzed by HPLC for Amadori product, hydroxymethyl furfural and maltol. The effects of the extrusion parameters on product formation were assessed by the generalized distance approach to derive individual and compromise optima and their associated extrusion conditions. Product yields were similar to those obtained by aqueous refluxing, but their rate of formation was -8 times faster. The results indicate that a controlled continuous production of Maillard reaction flavour precursors by extrusion is possible. Keywords: extrusion,
Maillard, optimization,
INTRODUCTION
flavour, Amadori.
tures. There is no information in the literature regarding the application of extrusion processing to producing Maillard reaction flavours per se; most studies deal with the stability of added aroma compounds (Maga & Sizer, 1979; Crouzet et al., 1984). Some work has been done by Ho et uf. (1989) on the formation of volatile compounds in corn-based model systems and determining Maillard reaction products by gas chromatographymass spectrometry (GC-MS). They noted that formation of flavour-related compounds, including pyrazines, were favoured at higher temperatures and lower moisture conditions. Cheftel (1986) indicated that the Maillard reaction is favoured at temperatures > 180°C and higher shear conditions (~100 rev/min) in combination with moisture contents of 1.5% or less. A recent review by Maga (1989) summarizes flavour formation and retention during extrusion. Sugar/amino acid model systems have served as a common means of studying the development of Maillard flavour and color precursors, specifically the Amadori rearrangement product (ARP). The objective of this study was: (i) to assess production and yield of tryptophan ARP and its key degradation products in a model system using the extruder as a continuous reactor; (ii) to assess the effect of key process variables (i.e. temperature,
Extrusion combines various unit operations such as mixing, cooking and texturizing into a single continuous process (Fichtali & van de Voort, 1989), which makes it an attractive option for the food processor and, as a result, it has been exploited extensively by the food industry (Harper, 1981). In terms of flavour, extrusion tends to be limiting because of chemical degradation due to oxidation, hydrolysis and other reactions occurring under high temperature short time (HTST) extruder operating conditions (Maga, 1989) and also volatiles being flashed off at the die. On the other hand, systems containing reducing sugars and proteins or amino acids can rapidly undergo the Maillard reactions (Maga & Kim, 1989), one of the key flavour-producing reactions occurring in food systems. As such, the extruder could serve as a continuous reactor for the production of flavour precursors from selected sugar/amino acid mix-
* Present address: Department of Food Science, BioResource Engineering, University of British Columbia, Vancouver. BC, Canada, V6T lW5 Food Reseurch lnternationulO963-9969/921$05.00 0 1992 Canadian Institute of Food Science and Technology
175
V. A. Yaylayan, J. Fichtali. F. R. van de Voort
176
moisture content, screw speed) on extrusion dependent variables (product temperature, pressure at the die, specific energy consumption) and Maillard reaction product yield; and (iii) to predict the optimum reaction conditions for product formation using the generalized distance approach (Khuri & Conlon, 1981).
MATERIALS
AND METHODS
Tryptophan, D-glucose, hydroxymethyl furfural (HMF), malt01 and microcrystalline cellulose were obtained from Aldrich Chemical Company (Wisconsin, USA) and used without further purification. All the solvents were of HPLC grade (BDH, USA), water was obtained from a Mini-Q reagent grade water system (Millipore Corp., Bedford, MA) and all HPLC mobile phases were degassed under vacuum. Tryptophan Amadori product was synthesized as reported by Yaylayan & Forage (1991) and the extrudates analyzed by HPLC. Extrusion processing and optimization Tryptophan (5 kg), glucose (15 kg) and cellulose (80 kg) were mixed in a vacuum dispersion mixer (Day Mixing, Cincinnati, Ohio, USA) to obtain a homogeneous mass. The moisture content of the mixture was determined to be 4.2% by oven drying (2 h at 60°C). The extruder used was a Baker Perkins MPF-SOD (APV Baker, Inc., Grand Rapids, MI, USA) co-rotating intermeshing twin screw extruder with the barrel configured in a 2O:l L/D ratio (barrel length to screw diameter), and the screw profile was designed to obtain good mixing and conveying performance. The screw configuration in sequence from the feeder to the die consisted of: 300 mm, feed screw; 50 mm, 30” forwarding paddles; 50 mm, short pitch screw; 50 mm, single lead screw; 37.5 mm, 60” forwarding paddles; 37.5 mm, 30” reversing paddles; 100 mm, single lead; 50 mm, 60” forwarding paddles; 37.5 mm, 30” reversing paddles; 100 mm, single lead; 50 mm, 90” paddles; 75 mm, single lead; 37.5 mm, 60” forwarding paddles; and 75 mm, single lead. A die having two 9 mm circular orifices was used and the temperature was controlled over nine zones along the barrel. Pressure and product temperature at the die and torque were measured and the specific energy consumption (SEC) was calculated as follows:
where
where
NHP T OS MS MHP
NHP = T(OS)/(MS)(MHP) net horsepower torque (units) operating speed (rev/min) maximum speed (500 rev/min) machine horsepower
(1)
HSEC = MHP/FR KSEC = HSEC(0.7457)(3600)
(2) (3)
HSEC specific energy consumption (hp h/kg) KSEC specific energy consumption
(kJW FR
feed rate (kg/h)
Preliminary experiments were carried out to determine viable operating ranges (i.e. screw speed, temperature, moisture) from which the basic experimental design was developed, which lays out the operating conditions for the extrusion experiments. A simple coded factorial design varying only moisture and temperature (Table 1) was used for two separate extrusion runs having distinct temperature ranges (70-90 and 80-100°C) but common moisture variation (55-63X), with the feed rate and screw speed held constant. A separate experiment was carried out to study the effect of screw speed. The extruder was run at each set of design conditions until the operating conditions stabilized. Samples were then taken after cooling, placed in plastic bags and frozen at -25°C until analyzed. The response data (i.e. the concentration of ARP, malt01 and HMF produced) was then used to determine the optimal conditions for their formation by the generalized distance approach (GDA) using a design radius of 1.O (Khuri & Conlon, 1981; Khuri & Cornell, 1987). Compromise optima in turn were obtained from the GDA data using the multiple response (MR) program developed by Conlon & Khuri (1988). Table 1. Coded (X) and real values of moisture (&I) and temperature (T,,J used in the experiment design for producing Maillard reaction products Xl 1 -1
I -1 1 -1 0 0 0
M(x) 59 55 63 55 63 55 59 59 59
x2, -1 -1 1 1 0 0 -1 1 0
T,(“C)
X2,
T,(“C)
70 70 90 90 80 80 70 90 80
-1
80 80 100 100 90 90 80 100 90
-1
1 1 0 0 -1 1 0
177
Production of Maillard reaction JEavour precursors by extrusion processing
HPLC analysis Extrudate samples were taken under selected conditions of temperature, moisture content, screw speed and feed rate for analysis of HMF, maltol and ARP. Two grams of each sample taken were diluted to 100 ml with distilled water and filtered through a 0.45-pm, type HA Millipore filter (Waters Scientific) before injection on to the HPLC column equipped with a Rheodyne injector having a 20-~1 loop. A Beckman System Gold Modular HPLC was used for analyses, consisting of a Model 166 variable wavelength UV detector set at 280 nm and a model 110B solvent delivery system controlled by a NEC lap-top computer and connected to a Shimadzu CR- 18 integrator (IO-mV full scale). Analyte HPLC separation was carried out using either a 5 m, 2.0 X 150 mm Beckman C18 Ultrasphere column or a Merck 5 m, 2.0 X 150 mm C- 18 Lichrosphere, 1OO-RP-18 column, equipped with a Licro CART 4-4 guard column. Both columns were operated at ambient temperature and 20 ~1 were injected for analysis; the values obtained represented the average of three injections. The analyte peaks were identified and confirmed by comparison of their retention times with commercial or synthesized standards using two mobile phases, wavelength ratioing and spiking (for details see Yaylayan & Forage, 1991). Quantitation was performed by the use of appropriate calibration curves.
tures and low moisture conditions, as suggested by Cheftel (1986); however, preliminary trials indicated that viable extrusion conditions could only be obtained using >55’%1moisture and temperatures lower than 110°C. These limiting conditions were likely due to the effect of the microcrystalline cellulose matrix (-80%) rather than the amino acid/sugar mixture. Based on the experimental design conditions specified in Table 1, barrel temperature did not affect torque, pressure at the die or specific energy consumption (SEC); however, moisture was a limiting variable (Table 2). Temperature, which normally affects viscosity, has no effect in this system as cellulose is insoluble and does not melt. Moisture acts as a lubricant, reducing torque, pressure and energy consumption. The effect of screw speed on torque, SEC and pressure was studied Table 2. Average torque, pressure, specific energy consumption (SEC) and die product temperature values obtained over the temperature range of 70-100°C (screw speed = 150 revlmin, feed rate = 20 kg/h) Moisture (‘I%)
Torque (‘X,)
Pressure (lb/in’)
55
25
800
59 63
11 8.5
(1dT = difference temperature
between setting.
SEC
dFI
(kJ/kg)
(“Cl
200 200
251 110 86
+8 +5
die product
temperature
and barrel
I
-Torque + SEC
RESULTS AND DISCUSSION
m
xl00 x2
m Pressure
m
m
Processing conditions The model system chosen was tryptophan/glucose, which has been studied previously by the authors (Yaylayan & Forage, 1991, 1992) in an aqueous matrix. Because of the extensive raw material requirements and inherent expense imposed by full scale extrusion experimentation, the sugar/amino acid mixture was diluted with microcrystalline cellulose which does not interfere with the Maillard reaction (Eichner & Karel, 1972) and is insoluble in water, making it simple to extract the reaction products from the extrudate. Tryptophan Amadori compound, a reactive intermediate, and its breakdown products, HMF and maltol, were used as indicator compounds of the extent of the Maillard reaction. Initial experimental designs were predicated on using relatively high tempera-
Screw
speed
(rev I min)
Fig. 1. The effect of screw speed on percent torque, pressure at the die and specific energy consumption (temperature = 87”C, feed rate = 20 kg/h, moisture = 55%).
178
V. A. Yaylayan, J. Fichtali. F. R. van de Voort -
ARP
+ MAL O-08
t
;: 0.06 al .’ Z .T?
-
m HMF
\
5 ‘2
‘\\
\
\
\
\
\ ‘+_______--+, \
0.04
-
/
5
*
fi&
5 s 0
\ 0.02
\ \ , ‘+
-
m
x
m
’ ,’
m
m
t 100
140
180
Screw
220
260
300
speed (rev/ min)
Fig. 2. The effect of screw speed on the production of ARP, HMF and maltol (temperature = 86°C feed rate = 20 Kg/h, moisture = 55%).
independently and is illustrated in Fig. 1. Although the torque decreases exponentially as screw speed increases, SEC reaches a minimum around 200 rev/min while the pressure tends to maximize at the screw speed. Screw speed was also studied (Fig. 1) and has a complex effect on torque, specific energy and pressure, which were related to Maillard product formation (Fig. 2). Since the screw speed effects on product formation were neither linear nor quadratic, they could not be included as a variable in the multiresponse optimization procedure. ARP and HMF are generally independent of screw speed, while maximum malt01 production was obtained at minimum revolutions per minute, corresponding to a longer residence (reaction) time in the extruder. Additional malt01 formation would require further screw speed reductions or the application of higher temperatures. Maillard reaction
The Maillard reaction, known to be an important flavor-producing reaction in food systems, is usually associated with baked and roasted products, and is used commercially by the flavour industry to produce reaction flavours (Reynolds, 1970). The flavour characteristics produced depend primarily on the precise nature and ratio of the amino acids
and sugars, reaction conditions (primarily time, temperature and moisture) and the nature of any thermal degradation reactions that may contribute to the overall flavour profile (Macleod & Seyeddain-Ardebili, 1981). Some of the steps of the reaction mechanism are well known, based on model system studies; however the accurate production of flavours produced is still largely an art. The reaction kinetics of glucose with tryptophan in an aqueous system to form tryptophan Amadori compound and its subsequent decomposition products HMF and malt01 has been studied (Yaylayan & Forage, 1991). Refluxing a 3: 1 mixture of glucose/ tryptophan at 110°C for 6 h in water yields -1.7% ARP, 0.15% HMF and 4.3% maltol, based on the initial tryptophan concentration. The main source of HMF and malt01 in this reaction mixture is the tryptophan Amadori compound, although they may also be formed at much slower rates from the decomposition of glucose alone (Yaylayan & Forage, 1991). ARP decomposition basically takes place via two well-established pathways, 1,2-enolization producing mainly HMF, leading to browning, while 2,3-enolization produces maltol, leading mainly to flavour formation (Reynolds, 1970). Hence the concentrations of ARP, HMF and malt01 produced are interrelated, ARP accumulation and decomposition being a dynamic process, and the optimization of any one compound automatically implies reduced levels of the others. distance approach Using the generalized (GDA), one can predict the optimum concentrations and processing conditions for each component from the concentrations of the selected Maillard intermediates and the experimental design process conditions used. The mathematical basis for this procedure was developed over a decade ago (Khuri & Conlon, 1981); however, it has only recently been applied to assist in the development and optimization of multivariate extrusion processes (Fichtali et al., 1990~1). GDA permits one to find compromise conditions for the input variables that are somewhat favourable to all responses. This implies that the multiresponse function deviates as little as possible from the individual optima and the deviation is formulated as a distance function which is minimized over the experiment region. By evaluating the Maillard reaction system by this means, one can determine whether the processing conditions chosen favour the 1,2- or 2,3-enolization degradation pathway based on whether HMF or malt01 is dominant. The method also permits the prediction of compromise optimum
Production of Maillard reactionJlavour precursors by extrusion processing
conditions where all components are optimized simultaneously, by formulating responses as a distance function minimized over the experimental region. Table 3 provides a summary of the optimum concentrations and yield for each Maillard component and the respective processing conditions for each of the two temperature sets. It can be seen that in both temperature ranges ARP development is optimized at lower temperatures, although the optimum shifts from 70 to 80°C in the higher temperature range and the yield drops slightly. HMF production is favoured at about 16°C above the ARP optimum in both temperature ranges, while malt01 production is favoured at a temperature about 34°C lower than HMF production. Slightly higher moisture contents favour the production of ARP, while lower values favour HMF and maltol. Malt01 shows the largest increase, in the second temperature set, with the yield almost doubling, indicating the ARP is decomposing preferably to maltol, reducing the total ARP present. The yield data are quite good relative to the conventional aqueous reaction studied, basically producing 24 g ARP/h kg tryptophan (70-90°C range) in the continuous extrusion process vs. 17 g in 6 h or -2.83 g/h kg using the conventional batch reflux procedure. ARP can also be produced by refluxing a 3:l mixture of glucose and tryptophan in methanol for 4 h with roughly a 20% yield (-50 g/h kg tryptophan). Although extrudate yield is roughly half of that of the methanol-based reaction, one must keep in mind that extruder throughput was limited by the fact that 80% of the raw material was inert filler rather than the reaction mixture. Assuming similar yields Table 3. Optimum concentrations calculated by the generalized distance approach for each Maillard reaction component and the respective processing conditions required to produce the optima listed Response
Optimum (g/liter) Temperature
ARPl HMFh MALc
0.04760 0~00116 0~03880
ARP HMF MAL
0.03450 0.00 138 0.06970
Temperature
Yield (‘X1)
Moisture (‘X1)
Temperature (“C)
range 7OG90°C 2.70 0.16 5.50
58.6 55.9 55.3
70.0 86.3 83.4
range 80-100°C 1.95 0.19 9.87
58.6 56.1 55.2
u ARP = Amadori rearrangement product. h HMF = hydroxymethyl furfural. ( MAL = maltol
80.0 97.2 93.3
179
Table 4. Compromise optima calculated for the two temperature sets and the associated extrusion conditions required Response
Compromise 70-90°C
Temperature (“C) Moisture (‘%I) ARP (g/liter) HMF (g/liter) MAL (g/liter)
72.9 56.2 0.04200 0.00098 0.03 170
Yield (‘i/;l)
2.40 0.14 4.49
optimum
(g/liter)
80-I 00°C
97.2 56.2 0~03100 0.00 I 38 0.0631
Yield (‘X1)
I.75 0.19 8.94
without the filler at a throughput of 20 kg/h (3:l sugar/amino acid) the yield would be -125 g/h. Table 4 presents the calculated compromise optimum data, where all components are optimized simultaneously. In this situation, one is balancing the production of the three components against each other; however, similar trends are observed as before: ARP dropping somewhat in the higher temperature range, malt01 doubling and HMF increasing, but being produced only in relatively small amounts. Moisture and temperature are but two of many variables that can be controlled during the extrusion process (Fichtali & van de Voort, 1989) and the GDA procedure has been successfully applied to develop and optimize the production of sodium caseinate for three product characteristics dictated by six operating variables simultaneously (Fichtali & van de Voort, 1990; Fichtali et al., 1990h,c). As such, the GDA has been proven to be a very useful optimization technique; however, the data derived therefrom is only valid within the original experimental design parameters and cannot be extrapolated.
CONCLUSION A twin screw extruder can be used as a continuous reactor for the Maillard reaction. The composition of reaction products can be based on the judicious adjustment of extrusion conditions to optimize the formation of ARP, HMF and malt01 individually or simultaneously as a compromise optimum. One limitation of this work was the need to use microcrystalline cellulose as a diluent, which strongly affected the extruder operating conditions (torque, specific energy and pressure). It is unlikely that the basic reaction pattern would change dramatically without the filler if run under the same conditions; however, the elimination of
180
V. A. Yaylayan, J. Fichtali, F. R. van de Voort
the filler would undoubtedly expand the range of operating conditions (i.e. temperature and moisture), which could increase yield and provide more direction to controlling the reaction. In order to elucidate whether there would be commercial potential for the production of Maillard reaction intermediates for any particular sugar/amino acid mixture, additional work should be carried out, without the diluent, including a reaction kinetic study that requires the assessment of the residence time distribution in the extruder. Albeit somewhat limited, sufficient information has been obtained to confirm the basic premise that Maillard flavour intermediates may be produced by extrusion processing and that some control can be exercised over the reaction by manipulating the extruder parameters.
ACKNOWLEDGMENTS The authors gratefully acknowledge funding support from Conseil des recherches en p&he et en agro-alimentaire du Quebec (CORPAC MCA 2567) for this research. The authors also would like to thank the Food Research and Development Center of Agriculture Canada (Ste Hyacinthe, Quebec) for access to the extruder.
REFERENCES Cheftel, J. C. (1986). Nutritional
effects of extrusion-cooking.
Food Chem., 20,263-83.
Conlon, M. & Khuri, A. I. (1988). MR multiple response optimization. Technical report 322, Department of Statistics, University of Florida. Crouzet, J., Sadafian, A., Doko, B. & Chouvel, M. (1984). Retention des composes volatils au tours de la cuisson-extrusion. In Thermal processing and Quality of Foods. Elsevier Applied Science Publishers, pp. 212-60. Eichner, K. & Karel, M. (1972). The influence of water content and water activity on the sugar-amino browning reaction in model systems under various conditions. J. Agr. Food Chem., 20, 218-23.
Fichtali, J. & van de Voort, F. R. (1989). Fundamental and practical aspects of twin screw extrusion. Cereal Foods World, 34, 921-9.
Fichtali, J. & van de Voort, F. R. (1990). Pilot plant production of caseins using extrusion processing. I. Acid casein production. Milchwissenschaft, 45, 56011. Fichtali, J., van de Voort, F. R. & Khuri, A. I. (1990a). Multiresponse optimization of acid casein production. J. Food Proc. Eng., 12, 247-58.
Fichtali, J., van de Voort, F. R. & Toupin, C. J. (1990b). Effect of coagulation and washing conditions on fines, water holding capacity and microstructure of acid casein curd. J. Dairy Res., 57, 527740. Fichtali, J., van de Voort, F. R. & Toupin, C. J. (1990~). Coagulation and washing conditions for acid casein production from skim milk powder. Int. J. Food Sci. Technol., 25, 377-88.
Harper, H. M. (1981). Exfrusion of Foods, Vol. I. CRC Press, Boca Raton, FL. Ho, C. T., Bruechert, L. J., Kuo, M. C. & Izzo, M. (1989). Formation of volatile compounds from extruded cornbased model systems. In Thermal Generation of Aromas, ed. T. Parliment, R. McGorrin & C. T. Ho. American Chemical Society, Washington, DC, pp. 504-l 1. Khuri, A. I. & Conlon, M. (1981). Simultaneous optimization of multiple responses represented by polynomial regression functions. Technometrics, 23, 363-75. Khuri, A. I. & Cornell, J. A. (1987). Response Surfaces, Designs and Analysis. Marcel Dekker, New York, NY. Macleod, G. & Seyeddain-Ardebili, M. (1981). Natural and simulated meat flavors (with particular reference to beef). CRC Crit. Rev. Food Sci. Nutr., 14, 3099437.
Maga, J. A. (1989). Flavor formation and retention during extrusion. In Extrusion Cooking, ed. C. Mercier, P. Linko & J. Harper. American Association of Cereal Chemists, Minnesota, p. 387798. Maga, J. A. & Kim, C. H. (1989). Protein-generated extrusion flavors. In Thermal Generation of Aromas, ed. T. Parliment, R. McGorrin & C. T. Ho. American Chemical Society, Washington, DC, pp. 494-503. Maga, J. A. & Sizer, C. E. (1979). Pyrazine formation during the extrusion of potato flakes. Lebensm. Wiss. Technol., 12, 15-21.
Reynolds, T. M. (1970). Flavors from nonenzymic browning reactions. Food Tech. Aust., 22, 610-19. Yaylayan, V. & Forage, N. (1991). Determination of the kinetics and mechanism of decomposition of tryptophan Amadori rearrangement products by RP-HPLC analysis. J. Agric. Food Chem., 39, 364-9.
Yaylayan, V. & Forage, N. (1992). A kinetic model for the reaction of tryptophan with glucose and mannose - the role of diglycation in the Maillard reaction. Food Chem., 44, 201-8.
(1992) 1755180
Production of Maillard reaction flavour precursors by extrusion processing V. A. Yaylayan, J. Fichtali” & F. R. van de Voort Department qf Food Science und Agricultural Chemistry, McGill University, Mucdonuld Cumpus, St0 Anne de Brllrvuc. Quebec, Canada, H9X I CO
In-situ generation of Maillard reaction flavours by extrusion was investigated using a model system consisting of 15% (w/w) glucose, 5% (w/w) tryptophan and 80% (w/w) inert microcrystalline cellulose under preselected conditions of temperature, moisture, screw speed and feed rate. Samples were analyzed by HPLC for Amadori product, hydroxymethyl furfural and maltol. The effects of the extrusion parameters on product formation were assessed by the generalized distance approach to derive individual and compromise optima and their associated extrusion conditions. Product yields were similar to those obtained by aqueous refluxing, but their rate of formation was -8 times faster. The results indicate that a controlled continuous production of Maillard reaction flavour precursors by extrusion is possible. Keywords: extrusion,
Maillard, optimization,
INTRODUCTION
flavour, Amadori.
tures. There is no information in the literature regarding the application of extrusion processing to producing Maillard reaction flavours per se; most studies deal with the stability of added aroma compounds (Maga & Sizer, 1979; Crouzet et al., 1984). Some work has been done by Ho et uf. (1989) on the formation of volatile compounds in corn-based model systems and determining Maillard reaction products by gas chromatographymass spectrometry (GC-MS). They noted that formation of flavour-related compounds, including pyrazines, were favoured at higher temperatures and lower moisture conditions. Cheftel (1986) indicated that the Maillard reaction is favoured at temperatures > 180°C and higher shear conditions (~100 rev/min) in combination with moisture contents of 1.5% or less. A recent review by Maga (1989) summarizes flavour formation and retention during extrusion. Sugar/amino acid model systems have served as a common means of studying the development of Maillard flavour and color precursors, specifically the Amadori rearrangement product (ARP). The objective of this study was: (i) to assess production and yield of tryptophan ARP and its key degradation products in a model system using the extruder as a continuous reactor; (ii) to assess the effect of key process variables (i.e. temperature,
Extrusion combines various unit operations such as mixing, cooking and texturizing into a single continuous process (Fichtali & van de Voort, 1989), which makes it an attractive option for the food processor and, as a result, it has been exploited extensively by the food industry (Harper, 1981). In terms of flavour, extrusion tends to be limiting because of chemical degradation due to oxidation, hydrolysis and other reactions occurring under high temperature short time (HTST) extruder operating conditions (Maga, 1989) and also volatiles being flashed off at the die. On the other hand, systems containing reducing sugars and proteins or amino acids can rapidly undergo the Maillard reactions (Maga & Kim, 1989), one of the key flavour-producing reactions occurring in food systems. As such, the extruder could serve as a continuous reactor for the production of flavour precursors from selected sugar/amino acid mix-
* Present address: Department of Food Science, BioResource Engineering, University of British Columbia, Vancouver. BC, Canada, V6T lW5 Food Reseurch lnternationulO963-9969/921$05.00 0 1992 Canadian Institute of Food Science and Technology
175
V. A. Yaylayan, J. Fichtali. F. R. van de Voort
176
moisture content, screw speed) on extrusion dependent variables (product temperature, pressure at the die, specific energy consumption) and Maillard reaction product yield; and (iii) to predict the optimum reaction conditions for product formation using the generalized distance approach (Khuri & Conlon, 1981).
MATERIALS
AND METHODS
Tryptophan, D-glucose, hydroxymethyl furfural (HMF), malt01 and microcrystalline cellulose were obtained from Aldrich Chemical Company (Wisconsin, USA) and used without further purification. All the solvents were of HPLC grade (BDH, USA), water was obtained from a Mini-Q reagent grade water system (Millipore Corp., Bedford, MA) and all HPLC mobile phases were degassed under vacuum. Tryptophan Amadori product was synthesized as reported by Yaylayan & Forage (1991) and the extrudates analyzed by HPLC. Extrusion processing and optimization Tryptophan (5 kg), glucose (15 kg) and cellulose (80 kg) were mixed in a vacuum dispersion mixer (Day Mixing, Cincinnati, Ohio, USA) to obtain a homogeneous mass. The moisture content of the mixture was determined to be 4.2% by oven drying (2 h at 60°C). The extruder used was a Baker Perkins MPF-SOD (APV Baker, Inc., Grand Rapids, MI, USA) co-rotating intermeshing twin screw extruder with the barrel configured in a 2O:l L/D ratio (barrel length to screw diameter), and the screw profile was designed to obtain good mixing and conveying performance. The screw configuration in sequence from the feeder to the die consisted of: 300 mm, feed screw; 50 mm, 30” forwarding paddles; 50 mm, short pitch screw; 50 mm, single lead screw; 37.5 mm, 60” forwarding paddles; 37.5 mm, 30” reversing paddles; 100 mm, single lead; 50 mm, 60” forwarding paddles; 37.5 mm, 30” reversing paddles; 100 mm, single lead; 50 mm, 90” paddles; 75 mm, single lead; 37.5 mm, 60” forwarding paddles; and 75 mm, single lead. A die having two 9 mm circular orifices was used and the temperature was controlled over nine zones along the barrel. Pressure and product temperature at the die and torque were measured and the specific energy consumption (SEC) was calculated as follows:
where
where
NHP T OS MS MHP
NHP = T(OS)/(MS)(MHP) net horsepower torque (units) operating speed (rev/min) maximum speed (500 rev/min) machine horsepower
(1)
HSEC = MHP/FR KSEC = HSEC(0.7457)(3600)
(2) (3)
HSEC specific energy consumption (hp h/kg) KSEC specific energy consumption
(kJW FR
feed rate (kg/h)
Preliminary experiments were carried out to determine viable operating ranges (i.e. screw speed, temperature, moisture) from which the basic experimental design was developed, which lays out the operating conditions for the extrusion experiments. A simple coded factorial design varying only moisture and temperature (Table 1) was used for two separate extrusion runs having distinct temperature ranges (70-90 and 80-100°C) but common moisture variation (55-63X), with the feed rate and screw speed held constant. A separate experiment was carried out to study the effect of screw speed. The extruder was run at each set of design conditions until the operating conditions stabilized. Samples were then taken after cooling, placed in plastic bags and frozen at -25°C until analyzed. The response data (i.e. the concentration of ARP, malt01 and HMF produced) was then used to determine the optimal conditions for their formation by the generalized distance approach (GDA) using a design radius of 1.O (Khuri & Conlon, 1981; Khuri & Cornell, 1987). Compromise optima in turn were obtained from the GDA data using the multiple response (MR) program developed by Conlon & Khuri (1988). Table 1. Coded (X) and real values of moisture (&I) and temperature (T,,J used in the experiment design for producing Maillard reaction products Xl 1 -1
I -1 1 -1 0 0 0
M(x) 59 55 63 55 63 55 59 59 59
x2, -1 -1 1 1 0 0 -1 1 0
T,(“C)
X2,
T,(“C)
70 70 90 90 80 80 70 90 80
-1
80 80 100 100 90 90 80 100 90
-1
1 1 0 0 -1 1 0
177
Production of Maillard reaction JEavour precursors by extrusion processing
HPLC analysis Extrudate samples were taken under selected conditions of temperature, moisture content, screw speed and feed rate for analysis of HMF, maltol and ARP. Two grams of each sample taken were diluted to 100 ml with distilled water and filtered through a 0.45-pm, type HA Millipore filter (Waters Scientific) before injection on to the HPLC column equipped with a Rheodyne injector having a 20-~1 loop. A Beckman System Gold Modular HPLC was used for analyses, consisting of a Model 166 variable wavelength UV detector set at 280 nm and a model 110B solvent delivery system controlled by a NEC lap-top computer and connected to a Shimadzu CR- 18 integrator (IO-mV full scale). Analyte HPLC separation was carried out using either a 5 m, 2.0 X 150 mm Beckman C18 Ultrasphere column or a Merck 5 m, 2.0 X 150 mm C- 18 Lichrosphere, 1OO-RP-18 column, equipped with a Licro CART 4-4 guard column. Both columns were operated at ambient temperature and 20 ~1 were injected for analysis; the values obtained represented the average of three injections. The analyte peaks were identified and confirmed by comparison of their retention times with commercial or synthesized standards using two mobile phases, wavelength ratioing and spiking (for details see Yaylayan & Forage, 1991). Quantitation was performed by the use of appropriate calibration curves.
tures and low moisture conditions, as suggested by Cheftel (1986); however, preliminary trials indicated that viable extrusion conditions could only be obtained using >55’%1moisture and temperatures lower than 110°C. These limiting conditions were likely due to the effect of the microcrystalline cellulose matrix (-80%) rather than the amino acid/sugar mixture. Based on the experimental design conditions specified in Table 1, barrel temperature did not affect torque, pressure at the die or specific energy consumption (SEC); however, moisture was a limiting variable (Table 2). Temperature, which normally affects viscosity, has no effect in this system as cellulose is insoluble and does not melt. Moisture acts as a lubricant, reducing torque, pressure and energy consumption. The effect of screw speed on torque, SEC and pressure was studied Table 2. Average torque, pressure, specific energy consumption (SEC) and die product temperature values obtained over the temperature range of 70-100°C (screw speed = 150 revlmin, feed rate = 20 kg/h) Moisture (‘I%)
Torque (‘X,)
Pressure (lb/in’)
55
25
800
59 63
11 8.5
(1dT = difference temperature
between setting.
SEC
dFI
(kJ/kg)
(“Cl
200 200
251 110 86
+8 +5
die product
temperature
and barrel
I
-Torque + SEC
RESULTS AND DISCUSSION
m
xl00 x2
m Pressure
m
m
Processing conditions The model system chosen was tryptophan/glucose, which has been studied previously by the authors (Yaylayan & Forage, 1991, 1992) in an aqueous matrix. Because of the extensive raw material requirements and inherent expense imposed by full scale extrusion experimentation, the sugar/amino acid mixture was diluted with microcrystalline cellulose which does not interfere with the Maillard reaction (Eichner & Karel, 1972) and is insoluble in water, making it simple to extract the reaction products from the extrudate. Tryptophan Amadori compound, a reactive intermediate, and its breakdown products, HMF and maltol, were used as indicator compounds of the extent of the Maillard reaction. Initial experimental designs were predicated on using relatively high tempera-
Screw
speed
(rev I min)
Fig. 1. The effect of screw speed on percent torque, pressure at the die and specific energy consumption (temperature = 87”C, feed rate = 20 kg/h, moisture = 55%).
178
V. A. Yaylayan, J. Fichtali. F. R. van de Voort -
ARP
+ MAL O-08
t
;: 0.06 al .’ Z .T?
-
m HMF
\
5 ‘2
‘\\
\
\
\
\
\ ‘+_______--+, \
0.04
-
/
5
*
fi&
5 s 0
\ 0.02
\ \ , ‘+
-
m
x
m
’ ,’
m
m
t 100
140
180
Screw
220
260
300
speed (rev/ min)
Fig. 2. The effect of screw speed on the production of ARP, HMF and maltol (temperature = 86°C feed rate = 20 Kg/h, moisture = 55%).
independently and is illustrated in Fig. 1. Although the torque decreases exponentially as screw speed increases, SEC reaches a minimum around 200 rev/min while the pressure tends to maximize at the screw speed. Screw speed was also studied (Fig. 1) and has a complex effect on torque, specific energy and pressure, which were related to Maillard product formation (Fig. 2). Since the screw speed effects on product formation were neither linear nor quadratic, they could not be included as a variable in the multiresponse optimization procedure. ARP and HMF are generally independent of screw speed, while maximum malt01 production was obtained at minimum revolutions per minute, corresponding to a longer residence (reaction) time in the extruder. Additional malt01 formation would require further screw speed reductions or the application of higher temperatures. Maillard reaction
The Maillard reaction, known to be an important flavor-producing reaction in food systems, is usually associated with baked and roasted products, and is used commercially by the flavour industry to produce reaction flavours (Reynolds, 1970). The flavour characteristics produced depend primarily on the precise nature and ratio of the amino acids
and sugars, reaction conditions (primarily time, temperature and moisture) and the nature of any thermal degradation reactions that may contribute to the overall flavour profile (Macleod & Seyeddain-Ardebili, 1981). Some of the steps of the reaction mechanism are well known, based on model system studies; however the accurate production of flavours produced is still largely an art. The reaction kinetics of glucose with tryptophan in an aqueous system to form tryptophan Amadori compound and its subsequent decomposition products HMF and malt01 has been studied (Yaylayan & Forage, 1991). Refluxing a 3: 1 mixture of glucose/ tryptophan at 110°C for 6 h in water yields -1.7% ARP, 0.15% HMF and 4.3% maltol, based on the initial tryptophan concentration. The main source of HMF and malt01 in this reaction mixture is the tryptophan Amadori compound, although they may also be formed at much slower rates from the decomposition of glucose alone (Yaylayan & Forage, 1991). ARP decomposition basically takes place via two well-established pathways, 1,2-enolization producing mainly HMF, leading to browning, while 2,3-enolization produces maltol, leading mainly to flavour formation (Reynolds, 1970). Hence the concentrations of ARP, HMF and malt01 produced are interrelated, ARP accumulation and decomposition being a dynamic process, and the optimization of any one compound automatically implies reduced levels of the others. distance approach Using the generalized (GDA), one can predict the optimum concentrations and processing conditions for each component from the concentrations of the selected Maillard intermediates and the experimental design process conditions used. The mathematical basis for this procedure was developed over a decade ago (Khuri & Conlon, 1981); however, it has only recently been applied to assist in the development and optimization of multivariate extrusion processes (Fichtali et al., 1990~1). GDA permits one to find compromise conditions for the input variables that are somewhat favourable to all responses. This implies that the multiresponse function deviates as little as possible from the individual optima and the deviation is formulated as a distance function which is minimized over the experiment region. By evaluating the Maillard reaction system by this means, one can determine whether the processing conditions chosen favour the 1,2- or 2,3-enolization degradation pathway based on whether HMF or malt01 is dominant. The method also permits the prediction of compromise optimum
Production of Maillard reactionJlavour precursors by extrusion processing
conditions where all components are optimized simultaneously, by formulating responses as a distance function minimized over the experimental region. Table 3 provides a summary of the optimum concentrations and yield for each Maillard component and the respective processing conditions for each of the two temperature sets. It can be seen that in both temperature ranges ARP development is optimized at lower temperatures, although the optimum shifts from 70 to 80°C in the higher temperature range and the yield drops slightly. HMF production is favoured at about 16°C above the ARP optimum in both temperature ranges, while malt01 production is favoured at a temperature about 34°C lower than HMF production. Slightly higher moisture contents favour the production of ARP, while lower values favour HMF and maltol. Malt01 shows the largest increase, in the second temperature set, with the yield almost doubling, indicating the ARP is decomposing preferably to maltol, reducing the total ARP present. The yield data are quite good relative to the conventional aqueous reaction studied, basically producing 24 g ARP/h kg tryptophan (70-90°C range) in the continuous extrusion process vs. 17 g in 6 h or -2.83 g/h kg using the conventional batch reflux procedure. ARP can also be produced by refluxing a 3:l mixture of glucose and tryptophan in methanol for 4 h with roughly a 20% yield (-50 g/h kg tryptophan). Although extrudate yield is roughly half of that of the methanol-based reaction, one must keep in mind that extruder throughput was limited by the fact that 80% of the raw material was inert filler rather than the reaction mixture. Assuming similar yields Table 3. Optimum concentrations calculated by the generalized distance approach for each Maillard reaction component and the respective processing conditions required to produce the optima listed Response
Optimum (g/liter) Temperature
ARPl HMFh MALc
0.04760 0~00116 0~03880
ARP HMF MAL
0.03450 0.00 138 0.06970
Temperature
Yield (‘X1)
Moisture (‘X1)
Temperature (“C)
range 7OG90°C 2.70 0.16 5.50
58.6 55.9 55.3
70.0 86.3 83.4
range 80-100°C 1.95 0.19 9.87
58.6 56.1 55.2
u ARP = Amadori rearrangement product. h HMF = hydroxymethyl furfural. ( MAL = maltol
80.0 97.2 93.3
179
Table 4. Compromise optima calculated for the two temperature sets and the associated extrusion conditions required Response
Compromise 70-90°C
Temperature (“C) Moisture (‘%I) ARP (g/liter) HMF (g/liter) MAL (g/liter)
72.9 56.2 0.04200 0.00098 0.03 170
Yield (‘i/;l)
2.40 0.14 4.49
optimum
(g/liter)
80-I 00°C
97.2 56.2 0~03100 0.00 I 38 0.0631
Yield (‘X1)
I.75 0.19 8.94
without the filler at a throughput of 20 kg/h (3:l sugar/amino acid) the yield would be -125 g/h. Table 4 presents the calculated compromise optimum data, where all components are optimized simultaneously. In this situation, one is balancing the production of the three components against each other; however, similar trends are observed as before: ARP dropping somewhat in the higher temperature range, malt01 doubling and HMF increasing, but being produced only in relatively small amounts. Moisture and temperature are but two of many variables that can be controlled during the extrusion process (Fichtali & van de Voort, 1989) and the GDA procedure has been successfully applied to develop and optimize the production of sodium caseinate for three product characteristics dictated by six operating variables simultaneously (Fichtali & van de Voort, 1990; Fichtali et al., 1990h,c). As such, the GDA has been proven to be a very useful optimization technique; however, the data derived therefrom is only valid within the original experimental design parameters and cannot be extrapolated.
CONCLUSION A twin screw extruder can be used as a continuous reactor for the Maillard reaction. The composition of reaction products can be based on the judicious adjustment of extrusion conditions to optimize the formation of ARP, HMF and malt01 individually or simultaneously as a compromise optimum. One limitation of this work was the need to use microcrystalline cellulose as a diluent, which strongly affected the extruder operating conditions (torque, specific energy and pressure). It is unlikely that the basic reaction pattern would change dramatically without the filler if run under the same conditions; however, the elimination of
180
V. A. Yaylayan, J. Fichtali, F. R. van de Voort
the filler would undoubtedly expand the range of operating conditions (i.e. temperature and moisture), which could increase yield and provide more direction to controlling the reaction. In order to elucidate whether there would be commercial potential for the production of Maillard reaction intermediates for any particular sugar/amino acid mixture, additional work should be carried out, without the diluent, including a reaction kinetic study that requires the assessment of the residence time distribution in the extruder. Albeit somewhat limited, sufficient information has been obtained to confirm the basic premise that Maillard flavour intermediates may be produced by extrusion processing and that some control can be exercised over the reaction by manipulating the extruder parameters.
ACKNOWLEDGMENTS The authors gratefully acknowledge funding support from Conseil des recherches en p&he et en agro-alimentaire du Quebec (CORPAC MCA 2567) for this research. The authors also would like to thank the Food Research and Development Center of Agriculture Canada (Ste Hyacinthe, Quebec) for access to the extruder.
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Yaylayan, V. & Forage, N. (1992). A kinetic model for the reaction of tryptophan with glucose and mannose - the role of diglycation in the Maillard reaction. Food Chem., 44, 201-8.