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Lipid migration in two-phase chocolate systems investigated by NMR and DSC
Food Research International 35 (2002) 761–767 www.elsevier.com/locate/foodres
Lipid migration in two-phase chocolate systems investigated by NMR and DSC§ Peggy Walter, Paul Cornillon* Purdue University-Department of Food Science, Whistler Center for Carbohydrate Research, 1160 Food Science Building-West Lafayette, IN 47907-1160, USA Received 30 September 2000; accepted 6 December 2001
Abstract The migration of lipids in two-phase chocolate systems (i.e. lauric acid+chocolate and peanut butter+chocolate) was analyzed by magnetic resonance and differential scanning calorimetry. Kinetics of fat migration was evaluated and the diffusion coefficient of lauric acid in chocolate was found to be dependent on migration time. This may be due to the capillary nature of fat migration in chocolate. Fat bloom characteristics were determined and related to the thermal history of samples and the presence of fat in the chocolate layer. Both lauric acid and peanut butter increased the liquid-to-solid ratio of chocolate and helped prevent fat bloom. # 2002 Elsevier Science Ltd. All rights reserved. Keywords: Chocolate; DSC; Fat bloom; Fat migration; MRI; Relaxation times
1. Introduction Fat bloom is an important issue for chocolate confectioners. Without proper tempering, it can appear on chocolate samples stored at room temperature after different thermal histories. This physical defect occurred in less than 2 days after storage at 21 C with no tempering prior to cooling (Walter & Cornillon, 2001). The lack of gloss and the presence of white areas that characterize fat bloom often occur for confectionery products containing a layer of chocolate and one or more layers of fat fillings. To prevent fat bloom, additives like triglycerides (e.g. B.O.B. or 1–3 dibehenoyl, 2-oleoylglycerol) could be used (Hachiya, Koyano, & Sato, 1989; Walter & Cornillon, 2001). Former studies confirmed that migration of liquidlike lipid could be easily detected using spin-echo magnetic resonance imaging (MRI) experiments. Migration resulted in a variation of signal intensity of both chocolate and fatty fillings (Guiheneuf, Couzens, Wille, & Hall, 1997; Walter & Cornillon, 2001). Migration of liquid triacyl-glycerol was faster at 28 C than at 19 C
because of an Arrhenius relationship. The mechanism of fat migration involved both diffusion and capillary attraction phenomena (Guiheneuf et al., 1997). However, there is almost no literature on the migration properties and mechanisms of foreign liquids from fillings into chocolate samples. The present study focused on determining the migration mechanism of foreign liquid fats into chocolate at 50 C. This temperature provided a way to accelerate lipid migration into chocolate. Lauric acid was selected as a model system for studying the migration of lipids from fatty fillings into chocolate. Such choice was justified because of its price as compare to that of some triacylglycerols and quantity used for the experiments. After accelerated migration, the appearance of fat bloom on chocolate samples infiltrated by lauric acid was analyzed. Peanut butter was also used to determine the migration mechanism of fat, from a common filling used in industry, into chocolate and how such mass transfer could influence the appearance of fat bloom.
2. Experimental procedures § Journal paper No. 16246 of the Purdue University Agricultural Experiment Station. * Corresponding author. Tel.: +1-765-494-1749; fax: +1-765-4947953. E-mail address: [email protected] (P. Cornillon).
2.1. Sample preparation Dark chocolate was purchased at a local store. Its composition was 54% sugar, 44% cocoa liquor, and
0963-9969/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved. PII: S0963-9969(02)00072-8
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P. Walter, P. Cornillon / Food Research International 35 (2002) 761–767
1.5% cocoa butter. Three samples were prepared and placed in 18 mm O.D. nuclear magnetic resonance (NMR) tubes. One tube was filled with a well-tempered dark commercial chocolate (sample A), the second tube with a layer of well-tempered dark commercial chocolate (2.3 cm) under a layer of lauric acid (0.8 cm) (sample B), and the third tube with lauric acid only (sample C). In order to melt lauric acid (M.P. 42.7 C) and the chocolate, each sample was placed in a water bath at 50 C. Lauric acid migrated into the molten chocolate layer during 25 days at 50 C. Then, samples were stored at 21 C to study the appearance of fat bloom upon storage. For experiments with peanut butter, three samples were prepared. Two NMR tubes were filled with the same commercial dark chocolate. One of them was stored at 21 C (sample D) and the other placed in a water bath at 28 C (sample E). Sample F was composed of a layer of commercial dark chocolate (2.1 cm) under a layer of peanut butter (1.6 cm). All ingredients were purchased at a local store. Note that peanut butter without any pieces of peanut in it was selected (i.e. creamy type).
spectrometer as described above. The linear magnetic field gradient was generated using gradient coils made by the vendor. Images corresponded to a longitudinal cut of the samples (i.e. axes X–Y). A standard spin-echo imaging pulse sequence was used with an echo time (TE) of 8 ms and a relaxation delay of 500 ms. For the study with lauric acid, 8 scans were used for signal averaging. The dwell time was 20 ms and the 128 128 data matrix images were acquired in about 8.5 min. The field of view of the images was 5.4 cm. For the migration study with peanut butter, 16 scans were necessary in order to acquire images with good signal-to-noise ratio. The acquisition time was about 17 min. The field of view of the images was 5.4 cm. The slice selection for each image was 5 mm. The two-dimensional images were processed with an image display macro written in the IDL programming language (Research Systems Inc., Boulder, CO). Further image processing was also carried out with the freely distributed software Scion PC (http://www.scioncorp.com).
2.2. NMR relaxometry experiments
A modulated differential scanning calorimeter (MDSC 2920, TA Instruments, New Castle, NJ) was used to measure the melting points for the following samples: (1) at the end of the lauric acid migration, i.e. after 25 days at 50 C, (2) after removal of lauric acid and storage for 25 additional days at 21 C to determine the effect of the presence of lauric acid in the chocolate on lipid crystal transformation, and (3) after 28 days (17 days at 28 C and 11 days at 21 C) for the migration study with peanut butter. Samples of about 10 mg were placed in hermetic aluminum pans. An empty pan was used as reference. Two different measurements were made from each sample previously used in the NMR experiments. After stabilization for 2 min, between 5 and 5 C, the chocolate samples were heated to 60 C at 10 C min 1. For each thermogram, the melting point was determined using the analysis software provided by the vendor.
The 1H-NMR experiments were carried out at 50 C for the study with lauric acid and 28 C for the study with peanut butter using a 15 MHz Maran benchtop NMR spectrometer (Resonance Instruments Ltd., Witney, UK). The pulse lengths for all samples were 42 and 84 ms for the 90 and 180 pulses, respectively. The dwell time was 0.5 ms and the receiver gain was set to obtain a maximum signal. The other NMR parameters were set so that only liquid-like protons would contribute to the signal. The inversion-recovery pulse sequence was used to determine the spin-lattice relaxation time (T1) of the samples. The relaxation delay was fixed to 2 s. Four scans were used for signal averaging and 7–10 pulse spacing times were selected between 1 ms and 4 s to describe all the recovery curve. Spin-spin relaxation times (T2) were determined using the Carr–Purcell– Meiboom–Gill (CPMG) pulse sequence using 4096 points and an echo time of 0.2 ms for the lauric acid study, and 2048 points with an echo time of 0.3 ms for the migration study with peanut butter. Four scans were acquired with a relaxation delay of 1 s between scans. The NMR relaxation data were fitted to a single exponential decay (for T1) and a multi-exponential function (for T2) using software provided by the vendor. T1 and T2 were collected in duplicate and the error was always smaller than 4%. 2.3. MRI experiments Magnetic resonance images were acquired at 50 and 28 C, respectively for each study, using i.e. the same
2.4. DSC analysis
3. Results and discussion Fig. 1 presents MRI images for the chocolate/lauric acid system collected over migration time. Fig. 1a is an image for a commercial chocolate sample. At day 0, the image confirms the heterogeneity of such sample as indicated elsewhere (Guiheneuf et al., 1997; Miguel & Hall, 1998). This heterogeneity is due to the internal distribution of liquid triacyl-glycerols. At day 10, a phase separation can be seen due to sedimentation of denser ingredients in chocolate during storage at 50 C. Fig. 1b shows the fairly slow migration of lauric acid into the chocolate layer, mainly due to diffusion across a
P. Walter, P. Cornillon / Food Research International 35 (2002) 761–767
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Fig. 1. MRI images of chocolate and lauric acid samples over time. 1a: sample A: commercial dark chocolate; 1b: sample B: commercial dark chocolate under a layer of lauric acid; 1c: sample C: lauric acid. Temperature=50 C.
concentration gradient at approximately 0.3 mm per day. This is confirmed by the average intensity data of specific areas indicated in Fig. 2 and reported in Table 1 (Maximum intensity=256). Image intensity of the sample containing chocolate and lauric acid was lower at 21 C due to less liquid-like lipids than at 50 C. After 10 days at 50 C, the intensity of the sample without lauric acid was about six units lower for the area at the bottom of the tube (area A), 30 units lower for the intermediate layer (area C) and more than 43 units lower for the intensity of the lauric acid layer (area B). However, lauric acid was expected to remain liquid during its storage at 50 C (Fig. 1c) so that the intensity should not have varied dramatically. The decrease of intensity in that region can be noticed in the images for sample B. One of the reasons for such decrease of intensity could be migration of some chocolate components in the lauric acid phase while lauric acid transferred to chocolate. This loss of intensity became apparent after 6 days at 50 C. On images of sample B at early times, low signal intensity zones due to air gaps inside the chocolate sample were observed (indicated by the arrow—Fig. 1b, day 1). A mercury porosimetry study of dark chocolate
Fig. 2. Schematic of the different areas of sample B (commercial dark chocolate under a layer of lauric acid) during migration of lauric acid.
structure has confirmed the presence of a porous matrix composed of empty spaces or air cavities (Loisel, Lecq, Ponchel, Keller, & Ollivon, 1997). However, after 6 days, these air gaps were not observed anymore. This might be explained by the capillary migration of lauric acid or some liquid lipids from chocolate into them. This is in accordance with previous results reporting that hazelnut oil had the ability to occupy air gaps (Guiheneuf et al., 1997). Relaxometry data also indicated that lauric acid migrated into chocolate at 50 C. Indeed, the spin-lattice relaxation time for lauric acid was about 480 ms whereas for chocolate, it was around 150 ms. Sample B, containing these two components in different proportions over time, had a T1 between 175
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Table 1 Mean intensity of the different areas of the chocolate-lauric acid sample obtained during migration time. Choc=chocolate; L.Ac.=lauric acid T ( C)
21
Sample
A—Chocolate—Day 0 A—Chocolate—Day 10 B—Choc+L.Ac.—Day 0 B—Choc+L.Ac.—Day 1 B—Choc+L.Ac.—Day 2 B—Choc+L.Ac.—Day 3 B—Choc+L.Ac.—Day 4 B—Choc+L.Ac.—Day 5 B—Choc+L.Ac.—Day 6 B—Choc+L.Ac.—Day 7 B—Choc+L.Ac.—Day 8 B—Choc+L.Ac.—Day 9 B—Choc+L.Ac.—Day 10 B—Choc+L.Ac.—Day 15 B—Choc+L.Ac.—Day 25 C—L.Ac.—Day 0 C—L.Ac.—Day 10 B—Choc+L.Ac. —Day 28 B—Choc+L.Ac.—Day 33
Area A
Area B
Area C
Area D
Mean
S.D.
Mean
S.D.
Mean
S.D.
Mean
S.D.
100.3 66.4 74.2 65.2 68.3 68.8 67.9 71.7 67.1 66.2 67.2 57.5 60.5 51.5 50.0 – – 92.36 77.30
25.7 14.7 12.3 14.5 12.2 12.9 15.0 14.2 19.7 27.8 31.6 12.7 15.7 17.7 23.0 – – 21.5 26.0
– – 195.8 211.8 190.8 210.4 197.8 197.8 200.4 180.0 181.5 191.4 193.8 184.2 156.7 210.4 213.9 – –
– – 31.6 33.7 34.2 28.7 32.4 39.7 42.4 39.7 32.9 33.6 30.7 39.3 42.3 25.2 23.1 – –
– 108.3 – – – – – – – 128.0 126.5 131.4 138.7 136.5 135.8 – – 150.02 145.51
– 18.3 – – – – – – – 21.4 28.4 26.4 23.8 19.4 28.1 – – 55.2 51.0
– 177.8 – – – – – – – 228.7 216.9 227.1 221.1 222.0 199.0 – – – –
– 43.6 – – – – – – – 15.1 23.5 20.6 24.0 24.1 37.8 – – – –
ms and 230 ms. It appears that the lipids have gained chain mobility in the molten chocolate system by the increased amount of lauric acid and possible interactions between the phases. Together, lauric acid and lipids in chocolate formed a eutectic similar to that formed between hazelnut oil and cocoa butter or other fat mixtures (Bigalli, 1988; Guiheneuf et al., 1997; Talbot, 1990). Therefore, by increasing the chain mobility in cocoa butter, the liquid-like nature of the sample is enhanced and thus the intensity of the images. The diffusion coefficient (D) of the lauric acid interface in chocolate was determined by solving the diffusion equation in a semi-infinite medium where the surface concentration between the two media was changing depending on migration time (Crank, 1975). The error made in the calculation of D was related to the resolution of the images and it was estimated to be around 5%. The values found for D were comparable to that of other products of the same type. The surface concentration dependence on time was justified because the intensity of the lauric acid phase changed during the experiment due to migration of chocolate components into it. As can be seen from Figs. 1 and 3, migration started to become visible by MRI after 5–7 days. At 5 days, lauric acid migration was very limited and no attempt was made to calculate the diffusion coefficient before day 7. Beyond day 7, the distance migrated increased almost linearly with time. Unexpectedly, the diffusion coefficient did not follow the same trend during the experiment. It increased at first until day 15, but then slightly decreased as the experiment proceeded. This trend could probably be due to the internal structure of chocolate (i.e. porosity factor and heterogeneity)
and fat migration inside chocolate. After migration of lauric acid, the eutectic could have been formed, limiting diffusion by capillary migration. Lauric acid migration monitored by relaxation times is shown in Fig. 4. Both relaxation times decreased steadily over the duration of the experiment and T2 was separated in a short and a long component indicating two spin species. These data suggest that migration of lauric acid and perhaps phase separation of the triglycerides of cocoa butter are not a one-phase phenomenon. It is possible that migration could only occur if a particular internal structure of chocolate is created. At the beginning of the experiments, lauric acid migrated rapidly into the area of contact with chocolate until reaching a pseudo-steady state distribution of liquid triacylglycerol in that region (Guiheneuf et al., 1997). After 2 days at 50 C, phase separation of some liquid triacylglycerols from a sample of chocolate occurred (data not shown - this result may suggest that both mobility differences and effect of gravity play a role in such separation; however, no evidence on the mobility differences of these components is available). When under lauric acid, the most mobile liquid-like lipids from cocoa butter reached the interface between chocolate and lauric acid. Even if the mechanism of this accumulation is still not clearly understood (Guiheneuf et al., 1997), an explanation could be that, with diffusion and migration of lauric acid, the different fats would form a phase (similar to a eutectic) through strong interactions between them inducing gradients of concentration in the system and migration of new triacylglycerols towards the interface. At the end of migration, the most mobile triacylglycerols will stay at the chocolate-lauric acid interface, which is confirmed by
P. Walter, P. Cornillon / Food Research International 35 (2002) 761–767
the MRI images in which this intermediate area (e.g. area C) is higher in intensity than the bottom layer (area A). DSC measurements were carried out 50 days after the first contact between lauric acid and chocolate, and 25 days after the removal of lauric acid for storage at room temperature. The average melting point of the top layer of chocolate was about 34.2 C whereas that of the bottom layer was around 35.1 C. This indicates that area A was composed mostly of more stable fat crystals. Indeed, since most of the mobile liquid-like triacylglycerols migrated towards the interface, the bottom layer
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tended to crystallize in a more stable form after being placed at room temperature. This result is in accordance with data reported elsewhere (Schlichter-Aronhime & Garti, 1988). Visual observation of this sample stored at 21 C in the NMR tube for more than 35 days indicated fat bloom slightly at the bottom layer. Therefore, lauric acid that migrated into chocolate and increased its liquid-to-solid ratio, prevented fat bloom in the upper layer of chocolate. The glass tube may have played a role in limiting the growth of the fat crystals beyond the layer of chocolate. However, quantification of the
Fig. 3. Migrated distance and diffusion coefficient of sample B (commercial dark chocolate under a layer of lauric acid) as a function of time.
Fig. 4. Relaxation times of sample B (commercial dark chocolate under a layer of lauric acid) as a function of time.
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changes in crystal sizes during storage influenced by the presence of lauric acid was not part of the objectives of this study. Fig. 5 presents NMR images for samples E and F placed at 28 C. Only one image is presented for sample E because there was no significant modification during the time of experiment. For sample E, the relaxation times were about 92.2, 62.3, and 18.1 ms for the T1, long T2 and short T2 components, respectively; whereas for sample F, they were about 112, 162.6, and 38.7 ms for
T1, long T2, and short T2 components, respectively. Moreover, for pure peanut butter, these three parameters were 119.6, 194.8, and 59.3 ms, respectively. These values indicate that peanut butter contributed most to T1 and T2 values for the whole sample. The relaxation times did not vary significantly over migration time. However, even though T1 and T2 did not vary, the migration of peanut butter fat into chocolate occurred as indicated in Fig. 5a. After 1 day, the signal arising from the chocolate region had a higher intensity
Fig. 5. MRI images obtained for migration time of fat from fatty fillings. 5a: sample F: commercial dark chocolate with a layer of peanut butter; 5b: sample E: commercial dark chocolate. Temperature=28 C.
P. Walter, P. Cornillon / Food Research International 35 (2002) 761–767 Table 2 DSC measurements for all samples of peanut butter+chocolate. Rate: 10 C.min 1. (–) No bloom; (+) Some bloom; (++); Average bloom. N/A: Not available Sample D Sample E Sample F Peanut butter Fresh Melting Point ( C) 34.2 S.D. 0.4 Bloom –
36.9 1.7 ++
34.13 0.07 –
Old
30.6 47.4 N/A 0.5 N/A N/A
due to migration of liquid fat from peanut butter. After the second day, a low signal intensity layer (area D in Fig. 2) appeared in the sample at the interface of peanut butter and chocolate. It is not clear yet why such interface would form during migration of fat. After 16 days of migration, fat from peanut butter was present in the whole section of chocolate sample. Since fat migrated to the chocolate layer, there was less and less fat in the peanut butter and a crack appeared in this layer due to a ‘‘drying’’ phenomenon associated with migration. After 19 days, the entire peanut butter layer was separated in two parts. DSC measurements (Table 2) show that between fresh peanut butter (prior to experiment) and old peanut butter (after experiment), there is more than 16 C difference in melting point, suggesting that the old peanut butter contained less low-melting fats, which correlates well with the liquid migration out of peanut butter indicated by the NMR data. For chocolate stored at 21 C after 28 days (sample D), the melting point was about 34.2 C, which was similar to a lipid crystal of form V. Sample E, kept at 28 C during 16 days and then at 21 C a few days, presented a thin white layer on its surface. Its melting point was around 36.9 C, similar to form VI (Walter & Cornillon, 2001). Therefore, thermal history in these samples induced fat bloom as indicated elsewhere (Walter & Cornillon, 2001). However, for sample F, DSC data showed that the melting point was around 34 C. Sample F still had a glossy surface without any appearance of fat bloom. This result could indicate that cocoa butter is still in form V (lack of bloom), or that lipids from peanut butter migrated in the chocolate layer and associated with cocoa butter lipids lowering its melting point. In any case, the determination of the polymorphic state in such system may be difficult without further X-ray analysis. In addition, peanut butter oil contains the same fatty acids as cocoa butter (Stauffer, 1996), so migration of the fatty acids from one phase to the other could have been enhanced. For sample F, after the migration
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experiment, chocolate had a phase separation at the interface, which was darker than that at the bottom of the NMR tube. The top layer had a melting point slightly lower than common stable chocolate (sample D) due to the presence of peanut butter lipids that had migrated in chocolate and prevented the appearance of fat bloom. Since fat bloom is a solid transformation, the liquid lipids from peanut butter had limited this phase change by increasing the liquid-to-solid ratio of chocolate. Fat bloom appeared in sample E even though its storage temperature was higher than that for sample F. Thus, the effect of the foreign fat migration in chocolate seems to have a greater influence on the presence of fat bloom as compare to a slight increase of storage temperature. In summary, fat bloom can be prevented by a good control of temperature of chocolate (tempering), addition of foreign fat (through proper formulation), or a combination of both. The results presented here highlight the benefits of using DSC and NMR relaxometry and imaging in analyzing the effects of experimental conditions and thermal history on fat bloom appearance and mechanism.
References Bigalli, G. L. (1988). Practical aspects of the eutectic effect on confectionery fats and their mixtures. Manufacturing Confectioners, 65– 68, 79–80. Crank, J. (1975). The mathematics of diffusion. Oxford: Oxford University Press. Guiheneuf, T. M., Couzens, P. J., Wille, H. J., & Hall, L. D. (1997). Visualization of liquid triacylglycerol migration in chocolate by magnetic resonance imaging. Journal of the Science of Food and Agriculture, 75, 265–279. Hachiya, I., Koyano, T., & Sato, K. (1989). Seeding effects on solidification behavior of cocoa butter and dark chocolate. II. Physical properties of dark chocolate. Journal of the American Oil Chemists Society, 66(2), 66–75. Loisel, C., Lecq, G., Ponchel, G., Keller, G., & Ollivon, M. (1997). Fat bloom and chocolate structure studied by mercury porosimetry. Journal of Food Science, 62(4), 788–791. Miguel, M. E., & Hall, L. D. (1998). A general survey of chocolate confectionery by magnetic resonance imaging. LebensmittelWissenchaft und Technologie, 31, 93–99. Schlichter-Aronhime, J., & Garti, N. (1988). Solidification and polymorphism in cocoa butter and the blooming problems. In N. Garti, & K. Sato (Eds.), Crystallization and polymorphism of fats and fatty acids, Surfactant Science series, Vol. 31 (pp. 363–393). New York: Marcel Dekker. Stauffer, C. E. (1996). Chocolate and confectionery coatings. Fats and oils. Eagan Press Handbook Series. Talbot, G. (1990). Fat migration in biscuits and confectionery systems. Confectionery Production, 4, 265–266, 268–270, 272. Walter P., Cornillon P. (2001). Influence of thermal conditions and additives on fat bloom in chocolate. Journal of the American Oil Chemists Society (in press).
Lipid migration in two-phase chocolate systems investigated by NMR and DSC§ Peggy Walter, Paul Cornillon* Purdue University-Department of Food Science, Whistler Center for Carbohydrate Research, 1160 Food Science Building-West Lafayette, IN 47907-1160, USA Received 30 September 2000; accepted 6 December 2001
Abstract The migration of lipids in two-phase chocolate systems (i.e. lauric acid+chocolate and peanut butter+chocolate) was analyzed by magnetic resonance and differential scanning calorimetry. Kinetics of fat migration was evaluated and the diffusion coefficient of lauric acid in chocolate was found to be dependent on migration time. This may be due to the capillary nature of fat migration in chocolate. Fat bloom characteristics were determined and related to the thermal history of samples and the presence of fat in the chocolate layer. Both lauric acid and peanut butter increased the liquid-to-solid ratio of chocolate and helped prevent fat bloom. # 2002 Elsevier Science Ltd. All rights reserved. Keywords: Chocolate; DSC; Fat bloom; Fat migration; MRI; Relaxation times
1. Introduction Fat bloom is an important issue for chocolate confectioners. Without proper tempering, it can appear on chocolate samples stored at room temperature after different thermal histories. This physical defect occurred in less than 2 days after storage at 21 C with no tempering prior to cooling (Walter & Cornillon, 2001). The lack of gloss and the presence of white areas that characterize fat bloom often occur for confectionery products containing a layer of chocolate and one or more layers of fat fillings. To prevent fat bloom, additives like triglycerides (e.g. B.O.B. or 1–3 dibehenoyl, 2-oleoylglycerol) could be used (Hachiya, Koyano, & Sato, 1989; Walter & Cornillon, 2001). Former studies confirmed that migration of liquidlike lipid could be easily detected using spin-echo magnetic resonance imaging (MRI) experiments. Migration resulted in a variation of signal intensity of both chocolate and fatty fillings (Guiheneuf, Couzens, Wille, & Hall, 1997; Walter & Cornillon, 2001). Migration of liquid triacyl-glycerol was faster at 28 C than at 19 C
because of an Arrhenius relationship. The mechanism of fat migration involved both diffusion and capillary attraction phenomena (Guiheneuf et al., 1997). However, there is almost no literature on the migration properties and mechanisms of foreign liquids from fillings into chocolate samples. The present study focused on determining the migration mechanism of foreign liquid fats into chocolate at 50 C. This temperature provided a way to accelerate lipid migration into chocolate. Lauric acid was selected as a model system for studying the migration of lipids from fatty fillings into chocolate. Such choice was justified because of its price as compare to that of some triacylglycerols and quantity used for the experiments. After accelerated migration, the appearance of fat bloom on chocolate samples infiltrated by lauric acid was analyzed. Peanut butter was also used to determine the migration mechanism of fat, from a common filling used in industry, into chocolate and how such mass transfer could influence the appearance of fat bloom.
2. Experimental procedures § Journal paper No. 16246 of the Purdue University Agricultural Experiment Station. * Corresponding author. Tel.: +1-765-494-1749; fax: +1-765-4947953. E-mail address: [email protected] (P. Cornillon).
2.1. Sample preparation Dark chocolate was purchased at a local store. Its composition was 54% sugar, 44% cocoa liquor, and
0963-9969/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved. PII: S0963-9969(02)00072-8
762
P. Walter, P. Cornillon / Food Research International 35 (2002) 761–767
1.5% cocoa butter. Three samples were prepared and placed in 18 mm O.D. nuclear magnetic resonance (NMR) tubes. One tube was filled with a well-tempered dark commercial chocolate (sample A), the second tube with a layer of well-tempered dark commercial chocolate (2.3 cm) under a layer of lauric acid (0.8 cm) (sample B), and the third tube with lauric acid only (sample C). In order to melt lauric acid (M.P. 42.7 C) and the chocolate, each sample was placed in a water bath at 50 C. Lauric acid migrated into the molten chocolate layer during 25 days at 50 C. Then, samples were stored at 21 C to study the appearance of fat bloom upon storage. For experiments with peanut butter, three samples were prepared. Two NMR tubes were filled with the same commercial dark chocolate. One of them was stored at 21 C (sample D) and the other placed in a water bath at 28 C (sample E). Sample F was composed of a layer of commercial dark chocolate (2.1 cm) under a layer of peanut butter (1.6 cm). All ingredients were purchased at a local store. Note that peanut butter without any pieces of peanut in it was selected (i.e. creamy type).
spectrometer as described above. The linear magnetic field gradient was generated using gradient coils made by the vendor. Images corresponded to a longitudinal cut of the samples (i.e. axes X–Y). A standard spin-echo imaging pulse sequence was used with an echo time (TE) of 8 ms and a relaxation delay of 500 ms. For the study with lauric acid, 8 scans were used for signal averaging. The dwell time was 20 ms and the 128 128 data matrix images were acquired in about 8.5 min. The field of view of the images was 5.4 cm. For the migration study with peanut butter, 16 scans were necessary in order to acquire images with good signal-to-noise ratio. The acquisition time was about 17 min. The field of view of the images was 5.4 cm. The slice selection for each image was 5 mm. The two-dimensional images were processed with an image display macro written in the IDL programming language (Research Systems Inc., Boulder, CO). Further image processing was also carried out with the freely distributed software Scion PC (http://www.scioncorp.com).
2.2. NMR relaxometry experiments
A modulated differential scanning calorimeter (MDSC 2920, TA Instruments, New Castle, NJ) was used to measure the melting points for the following samples: (1) at the end of the lauric acid migration, i.e. after 25 days at 50 C, (2) after removal of lauric acid and storage for 25 additional days at 21 C to determine the effect of the presence of lauric acid in the chocolate on lipid crystal transformation, and (3) after 28 days (17 days at 28 C and 11 days at 21 C) for the migration study with peanut butter. Samples of about 10 mg were placed in hermetic aluminum pans. An empty pan was used as reference. Two different measurements were made from each sample previously used in the NMR experiments. After stabilization for 2 min, between 5 and 5 C, the chocolate samples were heated to 60 C at 10 C min 1. For each thermogram, the melting point was determined using the analysis software provided by the vendor.
The 1H-NMR experiments were carried out at 50 C for the study with lauric acid and 28 C for the study with peanut butter using a 15 MHz Maran benchtop NMR spectrometer (Resonance Instruments Ltd., Witney, UK). The pulse lengths for all samples were 42 and 84 ms for the 90 and 180 pulses, respectively. The dwell time was 0.5 ms and the receiver gain was set to obtain a maximum signal. The other NMR parameters were set so that only liquid-like protons would contribute to the signal. The inversion-recovery pulse sequence was used to determine the spin-lattice relaxation time (T1) of the samples. The relaxation delay was fixed to 2 s. Four scans were used for signal averaging and 7–10 pulse spacing times were selected between 1 ms and 4 s to describe all the recovery curve. Spin-spin relaxation times (T2) were determined using the Carr–Purcell– Meiboom–Gill (CPMG) pulse sequence using 4096 points and an echo time of 0.2 ms for the lauric acid study, and 2048 points with an echo time of 0.3 ms for the migration study with peanut butter. Four scans were acquired with a relaxation delay of 1 s between scans. The NMR relaxation data were fitted to a single exponential decay (for T1) and a multi-exponential function (for T2) using software provided by the vendor. T1 and T2 were collected in duplicate and the error was always smaller than 4%. 2.3. MRI experiments Magnetic resonance images were acquired at 50 and 28 C, respectively for each study, using i.e. the same
2.4. DSC analysis
3. Results and discussion Fig. 1 presents MRI images for the chocolate/lauric acid system collected over migration time. Fig. 1a is an image for a commercial chocolate sample. At day 0, the image confirms the heterogeneity of such sample as indicated elsewhere (Guiheneuf et al., 1997; Miguel & Hall, 1998). This heterogeneity is due to the internal distribution of liquid triacyl-glycerols. At day 10, a phase separation can be seen due to sedimentation of denser ingredients in chocolate during storage at 50 C. Fig. 1b shows the fairly slow migration of lauric acid into the chocolate layer, mainly due to diffusion across a
P. Walter, P. Cornillon / Food Research International 35 (2002) 761–767
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Fig. 1. MRI images of chocolate and lauric acid samples over time. 1a: sample A: commercial dark chocolate; 1b: sample B: commercial dark chocolate under a layer of lauric acid; 1c: sample C: lauric acid. Temperature=50 C.
concentration gradient at approximately 0.3 mm per day. This is confirmed by the average intensity data of specific areas indicated in Fig. 2 and reported in Table 1 (Maximum intensity=256). Image intensity of the sample containing chocolate and lauric acid was lower at 21 C due to less liquid-like lipids than at 50 C. After 10 days at 50 C, the intensity of the sample without lauric acid was about six units lower for the area at the bottom of the tube (area A), 30 units lower for the intermediate layer (area C) and more than 43 units lower for the intensity of the lauric acid layer (area B). However, lauric acid was expected to remain liquid during its storage at 50 C (Fig. 1c) so that the intensity should not have varied dramatically. The decrease of intensity in that region can be noticed in the images for sample B. One of the reasons for such decrease of intensity could be migration of some chocolate components in the lauric acid phase while lauric acid transferred to chocolate. This loss of intensity became apparent after 6 days at 50 C. On images of sample B at early times, low signal intensity zones due to air gaps inside the chocolate sample were observed (indicated by the arrow—Fig. 1b, day 1). A mercury porosimetry study of dark chocolate
Fig. 2. Schematic of the different areas of sample B (commercial dark chocolate under a layer of lauric acid) during migration of lauric acid.
structure has confirmed the presence of a porous matrix composed of empty spaces or air cavities (Loisel, Lecq, Ponchel, Keller, & Ollivon, 1997). However, after 6 days, these air gaps were not observed anymore. This might be explained by the capillary migration of lauric acid or some liquid lipids from chocolate into them. This is in accordance with previous results reporting that hazelnut oil had the ability to occupy air gaps (Guiheneuf et al., 1997). Relaxometry data also indicated that lauric acid migrated into chocolate at 50 C. Indeed, the spin-lattice relaxation time for lauric acid was about 480 ms whereas for chocolate, it was around 150 ms. Sample B, containing these two components in different proportions over time, had a T1 between 175
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Table 1 Mean intensity of the different areas of the chocolate-lauric acid sample obtained during migration time. Choc=chocolate; L.Ac.=lauric acid T ( C)
21
Sample
A—Chocolate—Day 0 A—Chocolate—Day 10 B—Choc+L.Ac.—Day 0 B—Choc+L.Ac.—Day 1 B—Choc+L.Ac.—Day 2 B—Choc+L.Ac.—Day 3 B—Choc+L.Ac.—Day 4 B—Choc+L.Ac.—Day 5 B—Choc+L.Ac.—Day 6 B—Choc+L.Ac.—Day 7 B—Choc+L.Ac.—Day 8 B—Choc+L.Ac.—Day 9 B—Choc+L.Ac.—Day 10 B—Choc+L.Ac.—Day 15 B—Choc+L.Ac.—Day 25 C—L.Ac.—Day 0 C—L.Ac.—Day 10 B—Choc+L.Ac. —Day 28 B—Choc+L.Ac.—Day 33
Area A
Area B
Area C
Area D
Mean
S.D.
Mean
S.D.
Mean
S.D.
Mean
S.D.
100.3 66.4 74.2 65.2 68.3 68.8 67.9 71.7 67.1 66.2 67.2 57.5 60.5 51.5 50.0 – – 92.36 77.30
25.7 14.7 12.3 14.5 12.2 12.9 15.0 14.2 19.7 27.8 31.6 12.7 15.7 17.7 23.0 – – 21.5 26.0
– – 195.8 211.8 190.8 210.4 197.8 197.8 200.4 180.0 181.5 191.4 193.8 184.2 156.7 210.4 213.9 – –
– – 31.6 33.7 34.2 28.7 32.4 39.7 42.4 39.7 32.9 33.6 30.7 39.3 42.3 25.2 23.1 – –
– 108.3 – – – – – – – 128.0 126.5 131.4 138.7 136.5 135.8 – – 150.02 145.51
– 18.3 – – – – – – – 21.4 28.4 26.4 23.8 19.4 28.1 – – 55.2 51.0
– 177.8 – – – – – – – 228.7 216.9 227.1 221.1 222.0 199.0 – – – –
– 43.6 – – – – – – – 15.1 23.5 20.6 24.0 24.1 37.8 – – – –
ms and 230 ms. It appears that the lipids have gained chain mobility in the molten chocolate system by the increased amount of lauric acid and possible interactions between the phases. Together, lauric acid and lipids in chocolate formed a eutectic similar to that formed between hazelnut oil and cocoa butter or other fat mixtures (Bigalli, 1988; Guiheneuf et al., 1997; Talbot, 1990). Therefore, by increasing the chain mobility in cocoa butter, the liquid-like nature of the sample is enhanced and thus the intensity of the images. The diffusion coefficient (D) of the lauric acid interface in chocolate was determined by solving the diffusion equation in a semi-infinite medium where the surface concentration between the two media was changing depending on migration time (Crank, 1975). The error made in the calculation of D was related to the resolution of the images and it was estimated to be around 5%. The values found for D were comparable to that of other products of the same type. The surface concentration dependence on time was justified because the intensity of the lauric acid phase changed during the experiment due to migration of chocolate components into it. As can be seen from Figs. 1 and 3, migration started to become visible by MRI after 5–7 days. At 5 days, lauric acid migration was very limited and no attempt was made to calculate the diffusion coefficient before day 7. Beyond day 7, the distance migrated increased almost linearly with time. Unexpectedly, the diffusion coefficient did not follow the same trend during the experiment. It increased at first until day 15, but then slightly decreased as the experiment proceeded. This trend could probably be due to the internal structure of chocolate (i.e. porosity factor and heterogeneity)
and fat migration inside chocolate. After migration of lauric acid, the eutectic could have been formed, limiting diffusion by capillary migration. Lauric acid migration monitored by relaxation times is shown in Fig. 4. Both relaxation times decreased steadily over the duration of the experiment and T2 was separated in a short and a long component indicating two spin species. These data suggest that migration of lauric acid and perhaps phase separation of the triglycerides of cocoa butter are not a one-phase phenomenon. It is possible that migration could only occur if a particular internal structure of chocolate is created. At the beginning of the experiments, lauric acid migrated rapidly into the area of contact with chocolate until reaching a pseudo-steady state distribution of liquid triacylglycerol in that region (Guiheneuf et al., 1997). After 2 days at 50 C, phase separation of some liquid triacylglycerols from a sample of chocolate occurred (data not shown - this result may suggest that both mobility differences and effect of gravity play a role in such separation; however, no evidence on the mobility differences of these components is available). When under lauric acid, the most mobile liquid-like lipids from cocoa butter reached the interface between chocolate and lauric acid. Even if the mechanism of this accumulation is still not clearly understood (Guiheneuf et al., 1997), an explanation could be that, with diffusion and migration of lauric acid, the different fats would form a phase (similar to a eutectic) through strong interactions between them inducing gradients of concentration in the system and migration of new triacylglycerols towards the interface. At the end of migration, the most mobile triacylglycerols will stay at the chocolate-lauric acid interface, which is confirmed by
P. Walter, P. Cornillon / Food Research International 35 (2002) 761–767
the MRI images in which this intermediate area (e.g. area C) is higher in intensity than the bottom layer (area A). DSC measurements were carried out 50 days after the first contact between lauric acid and chocolate, and 25 days after the removal of lauric acid for storage at room temperature. The average melting point of the top layer of chocolate was about 34.2 C whereas that of the bottom layer was around 35.1 C. This indicates that area A was composed mostly of more stable fat crystals. Indeed, since most of the mobile liquid-like triacylglycerols migrated towards the interface, the bottom layer
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tended to crystallize in a more stable form after being placed at room temperature. This result is in accordance with data reported elsewhere (Schlichter-Aronhime & Garti, 1988). Visual observation of this sample stored at 21 C in the NMR tube for more than 35 days indicated fat bloom slightly at the bottom layer. Therefore, lauric acid that migrated into chocolate and increased its liquid-to-solid ratio, prevented fat bloom in the upper layer of chocolate. The glass tube may have played a role in limiting the growth of the fat crystals beyond the layer of chocolate. However, quantification of the
Fig. 3. Migrated distance and diffusion coefficient of sample B (commercial dark chocolate under a layer of lauric acid) as a function of time.
Fig. 4. Relaxation times of sample B (commercial dark chocolate under a layer of lauric acid) as a function of time.
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changes in crystal sizes during storage influenced by the presence of lauric acid was not part of the objectives of this study. Fig. 5 presents NMR images for samples E and F placed at 28 C. Only one image is presented for sample E because there was no significant modification during the time of experiment. For sample E, the relaxation times were about 92.2, 62.3, and 18.1 ms for the T1, long T2 and short T2 components, respectively; whereas for sample F, they were about 112, 162.6, and 38.7 ms for
T1, long T2, and short T2 components, respectively. Moreover, for pure peanut butter, these three parameters were 119.6, 194.8, and 59.3 ms, respectively. These values indicate that peanut butter contributed most to T1 and T2 values for the whole sample. The relaxation times did not vary significantly over migration time. However, even though T1 and T2 did not vary, the migration of peanut butter fat into chocolate occurred as indicated in Fig. 5a. After 1 day, the signal arising from the chocolate region had a higher intensity
Fig. 5. MRI images obtained for migration time of fat from fatty fillings. 5a: sample F: commercial dark chocolate with a layer of peanut butter; 5b: sample E: commercial dark chocolate. Temperature=28 C.
P. Walter, P. Cornillon / Food Research International 35 (2002) 761–767 Table 2 DSC measurements for all samples of peanut butter+chocolate. Rate: 10 C.min 1. (–) No bloom; (+) Some bloom; (++); Average bloom. N/A: Not available Sample D Sample E Sample F Peanut butter Fresh Melting Point ( C) 34.2 S.D. 0.4 Bloom –
36.9 1.7 ++
34.13 0.07 –
Old
30.6 47.4 N/A 0.5 N/A N/A
due to migration of liquid fat from peanut butter. After the second day, a low signal intensity layer (area D in Fig. 2) appeared in the sample at the interface of peanut butter and chocolate. It is not clear yet why such interface would form during migration of fat. After 16 days of migration, fat from peanut butter was present in the whole section of chocolate sample. Since fat migrated to the chocolate layer, there was less and less fat in the peanut butter and a crack appeared in this layer due to a ‘‘drying’’ phenomenon associated with migration. After 19 days, the entire peanut butter layer was separated in two parts. DSC measurements (Table 2) show that between fresh peanut butter (prior to experiment) and old peanut butter (after experiment), there is more than 16 C difference in melting point, suggesting that the old peanut butter contained less low-melting fats, which correlates well with the liquid migration out of peanut butter indicated by the NMR data. For chocolate stored at 21 C after 28 days (sample D), the melting point was about 34.2 C, which was similar to a lipid crystal of form V. Sample E, kept at 28 C during 16 days and then at 21 C a few days, presented a thin white layer on its surface. Its melting point was around 36.9 C, similar to form VI (Walter & Cornillon, 2001). Therefore, thermal history in these samples induced fat bloom as indicated elsewhere (Walter & Cornillon, 2001). However, for sample F, DSC data showed that the melting point was around 34 C. Sample F still had a glossy surface without any appearance of fat bloom. This result could indicate that cocoa butter is still in form V (lack of bloom), or that lipids from peanut butter migrated in the chocolate layer and associated with cocoa butter lipids lowering its melting point. In any case, the determination of the polymorphic state in such system may be difficult without further X-ray analysis. In addition, peanut butter oil contains the same fatty acids as cocoa butter (Stauffer, 1996), so migration of the fatty acids from one phase to the other could have been enhanced. For sample F, after the migration
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experiment, chocolate had a phase separation at the interface, which was darker than that at the bottom of the NMR tube. The top layer had a melting point slightly lower than common stable chocolate (sample D) due to the presence of peanut butter lipids that had migrated in chocolate and prevented the appearance of fat bloom. Since fat bloom is a solid transformation, the liquid lipids from peanut butter had limited this phase change by increasing the liquid-to-solid ratio of chocolate. Fat bloom appeared in sample E even though its storage temperature was higher than that for sample F. Thus, the effect of the foreign fat migration in chocolate seems to have a greater influence on the presence of fat bloom as compare to a slight increase of storage temperature. In summary, fat bloom can be prevented by a good control of temperature of chocolate (tempering), addition of foreign fat (through proper formulation), or a combination of both. The results presented here highlight the benefits of using DSC and NMR relaxometry and imaging in analyzing the effects of experimental conditions and thermal history on fat bloom appearance and mechanism.
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