Abstract
In this study, white pan breads part-baked 10, 15, and 20 min with and without added calcium propionate were stored at 20°C (room temperature) for 3, 5, and 7 days and at 4°C (refrigerator temperature) for 7, 14, and 21 days. After storage, the baking time of part-baked breads was completed to the baking time of control breads (25 min). Total aerobic mesophylic bacteria (TAMB), coliform bacteria, yeast and mold, and Bacillus spore counts of breads were determined before and after the second baking. While TAMB, yeast and mold counts were 8-log CFU/g in dough, it was measured as 6 and 2-log CFU/g before and after the rebaking process, respectively. Microorganism counts of the part-baked breads without Ca-propionate stored at room temperature increased in significant amounts. However, the second baking process after storage contributed to the re-freshness of breads and decreased the microorganism counts. The levels of water activity (aw) for breads with and without ca-propionate stored at different temperatures and time profiles approximately ranged from 0.92 to 0.89 after the rebaking process and did not significantly affect the microorganism counts. However, addition of calcium propionate in the bread formulation significantly decreased TAMB, coliform bacteria, Bacillus spore, and yeast and mold counts, depending on decrease of pH levels. It was found that the microbiological quality of the rebaking bread with Ca-propionate after part-baking for 10 and 15 mins and storage at both room and refrigerator temperature was much higher than that of the other.
INTRODUCTION
Bread is the main staple food meeting nutritional needs of humans in the world. Physico-chemical changes (staling and strength), and microbial spoilages such as rope and mold growth, are major factors limiting the shelf life of bread. The texture and aroma of bread is quickly lost. In considering the importance of bread in terms of nutrition and over consumption, this limited shelf life causes great economic losses in the world. The shelf life of bread has been extended by different product formulations, and the various process conditions and packaging techniques due to losing to its freshness quickly (desirable texture and aroma) and exposing to mould [Citation1–4].
The shelf life of bread is mainly influenced by water content and its distribution in bread that influence the softness of the crumb, crispness of the crust, and the quality of bread. As generally known, water activity (aw) and pH are also important factors that effect the microbial quality of foods. Water activity is defined as the ratio of the vapor pressure of water over a substrate compared to that over pure water at the same temperature and pressure. The lower levels of water activity for microbial growth are approximately 0.91 for bacteria, 0.88 for yeasts, 0.80 for fungi, 0.65 for xerophylic fungi, and 0.60 for osmophylic yeasts.[Citation5,Citation6,Citation7] Partial baking is a gradual baking process. This process has a great market potential, since bread is consumed as fresh and crust with a simple last baking. The disadvantage of the gradual baking process method is caused by the difficulties in the conservation of bread quality for a long time.[Citation8] The part-baked breads were marketed in the USA for 50 years with the slogan “more fry and ready to service.” The part-baked breads must be adequately baked in a traditional oven until the bread structure forms. The color of the bread crumb must be light and contain sufficient moisture for the formation of desirable characteristics when it is re-baked. In addition, in order to obtain a product that is ready to serve, a second baking process must be done.[Citation9,Citation10] Some fungi such as Rhizopus nigricans, Aspergillus niger, and Penicillium expansum, and bacteria such as Bacillus subtilis and Bacillus pumilus (rope) cause microbial spoilage in bread. Therefore, various preservative agents are added to the bread formulation to prevent fungal spoilage.[Citation11,Citation12] Leuschner et al.[Citation13] studied the effects on Bacillus spores of the rebaking process in part-baked soda breads and determined D values of Bacillus spores. Rope formation was observed at the end of 2 d at room temperature. D values of Bacillus subtilis, Bacillus pumilus, and Bacillus licheniformis spores isolated breads were also 14, 10, and 56 min at 100°C, respectively. In addition, they reported that Bacillus spores did not grow at 4°C and pH 10, and Bacillus licheniformis spores were active with the re-baking process. In another study, it was reported that the numbers of Bacillus spore were 1.8 CFU/g in wheat, 3.4 CFU/g in wheat flour, 12.4 CFU/g in wheat scurf, 14.9 CFU/g in whole wheat flour, 7.0 CFU/g in rye, 2.2 CFU/g in rye flour, 2.9 CFU/g in rye scurf, 7.3 CFU/g in whole rye flour, and 106 CFU/g in white and whole flour breads after storage of 2 d at 25-30°C. Of the genus Bacillus isolated from breads, 70% was Bacillus subtilis, 24 % Bacillus licheniformis, 2% Bacillus pumilus, and 2% Bacillus cereus. Bacillus subtilis strains were also found in higher levels than the others, because of its resistance to heat.[Citation14] The aim of this research was to evaluate the shelf life and the microbiological and consuming qualities of white pan breads re baked after part-baking and storage at various time-temperature profiles. Samples were analyzed in duplicate (n: 2), and the means of the results determined.
MATERIALS AND METHODS
Wheat flour (13.87% water, 12.83% protein, 0.53% ash, 30.7% gluten, 59.5% water absorption), fresh yeast, and salt were obtained from the local grocery stores. Calcium propionate was purchased from Fluka Inc. Bread was baked in a conventional oven according to the standard pup-loaf procedure, AACC method 10/09[Citation15] with some modifications. White bread was made according to the following dough formulation: 100 parts (weight basis) wheat flour; water, 59.5; salt, 1.5; yeast, 3; and calcium propionate, 0.2. All ingredients were poured into a mixer (Stephan UM-5) and mixed for 2 min using low speed (1500 rpm). The production diagram of part-baked white pan bread is presented in . For production of part-baked white bread (baking phase I), baking times of 10, 15, and 20 mins in a conventional oven at 230°C were used. After the baking process, breads were cooled for 1 hr at room temperature and then packaged in 20 × 30 cm double-folded nylon pouches and stored for 3, 5, and 7 days at room temperature and for 7, 14, and 21 days at refrigerator temperature. After storage, the part-baked breads were re baked in the same oven (230°C). The baking periods of the part-baked breads were completed to the baking time (25 min) of the control breads in the re baking process (baking phase II).
Figure 1 The production diagram of part-baked and rebaked white pan bread
Physical and Chemical Analysis
Moisture content of flour was determined by using an air circulation-drying chamber for 2.5 hr at 135°C.[Citation16] The amount of protein was determined with the Kjeldahl method as described by Elgün et al.[Citation16] Ash content was carried out at 910°C ±25.[Citation14] Wet gluten was determined using Glutomatic 2000.[Citation16] The pH was measured using an ATI-ORION 420A model pH-meter.[Citation16] Water activity (aw) of the bread crumbs was determined using the electrohigrometric method proposed by Münzing.[Citation17] Values for investigated parameters were means of duplicate.
Microbiological Analysis
The sample homogenizations and dilutions were first prepared for microbiological analysis. For this step, 25 g of sample was weighed in sterile stomacher bag and 225 ml steril physiological water (8.5 %NaCl) was added and then homogenized using a Stomacher 400 (Seward Medical London Se1 1PP UK). Thus, the first dilution of 10−1 was obtained. The other dilutions were prepared from the dilution of 10−1 to 10−6 and microbiological analyses were performed as follows.
Total aerobic mesophylic bacteria (TAMB) count
Total aerobic mesophylic bacteria count was determined as colony forming units (CFU/g) bread. Suitable dilutions of 0.1 ml (10−4 and 10−5) were spread on about 15 ml of plate count agar (PCA) (Merck). Inoculated petri plates were incubated at 30°C for 3 days. After incubation, the colonies forming in 10-300 CFU/g were enumerated using a colony counter (Dark field Quebect Model 3326, AO-American Optical).[Citation18]
Coliform bacteria count
The coliform bacteria count was determined as colony forming units (CFU/g) bread. Suitable dilutions of 0.1 ml (10−1 and 10−3) were spread on about 15 ml of violet bile agar (VRB) (Merck), and the petri plates were then incubated at 37°C for 1 day (18).
Yeast and Mold Count
The yeast and mold count was determined as colony forming units (CFU/g) bread. Suitable dilutions of 0.1 ml (10−2 and 10−4) spread on about 15 ml of potato dextrose agar (PDA) (Merck) in petri plates and then incubated at 20°C for 5 days (18).
Bacilusi spore count
Bacilli were determined as colony forming units (CFU/g) bread. The bread homogenizates were first transferred into sterile test tubes and incubated at 80°C for 10 min and then cooled to 45-50°C, and then spread on about 15 ml of plate count agar in petri plates. The petri plates were incubated at 30°C for 3 days (14). The experiments were performed twice.
RESULTS AND DISCUSSION
Results of Microbiological, Physical, and Chemical Analyses for Wheat Flour
Results of physical, chemical and microbiological analyses for the wheat flour are presented in . Results of microbiological analysis for the dough used in the production of wheat pan bread are presented in . As shown in , the numbers of TAMB, Bacillus spore and yeast and mold counts were found higher in the dough with added antimicrobial (calcium propionate) than in the dough without Ca-propionate. However, the coliform bacteria count was higher in the dough without added antimicrobial than bread samples. In a similar research, TAMB and yeast-mold counts were found more often in the dough samples than bread samples. Addition of antimicrobial significantly affected the numbers of microorganism.[Citation19].
Table 1 Results of chemical and microbiological analyses for the wheat flour.
Table 2 Results of microbiological analysis for the dough made from wheat flour.
Part-Baked White Pan Breads without Storage
Results of microbiological analyses, aw and pH for the part-baked breads without storage are presented in . The addition of antimicrobials and baking time did not affect the numbers of microorganisms and water activity levels of the breads. The coliform bacteria count was lower than log-2.00 CFU/g bread. The numbers of TAMB, yeast, and mold in dough samples also decreased from 8 to 6-log CFU/g bread with a decrease by 2-log CFU/g dough (). The microorganism counts increased significantly during storage of the breads without added antimicrobials at room temperature. However, the second baking process after storage proved the refreshness of breads and caused a decrease in the microorganism counts.
Table 3 Results of microbiological analysis for the part-baked white pan breads baked in different times.
The water activity of part-baked breads was about 0.92., which is appropriate for growing microorganism groups. As a matter of fact, the growth of all species is quite similar and, growth is faster at 0.90 a w and slower at the reduced water activity.[Citation20] The pH values of part-baking breads decreased with the addition of Ca-propionate. However, part-baking time did not significantly affect it. The results of analyses of the part baked white pan breads stored at various time-temperature profiles follow.
The breads stored at room temperature (20628C)
The numbers of TAMB, coliform bacteria, yeast, mold, and Bacillus spores for the part-baked breads stored at room temperature (20±2°C) before the second baking are shown in . The numbers of TAMB, yeast, mold, and Bacillus spores before the second baking increased as the storage period increased. The use of antimicrobial caused a decrease in the numbers of TAMB, coliform bacteria, yeast, mold, and Bacillus spores in the breads. The TAMB counts of the breads without added antimicrobial were about 8-log CFU/g at the end of storage of 7 days; however, in the breads without Ca-propinate, they were 2-log CFU/g with a decrease by 6-log CFU/g. In addition, a reduction of 5 and 3-log CFU/g in the numbers of Bacillus spores and yeast and mold counts of the breads were observed. The use of antimicrobial in the bread formulation significantly affected coliform bacteria counts. The coliform bacteria count was found in < 2,00 log-CFU/g of the breads after storage following the rebaking process.
Table 4 Results of microbiological analysis for the part-baked breads stored at room temperature (20±2°C).
Use of calcium propionate as an antimicrobial significantly affected the numbers of microorganisms in the breads. As a matter of fact, the Ca-propionate added to the bread formulation converts to calcium and propionate. Propionate reacts with the water and then forms propionic acid. Production of propionic acid is also stimulated with a low pH. Furthermore, Ca-propionate causes a the decrease of the intracellular pH and inhibits enzymes and microorganisms.[Citation21,Citation22] The numbers of TAMB, coliform, and Bacillus spores decreased with an increase of baking time in all the storage periods.
The numbers of TAMB, coliform bacteria, yeast and mould, Bacillus spores, water activity (aw) and pH values of the breads re baked after storage at room temperature following the part baking process are presented in . As shown in , the total baking time was completed to 25 min with the second baking process after storage. The number of microorganisms of the breads re baked was less than that of the breads without re baking. Because the second baking provided re freshness of the breads, the numbers of microorganisms increased by the part-baking process decreased significantly. The second baking process particularly caused a decrease in the yeast and mold counts of the breads.
When yeast and mold counts of the part-baked breads without added antimicrobial before the second baking were 2.00-6.64-log CFU/g, that of all baked breads without added antimicrobial after the second baking was 2.00-2.92-log CFU/g. Viljoen et al.[Citation19] also reported that TAMB counts were greater than yeast and mold counts for all part-baking and storage times. The addition of antimicrobial caused a decrease in the numbers of TAMB, coliform, Bacillus spore, and yeast and mold. When TAMB and Bacillus spore counts were about 7-log CFU/g on the 7th day of storage, a decrease of 5-log CFU/g in the breads with added antimicrobial was observed.
An increase in the part-baking times of the breads without added antimicrobial resulted in very little increase in TAMB and Bacillus spores. The breads baking with the time of 25 min after storage such as the control group breads were re-baked times for a short time (the second baking process). It has been reported that Bacillus spores could be resistant to temperatures of 97-101°C (14). Bacillus spores might survive, because of all the parts of bread did not reach to these temperatures. In order to prevent the rope formation in breads made with the part-baking method, dough bases must be used containing low bacterial load, and the pH of the dough must be reduced with antimicrobial additions. In addition, breads must be quickly cooled and kept at low temperatures. Furthermore, the oven atmosphere must be adequately sanitary.[Citation23,Citation24]
The second baking process significantly affected coliform bacteria counts. When the coliform bacteria count was 5-log CFU/g at the end of storage before re-baking (), this count had stayed below the detectable limits, except for the white pan breads without added antimicrobial baked 10 min after storage of 7 days. The part-baking time and storage period slightly changed the pH and aw of the breads re baked after storage at room temperature following the part-baking process. Water activity generally decreased as the storage period increased. This situation could be attributed to water lose in bread during the storage period. While the pH decreased with the addition of Ca-propionate, aw did not significantly change with the addition of an antimicrobial additive. It was found that the levels of pH for breads with and without Ca-propionate were approximately 6.3 and 5.8, respectively. This reduction of pH may also cause a decrease in the microorganism counts of breads. In general, the breads re baked after storage at room temperature, following the part-baking process, had aw values in the range 0.89-0.92 (). The pH, water activity, and temperature are the most important parameters that control the microbial growth responsible for the deterioration of foods [Citation20,Citation25].
The breads stored at refrigerator temperature (4628C)
The numbers of TAMB, coliform bacteria, Bacillus spore, and yeast and mold of the part-baked breads stored at refrigerator temperature (4±2°C) before the re-baking are presented in . As shown in this table, the microorganism counts of the part-baked breads stored at refrigerator temperature were less than those of the part-baked breads stored at room temperature, although the storage period was quite long.
The use of antimicrobial additives caused a decrease in the numbers of TAMB, coliform bacteria, and Bacillus spore in the part-baked breads stored at room temperature. The number of TAMB, coliform bacteria, and Bacillus spores decreased with an increase of the part-baking time. The number of TAMB, coliform bacteria, yeast and mold, Bacillus spore, aw, and pH values of the breads re baked after storage at refrigerator temperature following the part-baking process are presented in . As shown in the table, the storage temperature of the part-baked breads that were re baked significantly affected the microorganism counts in the final product. The microorganism counts of the breads stored at refrigerator temperature after the second baking were less than that of the breads stored at room temperature. Coliform bacteria counts were determined as < 2-log CFU/g in the different storage periods and temperatures. The numbers of TAMB, coliform bacteria and, yeast and mold were generally about 2-log CFU/g.
The effect of an antimicrobial additive, part-baking time, and storage period on water activity levels and pH values () of the breads re baked after storage at refrigerator temperature following the part-baking process were similar to that of the breads at room temperature (). As seen in , while aw levels ranged from 0.89 to 0.92, pH values of breads with and without Ca-propionate were about 6.3 and 5.9, respectively.
Table 5 Results of microbiological analysis for the part-baked breads stored at refrigerator temperature (4±2°C).
CONCLUSION
The results of this study indicated that the part-baking method was a suitable method for white pan bread production. The microbiological quality was found to be more useful in part-baking breads stored at refrigerator temperature than at room temperature. We may conclude that the use of an antimicrobial agent in part-baked breads is very important and, if not, the part-baked breads must be stored at refrigerator temperature (4±2°C). Antimicrobial additives limited the microbial growth due to a decrease of pH rather than aw.
Related Research Data
REFERENCES
- Knightly , W.H. 1977 . The staling of bread. A review . Baker's Dig. , 51 ( 5 ) : 52 – 56 .
- Knorr , D. and Tomlis , R.I. 1985 . Effect of carbon dioxide modified atmosphere on the compressibility of stored baked goods . J. Food Sci. , 50 : 1172 – 1176 .
- Corsetti , A. , Gobetti , M. , Balestrieri , F. , Russi , L. and Rossi , J. 1998 . Sourdough lactic acid bacteria effects on bread firmness and statling . J. Food Sci. , 63 ( 2 ) : 347 – 351 .
- Baik , M. and Chinachoti , P. 2000 . Moisture redistribution and phase transitions during bread staling. Cereal Chem. , 77 ( 4 ) : 484 – 488 .
- Beuchat , L. R. 1981 . Microbial stability as affected by water activity . Cereal Foods World , 26 ( 7 ) : 345 – 349 .
- Czuchajowska , Z. , Pomeranz , Y. and Jefers , H.C. 1989 . Water activity and moisture of dough and bread . Cereal Chem. , 66 ( 2 ) : 128 – 132 .
- Cazier , J.B. and Gekas , V. 2001 . Water activity and ist prediction: a review . International Journal of Food Properties. , 4 ( 1 ) : 35 – 43 . [CROSSREF]
- Pai , Y.Y. and Walker , C.E . 2001 . AACC Annual Meeting , October 14–18
- Osterwind , G. and Pagenstedt , B. 1982 . Herstellung und vertriben von vorgebackenen backwaren . Getreide Mehl und Brot , 36 : 97 – 102 .
- Leuschner , R.G. , Wehrle , K. and Arendt , E.K. 1996 . Development of a part-baked brown soda bread . J. Agric. and Food Research , 35 : 199 – 200 .
- Jackel , S. 1980 . Natural breads may cause microbiological problems . Bakery Production and Marketing , 15 : 138
- Bailey , C.P. and von Holy , A. 1993 . Bacillus spore contamination associated with commercial bread manufacture . Food Microbiol. , 10 : 287 – 294 . [CROSSREF]
- Leuschner , R.G.K. , O'Callaghan , M.J.A. and Erendt , E.K. 1998 . Bacilli spoilage in part-baked and rebaked brown soda bread . Int. J. Food Sci. Tech. , 65 ( 5 ) : 915 – 918 .
- Rosenkvist , H. and Hansen , A. 1995 . Contamination profiles and characteristics ofBacillus species in wheat bread and raw materials for bread production . Int. J. Of Food Mic. , 26 : 353 – 363 . [CROSSREF]
- Anonymous . American Association of Cereal Chemist. . 1983 . Approved Methods of the AACC. Methods 10/09, approved September 1985; 44-15, approved October 1975, revised October 1981; 44–18, approved April 1961, reviewed October 1982. The Association: St, Paul, M. N
- Elgün , A. , Ertugay , Z. , Certel , M. and Kotancilar , H.G. 1999 . Guide book for analytical quality control and laboratory for cereal and cereal products , 238 Erzurum : Ataturk Univ. Agric. Fac. Publication No:335 .
- Münzing , K. 1987 . Wasser ein wichtiger physikalllisc her qualitatsfaktor bei getreide . Getreide Mehl und Brot , 41 ( 12 ) : 362
- Baumgart , J. , Firnhaber , J. and Spacher , G. 1993 . Microbiologische Untersuchung von Lebensmittein , Hamburg, Germany : Behr's Verlang .
- Viljoen , C.R. , Holy , A.V. and Von , H.A. 1997 . Microbial population associated with commercial bread production . J. Microbiology , 37 ( 6 ) : 439 – 444 .
- Patriarca , A. , Vaamonde , G. , Fernández , V. , Comerio , P. and Comerio , R. 2001 . Influence of water activity and temperature on the growth of Wallemia sebi: application of a predictive model . International Journal of Food Microbiology , 68 : 61 – 67 . [PUBMED] [INFOTRIEVE] [CROSSREF]
- Lin , D.D. and Chen , T.C. 1995 . Relative antifungal afficacies of phosphoric acid and other compounds on fungi isolated from poultry feed . Animal Feed Sci. and Tech. , 54 : 217 – 226 . [CROSSREF]
- Anonymous . A guide to baking preservatives . 1996 . Lallemand Baking Update. American Yeast Sales. 1 (1)
- Ünlütürk , A. , Turantas , F. , Acar , J. , Karapinar , M. , Temiz , A. , Aktug Gönül , S and Tunçel , G. 1999 . Gida Mikrobiyolojisi . Mengi Tan Basimevi, Çinarli-Izmir , : 598
- Elgün , A. and Ertugay , Z. 2000 . Tahil Isleme Teknolojisi , 481 Erzurum : Atatürk Üniv. Ziraat . Fak., Yayin No: 97 .
- Beuchat , L.R. 1978 . Food and Beverage Mycology , 45 – 82 . Westport, Connecticut, USA : AVI Pub. Com. Inc. .