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Sensory and Nutritive Qualities of Food

Phytic Acid Degradation in Complementary Foods Using Phytase Naturally Occurring in Whole Grain Cereals
I. EGLI, L. DAVIDSSON, M.-A. JUILLERAT, D. BARCLAY, AND R. HURRELL ABSTRACT: Complementary foods based on cereals and legumes often contain high amounts of phytic acid, a potent inhibitor of mineral and trace element absorption. The possibility to degrade phytic acid during the production of complementary foods by using whole grain cereals as the phytase source was investigated. Whole grain rye, wheat, or buckwheat (10%) were added to cereal-legumebased complementary food mixtures, and phytic acid was shown to be completely degraded in a relatively short time (1.5 to 3 h) when incubated at optimal conditions for cereal phytase. The potential usefulness of the method for industrial production was demonstrated with a complementary food based on wheat and soybean. Keywords: phytic acid, phytase, complementary foods, cereals, food processing

Introduction
combined with milk, or with legumes in countries where milk is not readily available, to improve the protein quantity and nutritional quality. Cereal grains and legume seeds generally contain high amounts of phytic acid (myo-inositol 1,2,3,4,5,6 hexakis [dihydrogen phosphate]), which binds strongly to minerals and trace elements. Mineral and trace element requirements are high during early life due to rapid growth and development. To ensure adequate bioavailability of these nutrients from complementary foods should therefore be a priority. The enzyme phytase (myo-inositol hexakisphosphate phosphohydrolase) degrades phytic acid (IP6) to lower inositol phosphates (myo-inositol pentaphosphate [IP5], myo-inositol tetraphosphate [IP4], myo-inositol triphosphate [IP3], myo-inositol diphosphate [IP2], myo-inositol monophosphate [IP1]) and/or myo-inositol and inorganic phosphate. The inhibitory effect of phytic acid, resulting in reduced iron and zinc bioavailability in humans, was shown to be mainly caused by IP6 and IP5 (Sandstrm and Sandberg 1992; Sandberg and others 1999). Lower inositol phosphates bind less strongly to minerals and trace elements. The positive effect of phytic acid degradation or removal on iron absorption and zinc absorption from meals based on cereals or legumes has been shown in several studies in adults and infants (Kivist and others 1989; Davidsson and others 1994; Larsson and others 1996; Cook and others 1997; Hurrell and others 2002). Phytase occurs naturally in cereal grains and legume seeds as well as in microorganisms. The phytase activities of grains and seeds vary widely; rye and wheat show rather high activities, whereas legumes and the cereals maize, millet, oat, and sorghum were found to have rather low phytase activities (Bartnik and Szafranska 1987; Scott 1991; Egli and others 2002). The highest phytase activities have been reported in microorganisms of the Aspergillus species (Shieh and Ware 1968; Howson and Davies 1983). Microbial phytases, extracted or genetically engineered, are available but have not, to our knowledge, been used to degrade phytic acid in commercial products for human consumption. A more ac 2003 Institute of Food Technologists
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OMPLEMENTARY FOODS ARE OFTEN BASED ON CEREALS AND ARE

ceptable alternative in food production could be to use the enzymatic activity of phytase naturally occurring in the ingredients of cereal-based foods. Phytase in grains and seeds can be activated by traditional food processing methods such as soaking, germination, and fermentation to decrease the phytic acid content in complementary and other foods (Gupta and Sehgal 1991; Marero and others 1991; Svanberg and others 1993; Sanni and others 1999; Porres and others 2001). However, these processing methods change the composition, viscosity, and taste of the complementary foods considerably and might result in products with low consumer acceptability. In addition, the complete phytic acid degradation, necessary to improve iron absorption meaningfully (Hurrell and others 1992) generally requires prolonged fermentation and therefore might introduce problems of microbiological safety. In this study, the feasibility of complete phytic acid degradation, using phytase from whole grain cereals, was investigated by the inclusion of an incubation step in the production of complementary foods. Conditions were optimized during laboratory experiments to achieve complete phytic acid degradation, followed by the production of a phytic acid free complementary food in a pilot plant.

Materials and Methods

LL CHEMICALS AND REAGENTS WERE OF ANALYTICAL GRADE QUALITY.

For laboratory experiments, water was purified by reverse osmosis (Nanopure Cartridge System, Skan AG, Basel, Switzerland). Tap water was used for pilot plant trials.

Grains and seeds

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For laboratory experiments, whole grain rye and whole grain wheat were purchased from a commercial seed supplier (Fenaco, Winterthur, Switzerland). Whole grain maize, whole grain buckwheat, polished rice, and wheat flour with low extraction rate were purchased in supermarkets or health stores in Zurich, Switzerland. Nestl, Singapore, provided whole grain chickpea, and dehulled and toasted soybean was purchased from Zwicky AG, MllheimWigoltingen, Switzerland. The grains and seeds were stored at 4 2 C.

Phytic acid degradation . . .


For pilot plant trials, dehulled and toasted soybeans were purchased from Zwicky AG and whole grain wheat and wheat flour with low extraction rate from Moulins Cossonay SA, Cossonay-Ville, Switzerland. Wheat flour with low extraction rate was classified as wheat flour 550 according to the ash content (Arens 1992). All grains, seeds, and flour were stored at ambient temperature.
Table 1Composition (weight %) of the cereal-legume mixtures used for phytic acid degradation in laboratory experiments Cereal component Legume component Phytase source 10% whole grain wheat 10% whole grain rye 10% whole grain buckwheat None 10% whole grain wheat

Milling
Grains and seeds used in laboratory experiments were frozen in liquid nitrogen and milled with a centrifugal mill (0.5 mm mesh). Dehulled and toasted soybeans and whole grain wheat for pilot plant trials were milled at Nestl Product Technology Centre, Orbe, Switzerland.

90% whole grain maize None 90% whole grain maize None 30% polished rice 60% whole grain chickpea 33% polished rice 67% whole grain chickpea 70% wheat with low 20% dehulled, extraction rate toasted soybean

Phytic acid
The phytic acid content of the milled grains and seeds was determined according to Sandberg and Ahderinne (1986) and Sandberg and others (1989) with modifications (Egli and others 2002). Samples were analyzed in duplicate and results expressed as the sum of IP6 and IP5 in mg/g. The difference of duplicate analysis relative to the mean value was < 10%. Hard red wheat bran (Lot 195, 1995, American Assn. of Cereal Chemists, St. Paul, Minn., U.S.A.) was analyzed together with each series of samples and was used as an internal control material to monitor reproducibility. The determination limit for IP6 was 0.03 mg/g. terminated by the addition of 70 mL 0.57 M hydrochloric acid. Phytic acid and lower inositol phosphates were extracted under constant stirring (600 rpm) for 3 h at room temperature. About 30 mL of the sample solution were centrifuged at 3500 rpm for 10 min, and 15 mL of the supernatant were used for the inositol phosphate determination as described earlier. The contents of IP3, IP4, IP5, and IP6 were expressed as mg/g. The coefficient of variation based on triplicate analysis for each time point was < 15%.

Phytic acid degradation in complementary food production


Complementary food production was carried out in a pilot plant (Nestl Product Technology Center, Orbe). The complementary food mixture (total weight 40 kg) consisted of the milled components in the following proportions: 70% wheat with low extraction rate, 20% dehulled and toasted soybeans, and 10% whole grain wheat. Water (100 kg) was heated to about 55 C in a tank, and the complementary food mixture was added under constant stirring. The pH was adjusted with 1 M citric acid to pH 5.1. Aliquots (about 10 g of the slurry) were weighed into 100 mL polyethylene bottles containing 70 mL 0.57 M hydrochloric acid to terminate the enzymatic reaction. Aliquots were taken every 20 min, and the inositol phosphates were determined as described in the laboratory experiments. The difference of duplicate analysis relative to the mean value was < 15%. The pH value was measured at the beginning of the incubation and every 60 min thereafter. The temperature was monitored continuously. After 180 min, the slurry was heated by steam injection (about 135 C) and roller dried.

Phytase assay
The milled grains and seeds were screened for apparent phytase activity. About 1 g milled grains or seeds was added to 20 mL buffer (0.2 M acetate buffer, pH 5.0) containing 7.5 mM phytic acid prepared from phytic acid dodecasodium salt (Sigma-Aldrich Chemie GmbH, Steinheim, Germany). The measurement was performed for 1 h at 45 C under constant stirring with aliquots taken every 20 min. The reaction was terminated by adding 0.5 mL 0.9 M trichloroacetic acid to each 0.5 mL aliquot. Inorganic phosphate was determined colorimetrically (Van Veldhoven and Mannaerts 1987) at 4 times (0, 20, 40, 60 min). Inorganic phosphate was liberated at a constant rate, and apparent phytase activity was calculated by linear regression of the inorganic phosphate determined for each time point. Samples were analyzed in triplicate and the phytase activity expressed in phytase units (PU) per g. One phytase unit is equivalent to the enzymatic activity that liberates 1 mol inorganic phosphate per min. Additional experiments were performed to determine the pH and temperature conditions for maximum phytase activity of whole grain rye, whole grain wheat, and whole grain buckwheat. The pH was adjusted to values in the range of 4.0 to 6.0 using 0.2 M acetate buffer. The temperature varied between 25 C and 65 C. All other conditions and procedures were as described above.

Results and Discussion


Phytic acid content and phytase activity of experimental grains and seeds
The grains and seeds used for laboratory experiments and complementary food production were analyzed for their phytic acid content and their phytase activity (Table 2). The phytic acid content ranged from 1.3 mg/g for polished rice to 10.7 mg/g for soybean. Whole grain rye, wheat, and buckwheat were selected as phytase sources because they showed high apparent phytase activities in an earlier study (Egli and others 2002). In the present study, the phytase activities were comparable and were 6.0 PU/g for whole grain rye, 2.2 PU/g for whole grain buckwheat, 2.7 PU/g for whole grain wheat (laboratory experiments), and 3.4 PU/g (complementary food production). The cereals maize, rice, and wheat with low extraction rate and the legumes chickpea and soybean were chosen as ingredients for complementary food mixtures in the form they are commonly used in the food industry. All these ingredients showed low phytase activities in the range of 0.04 to 0.6 PU/
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Phytic acid degradation in laboratory experiments


Phytic acid degradation was investigated in various cereal and cereal-legume mixtures. Milled grains and seeds (total weight 50 to 100 g) were weighed into 500-mL polypropylene containers in the proportions shown in Table 1. They were mixed by vigorous shaking for about 5 min and aliquots (2 to 4 g) of the mixtures were weighed into Erlenmeyer flasks and heated to the incubation temperature (50 C). Citric acid (10 mL, 2 to 11 mM depending on the mixture and target pH) were heated to the incubation temperature and then added to form a slurry. The Erlenmeyer flasks were covered and placed in an incubator (50 C) under constant stirring (600 rpm). Incubation times varied between 20 and 240 min depending on the composition of the mixtures. The enzymatic reaction was
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Phytic acid degradation . . .


g. Because the differences in the phytic acid content and in the phytase activities between the grains and seeds used for laboratory experiments and for the complementary food production were very small, no substantial influence on phytic acid degradation was expected between the 2 sets of experiments.
Table 2Phytic acid content and phytase activity of grains and seeds used in laboratory experiments and complementary food production Grains and seeds Laboratory experiments Buckwheat (whole grain) Chickpea (whole grain) Maize (whole grain) Rice (polished) Rye (whole grain) Soybean (dehulled, toasted) Wheat (whole grain) Wheat (low extraction rate) Phytic acid (mg/g) a 10.0 4.4 9.0 1.3 6.8 9.0 8.9 1.9 Phytase activityb (PUc/g)d 2.17 0.02 0.22 0.02 0.11 0.00 0.04 0.00 6.01 0.35 0.22 0.03 2.65 0.15 0.60 0.02 3.42 0.01 0.52 0.02 0.16 0.01

Optimum reaction conditions for rye, wheat, and buckwheat phytase


For the phytase sources (whole grain rye, whole grain wheat, and whole grain buckwheat), the pH and temperature conditions were optimized so as to achieve maximum enzymatic activities. The optimum pH for whole grain rye (5.5) described by Greiner and others (1997) was confirmed, whereas the optimum temperature was found to be slightly higher, 55 C, as compared with 48 C (Greiner and others 1997). For whole grain wheat, the optimum pH (5.15) reported by Peers (1953) was confirmed. The optimum temperature was slightly higher than 55 C, as reported by Peers (1953). The optimum pH and temperature conditions for buckwheat phytase had not been reported previously and were found to be pH 5.0 and 55 C. In the following experiments, the incubation conditions were adjusted to the optimum pH for the respective phytase source and 50 C was used as the maximum temperature to avoid thermal denaturation of the enzymes.

Complementary food production Wheat (whole grain) 9.2 Wheat (low extraction rate) 1.5 Soybean (dehulled, toasted) 10.7

a Values represent mean of duplicate analysis b Measured at pH 5.0, 45 C c 1 phytase unit (PU) is equivalent to the enzymatic activity that liberates 1

mol inorganic phosphate per min.


d Values represent mean SD of triplicate analysis.

Phytic acid degradation in laboratory experiments


The phytic acid degradation was 1st tested in maize to compare the impact of 2 phytase sources: whole grain wheat and whole grain rye. Figure 1 and 2 show the phytic acid degradation in slurries based on whole grain maize with the addition of 10% whole grain wheat or 10% whole grain rye, respectively. The temperature (50 C) and the dry matter content of the slurries (15%) were identical, but the pH was adjusted to 5.2 in the maize-wheat and to 5.5 in the maize-rye mixture. Phytic acid degradation using whole grain wheat as the phytase source was relatively slow. After 180 min of incubation, 0.2 mg/g IP6 and 0.2 mg/g IP5 remained, as well as 2.1 mg/g IP4 and 2.8 mg/g IP3. Complete degradation required 6 h of incubation. When rye was used as the phytase source, IP5 and IP6 were

no longer detected after 120 min. After 180 min only 0.1 mg/g IP3 was left, and no IP3 was detected after 240 min. The results of the phytic acid degradation in the maize-wheat and maize-rye mixture were used to verify whether the rapid determination of phytase activity (as described in the phytase assay) might be useful to estimate the time required for phytic acid degradation in cereal-legume mixtures. Theoretically, the addition of 10% whole grain rye (phytase activity 6.0 PU/g) should almost completely degrade all phytic acid (about 10 mg/g) in the maize-rye mixture in about 150 min. The results demonstrate that complete phytic acid degradation took slightly longer as the reaction slowed down with prolonged incubation time, most probably due to available substrate (phytic acid) concentration. In the phytase assay the enzymatic activity is measured in the initial phase of the reaction; that is, when inorganic phosphate is liberated at a constant rate, and

Figure 1Phytic acid degradation in maize (90%) with added whole grain wheat (10%). Incubation conditions: pH 5.2, temperature 50 C.
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Figure 2Phytic acid degradation in maize (90%) with added whole grain rye (10%). Incubation conditions: pH 5.5, temperature 50 C. Vol. 68, Nr. 5, 2003JOURNAL OF FOOD SCIENCE 1857

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Phytic acid degradation . . .


the substrate is available in excess. In the experiments monitoring phytic acid degradation in cereal or cereal-legume mixtures, the free or available substrate becomes a limiting factor and therefore slows down the reaction. Taking limited substrate into account as well as differences in the incubation conditions, the results of phytic acid degradation in maize-wheat and maize-rye mixtures demonstrate that the determination of the phytase activity is useful to estimate the potential of a grain to degrade phytic acid, because the phytase activity of rye was more than double that of wheat (Table 2). In the following experiments, cereals and legumes were combined so as to achieve a mixture that could be used as the basis for potentially useful complementary foods with a protein content of about 15%. This is based on the Codex Alimentarius for processed cereal-based foods for infants and children (Food And Agriculture Organization/World Health Organization 1994) proposing a minimum protein content of 15% for dry cereals, intended to be mixed with water before consumption. The selected cereal-legume mixtures were used to test the methodology developed and are not meant to be representative for specific regions of the world. Phytic acid degradation was 1st monitored in a mixture based on polished rice (30%) and whole grain chickpea (60%) with 10% whole grain buckwheat as the phytase source. Buckwheat has lower phytase activity than wheat and rye, but is gluten-free and thus has other interesting potential. It was tested in a mixture based on rice as an example of a gluten-free cereal. The relatively low initial phytic acid content (3.6 mg/g) was completely degraded after 200 min incubation at 50 C, in a slurry with a dry matter content of 20% and pH 5.0 (Figure 3). In addition, phytic acid degradation was measured under the same conditions in the rice-chickpea mixture without addition of whole grain buckwheat as the phytase source (results not shown). After 200 min, the initial phytic acid content of 3 mg/g was degraded to 0.7 mg/g IP3, 0.7 mg/g IP4, 0.2 mg/g IP5, and 0.1 mg/g IP6, comparable to the content after 120 min in the mixture containing buckwheat. Thus, the low phytase activity of chickpea and rice significantly contributed to the inositol phosphate degradation. However, the addition of buckwheat shortened the incubation time for complete phytic acid degradation considerably. In the next experiment, the phytic acid degradation was investigated in a cereal-legume mixture that is widely used for complementary food production, wheat and soybean. Figure 4 shows the phytic acid degradation in a mixture based on wheat with low extraction rate (70%) and dehulled and toasted soybean (20%) with 10% whole grain wheat as the phytase source. The initial phytic acid content (4.0 mg/g) was decreased to 0.1 mg/g IP6 and 0.1 mg/g IP3 after 60 min incubation at 50 C, in a slurry with a dry matter content of 25% and pH 5.1. The reaction slowed down after 60 min. However, complete degradation was achieved in a relatively short time (120 min), probably due to the contribution of phytase activity in 1 of the major ingredients, wheat with low extraction rate (0.6 PU/g).

Phytic acid degradation in complementary food production


The feasibility of adapting the laboratory method for phytic acid degradation to industrial complementary food production was tested in a pilot plant using the mixture based on wheat and soybean. The conditions (pH, temperature, dry matter content of the slurry) were the same as in the laboratory experiment. The batch size was increased from 4 g to 40 kg. The 1st sample was taken after about 10 min, when the slurry was homogeneous and pH had been adjusted, and every 20 min thereafter. The initial inositol phosphate content was calculated based on the determination of the phytic acid

Figure 4Phytic acid degradation in wheat (70%) and soybean (20%) with added whole grain wheat (10%). Incubation conditions: pH 5.1, temperature 50 C.

Sensory and Nutritive Qualities of Food

Figure 3Phytic acid degradation in rice (30%) and chickpea (60%) with added whole grain buckwheat (10%). Incubation conditions: pH 5.0, temperature 50 C. 1858

Figure 5Phytic acid degradation during complementary food production in wheat (70%) and soybean (20%) with added whole grain wheat (10%). Incubation conditions: pH 5.1, temperature 50 C.
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Phytic acid degradation . . .


content of the different components of the complementary food. During the 1st 10 min IP5 and IP6 decreased to 48% of the initial calculated value (4.1 mg/g) (Figure 5). Complete degradation of inositol phosphates required 90 min. In the laboratory experiments, the dry matter content of the slurries was limited to about 25% due to the capacity of the mixing device. This is a relatively low dry matter content for industrial complementary food production, especially when products are dried by roller drying. Therefore, a pilot plant trial was carried out with increased dry matter content (about 35%), whereas all other conditions remained unchanged (results not shown). The phytic acid degradation during the 1st 50 min of the incubation was comparable with results from the slurry with 25% dry matter content. Complete degradation required a slightly longer incubation time of 150 min.

References
Arens F-J. 1992. Die neue mehltypen-regelung. AID-Verbraucherdienst 37(10):2103. Barclay D, Davidsson L, Egli I, Hurrell R, Juillerat M-A, inventors; Societe des Produits Nestl S.A., Federal Inst. of Technology Zurich, assignee. 2000 December 7. Cereal products having low phytic acid content. WO 0072700. Bartnik M, Szafranska I. 1987. Changes in phytate content and phytase activity during the germination of some cereals. J Cereal Sci 5(1):238. Cook JD, Reddy MB, Burri J, Juillerat M-A, Hurrell RF. 1997. The influence of different cereal grains on iron absorption from infant cereal foods. Am J Clin Nutr 65(4):9649. Davidsson L, Galan P, Kastenmeyer P, Cherouvrier F, Juillerat M-A, Hercberg S, Hurrell RF. 1994. Iron bioavailability studied in infants: the influence of phytic acid and ascorbic acid in infant formulas based on soy isolate. Pediatr Res 36(6):81622. Egli I, Davidsson L, Juillerat M-A, Barclay D, Hurrell RF. 2002. The influence of soaking and germination on the phytase activity and phytic acid content of grains and seeds potentially useful for complementary feeding. J Food Sci 67(9):34848. [FAO/WHO] Food and Agriculture Organization/World Health Organization Codex Alimentarius Commission. 1994. Codex Alimentarius, vol. 4: foods for special dietary uses (including foods for infants and children) Rome: FAO/ WHO. 131 p. Greiner R, Konietzny U, Jany K-D. 1997. Isolierung und Charakterisierung einer phytase aus roggen. Getreide Mehl Brot 51(2):7984. Gupta C, Sehgal S. 1991. Development, acceptability and nutritional value of weaning mixtures. Plant Foods Hum Nutr 41(2):10716. Howson SJ, Davis RP. 1983. Production of phytate-hydrolysing enzyme by some fungi. Enzyme Microb Technol 5(5):37782. Hurrell RF, Juillerat M-A, Reddy MB, Lynch SR, Dassenko SA, Cook JD. 1992. Soy protein, phytate, and iron absorption in humans. Am J Clin Nutr 56(3):5738. Hurrell RF, Reddy MB, Burri J, Cook JD. 2002. Phytate degradation determines the effect of industrial processing and home cooking on iron absorption from cereal-based foods. Br J Nutr 88(2):11723. Kivist B, Cederblad A, Davidsson L, Sandberg AS, Sandstrm B. 1989. Effect of meal composition and phytate content on zinc absorption in humans from an extruded bran product. J Cereal Sci 10(3):18997. Larsson M, Rossander-Hulthn L, Sandstrm B, Sandberg A-S. 1996. Improved zinc and iron absorption from breakfast meals containing malted oats with reduced phytate content. Br J Nutr 76(5):67788. Marero LM, Payumo EM, Aguinaldo AR, Matsumoto I, Homma S. 1991. Antinutritional factors in weaning foods prepared from germinated cereals and legumes. Lebensm Wiss Technol 24(2):17781. Peers FG. 1953. The phytase of wheat. Biochem J 53(1):10210. Porres JM, Etcheverry P Miller DD, Lei XG. 2001. Phytase and citric acid supple, mentation in whole-wheat bread improves phytate-phosphorus release and iron dialyzability. J Food Sci 66(4):6149. Sandberg AS, Ahderinne R. 1986. HPLC method for determination of inositol tri-, tetra-, penta-, and hexaphosphates in foods and intestinal contents. J Food Sci 51(3):54750. Sandberg A-S, Carlsson N-G, Svanberg U. 1989. Effects of inositol tri-, tetra-, penta-, and hexaphosphates on in vitro estimation of iron availability. J Food Sci 54(1):15986. Sandberg A-S, Brune M, Carlsson N-G, Hallberg L, Skoglund E, Rossander-Hulthn L. 1999. Inositol phosphates with different numbers of phosphate groups influence iron absorption in humans. Am J Clin Nutr 70(2):2406. Sandstrm B, Sandberg A-S. 1992. Inhibitory effects of isolated inositol phosphates on zinc absorption in humans. J Trace Elem Electrolytes Health Dis 6(2):99103. Sanni AI, Onilude AA, Ibidapo OT. 1999. Biochemical composition of infant weaning food fabricated from fermented blends of cereal and soybean. Food Chem 65(1):359. Scott JJ. 1991. Alkaline phytase activity in nonionic detergent extracts of legume seeds. Plant Physiol 95(4):1298301. Shieh TR, Ware JH. 1968. Survey of microorganisms for the production of extracellular phytase. Appl Microbiol 16(9):134851. Svanberg U, Lorri W, Sandberg A-S. 1993. Lactic fermentation of non-tannin and high-tannin cereals: Effects on in vitro estimation of iron bioavailability and phytate hydrolysis. J Food Sci 58(2):40812. Van Veldhoven PP, Mannaerts GP. 1987. Inorganic and organic phosphate measurements in the nanomolar range. Anal Biochem 161(1):458. MS 20030011 Submitted 1/7/03, Revised 2/11/03, Accepted 3/17/03, Received 3/26/03
Dr. Josef Burri, Dr. Johan de Reu, and the staff at the pilot plant at Nestl Product Technology Centre, Orbe, Switzerland, are gratefully acknowledged for technical assistance during the production of the complementary foods. The project was financially supported by Nestec SA, Vevey, Switzerland.

Advantages and perspectives of phytic acid degradation with phytase naturally occurring in cereals
This study demonstrates that the newly developed method to degrade phytic acid can be applied to industrial complementary food production by the inclusion of an incubation step, followed by roller drying. The processing conditions (temperature, pH) need to be adapted slightly, but are within acceptable manufacturing conditions for complementary food production. The overall composition of the complementary foods would not change significantly by the addition of 10% whole grain cereals as a source of phytase activity. A major advantage with this approach is that the enzymatic activity would be provided by the whole grain cereals. Thus, the addition of exogenous phytase, for example of microbial origin, is not required. Wheat is one of the most commonly consumed and readily available cereals worldwide and therefore an obvious choice as a source of phytase in the manufacture of phytic acid free complementary foods. For complementary food mixtures with high phytic acid content, the addition of a higher proportion of wheat or the use of rye as a phytase source might be alternatives to achieve complete phytic acid degradation in a relatively short time. Even though the methodology was evaluated in a limited number of cereal-legume mixtures, we would expect similar results in other mixtures potentially useful as complementary foods. Future development of the methodology could include the adaptation to small-scale industries producing low-cost complementary foods based on locally consumed cereals and legumes, especially in developing countries. The method has been published as an international patent application (Barclay and others 2000).

Conclusion
and wheat, can be used to degrade phytic acid completely in cereal-legume-based complementary foods during production. Processing conditions should be modified, based on optimal pH and temperature for the phytase source, to minimize the time required for phytic acid degradation. Results based on laboratory experiments provide useful information for phytic acid degradation in complementary food production, and the time required for enzymatic degradation can be estimated based on the phytase activity and the initial phytic acid content of the mixture. The nutritional benefit of phytic acid free complementary foods would be related to the expected significant increase in bioavailability of iron and zinc.

HOLE GRAIN CEREALS WITH HIGH PHYTASE ACTIVITY, SUCH AS RYE

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Authors Egli, Davidsson, and Hurrell are with the Laboratory for Human Nutrition, Inst. of Food Science, Swiss Federal Inst. of Technology, Zurich, PO Box 474, 8803 Rschlikon, Switzerland. Author Juillerat is with the Nestl Research Centre, Lausanne, Switzerland. Author Barclay is with Nestec Ltd., Vevey, Switzerland. Direct inquiries to author Davidsson (E-mail: lena.davidsson@ilw.agrl.ethz.ch).