New antimicrobial active package for bak

New antimicrobial active package for bakery products Laura Gutie´rreza, Cristina Sa´ncheza,b, Ramo´n Batllea,b and Cristina Nerı´na,* University of Zaragoza, Marı´a de Luna 3, 50018 Zara

New antimicrobial

active package for

bakery products

Laura Gutie´rreza, Cristina

Sa´ncheza,b, Ramo´n Batllea,b and

Cristina Nerı´na,*

University of Zaragoza, Marı´a de Luna 3, 50018

Zaragoza, Spain (Tel.: D34 976761873; fax: D34

12, 22600 Sabin˜a´nigo, Huesca, Spain

The use of essential oils as food preservatives has gained a great

deal of attention over the last years Specifically, the protective

potential achieved by their addition to the packaging material

has been demonstrated in vitro Nevertheless, there is very little

information available concerning their use with real, complex

food and no procedure for quality evaluation is readily available

This manuscript presents a number of alternatives useful for

active essential oil-based packaging as well as a procedure for

sensory evaluation that can be easily implemented in different

food products The use of a cinnamon-based active package

has been proven to increase more than three times the product

shelf-life of a complex bakery product with a minimal change

in the packaging and no additional manipulation steps The

quality and definition characteristics were not altered by the

use of this concept Product is nowadays in the final steps prior

to market introduction

Introduction

Fungi are the most common spoilers in bakery products

Commonly, a shelf-life around 3e4 days may be expected

when they are unpreserved Apart from the repelling sight

of visible growth, fungi are responsible for off-flavour

for-mation and the production of mycotoxins and allergenic

compounds The use of weak organic acids such as pro-pionic, benzoic, and sorbic, investigation on the packaging material, or modified atmosphere packaging in the last years have been the main choice for satisfying the market demands to extend the shelf-life of bakery products (Ferna´ndez, Vodorotz, Courtney, & Pascall, 2006; Legan, 1993; Pagani, Lucisano, Mariotti, & Limbo, 2006) Specif-ically, ethanol has been commercially used in the form of

a sachet or incorporated into the packaged material; however, this option has a negative public perception, raises the chances to develop a residual flavour and implies

preservatives, ultraviolet light and aseptic packaging have been proposed for controlling microbial growth in bread Nowadays, consumers show preferences for products without preservatives but keeping free from microbial growth, toxins and other quality deteriorating factors

challenge for the food industry is to fulfil these demands with minimum change in food quality and maximum

Devlieghere, van Beest, de Kruijf, & Debevere, 1999) Active packaging is a very interesting alternative to both the use of preservatives or modified atmosphere packaging MAP It involves the incorporation of agents in the packag-ing that can either interact directly with the packaged foodstuff or with the atmosphere inside the package The development of active materials with properties for enhanc-ing the shelf-life and safety of packaged food is nowadays one of the most challenging research activities Several ap-proaches have been proposed but only a few of them have

et al., 2006; Suppakul, Miltz, Sonneveld, & Bigger, 2006; Tovar, Salafranca, Sa´nchez, & Nerı´n, 2005) and

2005; Suhr & Nielsen, 2005) Some antimicrobial films have already been evaluated for food-packaging applica-tions, such as alpha- and beta-cyclodextrin encapsulated allyl isothiocyanate in polylactide-co-polycaprolactone

et al., 2008); blends of gliadins and chitosan ( Ferna´ndez-Saiz, Lagaro´n, Herna´ndez-Mun˜oz, & Ocio-Ferna´ndez-Saiz, 2008) or

& Debevere, 2000); polyethylene coated with an

* Corresponding author.

0924-2244/$ - see front matter Ó 2008 Published by Elsevier Ltd.

doi:10.1016/j.tifs.2008.11.003

nisin-incorporated cross-linked

Although the antimicrobial properties of natural extracts

have been known for centuries, only limited publications

Sanchez, Batlle, & Nerı´n, 2007b; Matanet al., 2006;

Rodrı´-guez, Nerı´n, & Batlle, 2008) The use of natural extracts,

such as essential oils (EO hereafter) and their constituents,

113/EC, 2002; 2004/1935/EC, 2004; 89/107/EEC, 1989);

and as GRAS (Generally Recognized as Safe) by the US

Food and Drug Administration, in plastic films to avoid

microbial food spoilage is an attractive option for both

packaging manufacturers and demanding consumers

In these concepts, the active substances are released

from the packaging material to the surface of the product

during the whole shelf-life of the packaged foodstuff

Anti-microbials incorporated into packaging materials can,

therefore, control microbial contamination by reducing

the growth rate and maximum growth population and/or

extending the lag-phase of the target microorganism or by

Vicini, 2002) The usefulness of these packaging

alterna-tives have been demonstrated at a laboratory scale using

2007a; Lo´pezet al., 2007b) and even some of them have

far more information is needed on their performance

when working with commercial products

Therefore, this paper has two main aims First, to test the

usefulness of the natural extract-active film developed for

its use with a commercial complex bakery product Second,

to fully evaluate the sensory characteristics of the proposed

setup in order to detect potential drawbacks or pitfalls not

related with food objective quality but to food subjective

perception Obviously, these results constitute the final

decision tool for launching into the market the packaged

food product

Essential oils and antimicrobial films

(cinnamon, Chemical Abstract Service, CAS, number:

8015-91-6) was supplied by Argolide Quı´mica S.L

(Barce-lona, Spain) The antimicrobial films were prepared in the

laboratory by incorporating known concentrations (w/w)

of the EO in films of polypropylene (PP hereafter) (30-mm

thick, supplied by Poligal, S.A., Naro´n, Spain) suitable for

a European Patent EP1657181 held by the company

ARTIBAL S.A (Sabin˜a´nigo, Spain)

Bakery product

A grand total of 54 independent samples of the bakery

product from different batches were used in this study All

of them were supplied by a small size company interested

in the study Because of the commercial interest of this

development, a confidential agreement applies and no more data about the company will be provided The product

is a complex mixture prone to microbial infestation and quality deterioration Yolk, almonds, nuts, raisins swilled

in rum as well as other common ingredients are included

in the formula The whole product is covered with a sugar layer

The main desirable characteristics of the product were defined as tenderness of the inner part while nuts and almonds kept crunchy and the yolk cannot be visually distinguished from the mixture and only the taste can be perceived This product has an additional challenge as the requirement of humidity to keep tenderness should not influence the crunching and flavouring of the other ingredi-ents This product is well balanced just made and the same properties are required for its shelf-life No preservatives are added during its baking In fact, this is considered as

a handmade product

Headspace-single drop microextraction (HS-SDME) The atmosphere generated inside the active bag was sampled using headspace-single drop microextraction

Nerı´n (2007) Briefly, a 5-mL microsyringe with a 26S type needle (Hamilton 85RN, Bonaduz, Switzerland) containing p-xylene as the extractive phase was introduced inside the bag The plunger was then depressed and a 2.5-mL drop was exposed to the sample for 5 min After exposure, the drop was retracted again into the syringe and then, it was transferred to the injection port of a GCeMS system Gas chromatographyemass spectrometric

(GCeMS) analysis GCeMS analysis were performed using a Hewlett-Pack-ard 6890 chromatograph (Wilmington, DE, USA) equipped with a 5973 mass selective detector and a HP-5 MS

The temperature program for the GC was as follows: initial

injection mode with the valve closed for 1 min Helium (99.9999% pure, Carburos Meta´licos, Zaragoza, Spain) was used as the carrier gas at a constant flow of 1 mL per

mode was electron impact (70 eV) The mass selective detec-tor was operated in the scan mode between 45 and 400 m/z Sensory testing

A trained panel of 12 individuals was used to evaluate the sensorial properties and possible changes in the bakery product by each different option of packaging under study This was made in order to verify that the extension in shelf-life would not affect the quality of the product The technique used in this case was the classification of the responses given by the tasters in a scale where several

descriptors, defined according their relevance in the final

quality, were qualified The perception of the tasters

concerning the magnitude of the differences, the intensity

of the descriptors related to taste, flavour, sight, color and

tender were evaluated in a scale ranging from 1 to 8 points,

being 8 the highest intensity Three experts (members of the

staff of the manufacturing company) were also included in

the panel as a reference

Testing was conducted over a 28-days period, covering

a time-span longer than the required shelf-life, established

by the manufacturing company in 10 days Currently the

shelf-life is no longer than three days The following

check-points were scheduled (reported as days from

manufacturing): 0, 3, 6, 10, 14, 21, and 28 As a reference,

recently just made products were introduced in the trials

Packaging options

The bakery samples were introduced in a bag made

using all the PP packaging materials and different options

evaluated Then, the plastic bags were thermosealed in all

cases The following options were evaluated in the tests

1 Paraffin paper This is the traditional packaging

material used to cover this specific product by the

manufacturing company (Blank)

2 PP non-active nor micro perforated (PPno)

3 PP non-active micro perforated with a density of 16

4 PP active (4% w/w) micro perforated with a density of

5 PP active (2% w/w) partially micro perforated (3 rows

of 5 cm each) (3 row)

6 PP active (2% w/w) partially micro perforated (2 rows

of 5 cm each) (2 row)

piece to separate the bottom of the product from the

board secondary package (coated board)

metalized and introduced into the active PP bag

(alumi-num tray)

In every test, fresh, just made products in the traditional

packaging material were used as blank samples All options

were kept inside the cardboard box used for the company as

secondary packaging for commercial and marketing

purposes

Development

In a preliminary step, the suitability of the packaging

material intended to come into contact with food was

evaluated As has been described in the introductory

section, no regulatory issues arise from the use of

essen-tial oils as preservative Nevertheless, the first stage

consisted of measuring the volatile compounds released

by the active material to get an idea about the potential

impact characteristics of the product The rationale for

Batlle, & Nerı´n, 2006), the release and identification

of the active compounds from the active film was determined

Fig 1 shows the chromatograms obtained when sam-pling the headspace generated inside the bag using the SDME GCeMS strategy described in the Experimental Section As can be seen, the very same chemicals were detected with (top graph) (PPM) or without (bottom graph) (PPnoM) active film and they are included in the character-istic flavour of the product Nevertheless, when the active packaging (PPM) was analysed in the absence of the bakery product, a clear signal corresponding to cinnamaldehyde (chemical responsible of the cinnamon characteristic flavour) was obtained Therefore, it is hypothesized that this chemical is readily absorbed by the product and as

a consequence, it was included in the flavour descriptors

as an undesirable characteristic

Different groups of descriptors were identified and used in the sensory evaluation They can be roughly divided into five groups: taste, odour (flavour), texture, general appearance and presence of moulds Every group was fully divided into more specific indicators, which were included into the evaluation sheet These indicators were as follows

1 Taste: the following individual markers were defined and included

 Characteristic taste, as a positive factor It repre-sents the similarity obtained by the individual between the test and the blank items

 Cinnamon taste This factor can be considered either positive or negative, since acceptance of this specific taste is very subjective Nevertheless, since it represents an alteration of the expected taste, it was included as negative

 Board taste This is a negative factor and repre-sents the alteration due to the board material used as secondary packaging

2 Flavour

 Board flavour This is also a negative factor and represents alteration due to the board material used as secondary packaging

 Cinnamon flavour As in the previous paragraph, it was considered as negative factor

3 Texture

 Tenderness Positive factor, representing the appro-priate resistance of the product when chewed, relative to the blank item

 Crunchiness and crispiness Positive factor, rela-tive to the consumer’s perception of the nuts when compared to the blank item

 Elasticity rubber-like Negative factor, referring to the change in the product overall texture It was defined by the expert members of the sensory eval-uation board as a clear indicative of aging

Fig 1 Typical gas-chromatograms obtained inside the PP bag in the final commercial disposition with packaged product Top panel, active film used

(PPM, option 4); bottom panel, raw PP used (PPnoM, option 3).

4 General appearance.

 Freshness Positive, description of the overall

perception of the product when compared to the

blank item

 Fat stains Negative, with the original packaging, it

was observed that stains appeared with time on the

bottom of the board secondary package They were

attributed to the fat migration from the packaged

product, and severely detract from marketability

5 Presence of moulds Zero-tolerance was defined for this

descriptor, meaning that it was considered as an

ONeOFF test, concerning the final acceptance of the different packaging options Nevertheless, for clarity purposes, the relative abundance of the moulds onto the packaged foodstuffs was represented by a numerical value: 1 represents total absence of visible growth, whereas 7 indicates complete contamination by moulds

Fig 2AeC illustrates the evaluation results of the sen-sory attributes obtained when using the active package approaches (three independent replicates were analyzed for each control point) As can be seen, a clear

A

0 1 2 3 4 5 6 7 8 9

3 Days

Fresh Appearance

Crunchy and Crispy

Characteristic Taste

Fresh Appearance

Crunchy and Crispy

Characteristic Taste

Fresh Appearance

Crunchy and Crispy

Characteristic Taste

B

0 1 2 3 4 5 6 7 8

Sample

Sample

Sample

7 Days

C

0 1 2 3 4 5 6 7 8

10 Days

Fig 2 Results obtained in the evaluation of the diverse active packages For package identification, please proceed to the experimental section All

the results represent the average of three replicates.

improvement with respect to the traditional package was

detectable even at the initial stages (day 3) of the evaluation

was demonstrated It is worth to highlight that the

punctu-ations obtained were consistently high for all the active

so-lutions at day 7 and that the decrease was only detected at

day 10, varying among the various active solutions Taking

into account the scale used for the different descriptors and

the expectancies, an average value of 5 was defined as an

acceptable product Only the aluminum tray packaging

(A Tray, option 8) gave these results at day 10 To explain

in detail these achievements, it is interesting to go deeper

into the results The next set of spider plots illustrate in

detail the results obtained for each packaging combination

at day 10 Spider plot was selected since it provides a very

intuitive and direct view of the results; factors were

arranged so that parameters considered as negative are at

the top and the positive attributes can be seen at the bottom

of the figures Every spider plot represents the average of

three independent replicates

(top) was changed to raw PP (middle graph) significant

wors-ening was obtained for the elasticity descriptor, whereas the

other factors remained somehow unchanged with only slights

improvement for characteristic taste and fat stains As a first

modification, a physical change in the packaging material

(micro perforation, as described in the materials and methods

section) was then evaluated (bottom graph) and proven only

successful in reduction of the rubber-like perception As can

be seen, very low numbers (2 or 3) were obtained for the

positive factors, whereas very high values were obtained

for the negative ones, reflecting the clear fact that the product

does not fit the required quality standards

No significant improvement in the positive factors other

than the tenderness (2 to 3) was found Considering these

results together with the ones described in the previous

paragraph it seems clear that the combination of different

mi-cro perforation densities to reduce elasticity rubber-like and

active packaging to reduce presence of moulds could

consti-tute a better approach Three different densities were tested

a reduction of presence of moulds from 7 without active

packaging to 1 for the active option was observed Middle

and bottom graphs gave best results for cinnamon perception

while maintaining the antifungal capabilities required

Moreover, these combinations increased the scores in the

positive factors, especially in characteristic taste, as could

be expected The last challenge was to reduce the high scores

obtained by the factors that can be related with the secondary

package-board interaction such as board taste and flavour, as

well and fat stains To do so, the best active plastic package

results were combined with different secondary package

tray coated with PE layer (package option 7) was introduced

between the secondary package and the product in order to

minimize the transference of board characteristics to the

success since a reduction in scores for these factors was obtained as well as a significant increase in positive factors such as tenderness and freshness with no worsening of any desirable performance In order to achieve a final improve-ment of the package, the PE-coated piece was metalized (package option 8) to further minimize the interaction between the secondary package and the product As depicted

in Fig 5 (bottom), the results obtained were very good, providing the lowest scores for the negative factors and the maximum scores for the positive ones (with the only excep-tion of the freshness which provides a score of 7 instead of 8)

0_Parafina

1_PP non active no microperforated 2_PP non active microperforated

Fig 3 Non-active packaging options: spider plots obtained after sen-sory evaluation at day 10 Top, traditional (blank, option 1) package; middle, PPno (option 2); bottom, micro perforated PPnoM (option 3).

This work has illustrated the successful design,

develop-ment and testing of an active package based on the use of

cinnamon essential oil in combination with micro

perfo-rated polypropylene The final design provided an increase

in shelf-life from 3 to 10 days with maximum quality and safety, as demonstrated by the sensory evaluation results The evaluation procedure described (including factor defi-nition and evaluation) can be easily adapted to any bakery product and could be established as a standard for this development Further work is currently under way for industrial scale-up Hopefully, the product will be in the market in the near future

Acknowledgments This work has been financed by the Research Projects Cal03-080 from INIA and FEDER; AGL2004-07545 from the Spanish Ministry of Education and University and FEDER and INTERREG IIIA-5-326 C L.G acknowl-edges the Spanish Ministry of Education and University for a grant (BES-2005-10186) R.B expresses his grati-tude to the former Spanish Ministry of Science and

Fig 4 Combination of micro perforation and active concentration:

spider plots obtained after sensory evaluation at day 10 Top, PP

(4%) active micro perforated (PPM, option 4); middle, PP (2%) active

partially micro perforated (3 Row, option 5); bottom, PP (2%) active

partially micro perforated (2 Row, option 6).

1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.0010.00

Fresh Appearence Tenderness

Board Taste

Cinnamon Taste

Presence of Molds

Elasticity Rubber Like Board Flavour

Characteristic Taste

Crunch and Crispy Fat Stains

1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00

Fresh Appearence Tenderness

Board Taste

Cinnamon Taste

Presence of Molds

Elasticity Rubber Like Board Flavour

Characteristic Taste

Crunch and Crispy Fat Stains

Fig 5 Different alternatives tested to minimise secondary packag-ing-food alterations: spider plots obtained after sensory evaluation

at day 10 Top, use of PE-coated board (option 7); bottom, metalized

PE-coated board (option 8).

Technology for personal funding through the Ramo´n y

Cajal program

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