Abstract
Fatty acids and cholesterol compositions, in both fresh and mature cheeses, prepared in accordance with the Manchego-type cheese with different fat compositions, using milk enriched with avocado oil were evaluated. Fresh cheese enriched with avocado oil showed an increase of 52% in polyunsaturated acids in full-fat milk and 98% in skimmed milk. Mature manchego style cheeses enriched with avocado oil showed an increase of 32% in polyunsaturated acids in full-fat milk and 61% in skimmed milk. Multivariate analysis (non-linear mapping, hierarchical cluster analysis, and linear discriminating analysis) of the fatty acids composition in newly prepared cheeses confirmed that they were different from the traditional cheeses and that a small set of fatty acids (myristoleic, lauroleic, eicosenoic, caproleic, stearic, palmitoleic, and margaroleic) can be used to develop classification rules.
INTRODUCTION
[Supplementary materials are available for this article. Go to the publisher's online edition of International Journal of Food Properties to view the free supplementary files.]
The Mediterranean diet, based in olive oil, is generally believed to reduce the risk of heart disease, Alzheimer's disease, diabetes, and changes in the oxidative status of tissues causing others diseases.[Citation1–3] Extracted from the pericarp of the olive fruits, it is the major source of edible lipids that are consumed throughout the world.[Citation4] Another important oily fruit is the avocado (Persea americana Mill.), with the main European production of this fruit located in the subtropical regions of Málaga and Granada, in southern Spain. Avocado oil, as a food substance, is used as an ingredient in recipes, as well as cooking oil.[Citation5] In addition, it is used for lubrication[Citation6] and is very sought after in cosmetics, where it is valued for its regenerative and moisturizing properties.[Citation7–9] From the nutritional point of view, avocado is a high calorie fruit with a high unsaturated fatty acids content.[Citation10] It is also rich in vitamin E, ascorbic acid, vitamin B6, β-carotene, and potassium.[Citation11–13] Thus, the industrial production of functional foods based on avocado shows high potential.
Milk products play an important part in a healthy diet, as they contribute to intakes of essential nutrients and protein of high nutritional value. In Spain, dairy milk products contribute more than 50% of the total calcium intake.[Citation14] However, due to the high content of cholesterol raising saturated fatty acids (SFA) in milk fat, a decrease in the intake of fat-rich dairy products is recommended. Although there is no doubt that long-chain SFA in milk fat increases plasma cholesterol,[Citation15,Citation16] there is some controversy regarding the specific effect of milk products. Thus, dairy milk preparations were formulated by the milk industry to improve its natural composition.
This study presents the fatty acids composition and cholesterol contents of a particular dairy prepared cheese, both fresh and mature (ripened for 4 months). It is one of the most popular varieties of cheese in Spain (known as ‘Manchego-type cheese’); it is made with pure ewe milk from the machega breed (Ovis Aries) and is enriched with 2 g avocado oil/100 mL milk. The objective was to obtain a cheese with low cholesterol and saturated fatty acids content and a high percentage of unsaturated fat by use of a milk preparation based in avocado oil.[Citation17] The fatty acids contents were subjected to a multivariate classification (non-linear mapping, hierarchical cluster analysis, and linear discriminating analysis) to check for similarities and differences between the various samples and fatty acids compositions.
MATERIAL AND METHODS
Reagents and Materials
All the chemicals used were of the highest purity available, betulin (Lup-20(29) ene-3β, 28-diol, 99%); N, O-Bis (trimethylsilyl) trifluoroacetamide (≥98%), pyridine, standard free fatty acids (Supelco® 37 component FAME Mix), and betulin (98%) were supplied by Sigma-Aldrich Química (Barcelona, Spain). Reference standards for fatty acids were used to quantify as methyl testers. Betulin (2.25 × 10-3 M) and KOH (2 M) were prepared in methanol, respectively. Avocado oil was purchased from Laboratorie Soetenaey and Sictia (Fécamp, France).
Cheeses Manufactured and Storage
Cheeses were manufactured according to traditional Manchego type methods. Two types of natural milk are used. One with low fat milk (4.1%), obtained from ewes during three weeks subsequent to the birth of their lambs, to elaborate the Type I cheeses; and the other to elaborate Type II-F and Type III-F with a fat content of 6.5%. Raw ewe milk was filtered through a double membrane piece of porous paper that eliminated any possible remnants, thus avoiding contamination by external germs and the consequent transmission of undesirable flavours. It was then refrigerated at 4°C. Three different ewe cheeses were manufactured from unblended ewe milk in accordance with the following procedure: a commercial starter culture (Lactococcus lactis spp. lactis plus Lactococcus lactis spp. cremoris, from Degussa BioActives Deutschland GmbH & Co.KG, Freising, Germany) was added at the ratio of 2 g per 100 L of milk, and after 35 min at 30°C was cut using a 20-mm knife and stirred at 38°C. The curd was molded and pressed for 2 h and salted by immersion in 18% NaCl solution (pH 5.5) for 8 h at 14°C and stored in a refrigerated chamber at 4°C. Cheeses from every treatment were weighed and chemically analysed.
Two different cheese types were prepared according to the raw milk fat content used. For each sample, two types of cheeses were made: one was analysed fresh, while the other one was stored for 4 months to obtain the typical Manchego-type cheese and then analysed. In a preliminary test, several batches (B) of matured (M) cheeses were made to check the behaviour of the starter preparations with the supplement of the avocado oil. The fatty acids, fat, and cholesterol content results of these preliminary samples were analysed and used in the chemometric classification (shown as supplementary information) where Type I-M-B is made with low fat milk; Type II-M-B is made with full fat milk by means of a mechanical procedure; and Type II-M-B* and Type II-M-B** are elaborated by different mechanical procedures; according to this table, the incorporation of monounsaturated acids are most efficient by mechanical procedure. As reference of them, was made the traditional elaboration (see table in supplementary information for fatty matter, cholesterol, and fatty acids composition [ and ]).
Table 1 Fatty acids composition of the fresh cheese samples (%)
Table 2 Fatty acid composition of the mature (M) cheese samples (%)
After that a different set of cheeses were elaborated to analyze the composition before the ripening, fresh (F), and after 4 months:
| • | Type I-F: Prepared with 4.5 L milk (fat matter content: 4.1%) enriched with 2 g avocado oil/100 mL milk), with a final fat content of 5.9%. After it was made, the final weight was 742 ± 10 g. After ripening it is labeled as Type I-M. | ||||
| • | Type II-F: Prepared with 4.5 L full-fat milk (fat matter content: 6.5%) enriched with 2 g avocado oil/100 mL milk), presented a final fat content of the milk 8.1%. After that the cheese obtained weighed 960 ± 10 g. After ripening it is labeled as Type II-M. | ||||
| • | Type III-F: Finally, a batch of cheese made with full-fat milk (fat matter content of 6.5%) and used as reference, corresponding to commercial cheese. After ripening it is labeled as Type III-M. | ||||
Incorporation of Avocado Oil into Milk Fat-Globules
The implementation of avocado oil from embedded products, such as milk and cheese, required a homogeneous mixture of oil and milk. It was, therefore, necessary that the molecules of oil were incorporated into the fat globules of milk. Thus, optical microscopy was used to monitor the incorporation process of the avocado oil and the optimal concentration to obtain a homogeneous solution. Two methods of incorporation of avocado oil into the fat globule were tested: stirring for 1 min at 1000 rpm and by ultrasounds for 15 min at room temperature. In the case of the stirred procedure, two phases are observed with up to 4% after 4 days. However, for milks under ultrasound and for all the concentrations tested, two phases were observed with a low incorporation of avocado oil. Thus, the process used was mechanical stirring.
Determination of Total Fatty Matter
Respective samples of cheeses (3 g) were heated at 60°C with 10 mL of concentrated HCl (10 M). After that, three extractions with n-hexane were made with 2 × 20 mL and 1 × 10 mL, and when solvent evaporation was carried out, the amount of fatty matter was determined by weight.[Citation18]
Cholesterol Analysis
Five hundred grams of cheese, finely chopped and crushed, were extracted with n-hexane for 10 min at room temperature; after that 5 g of the extract were treated under gentle reflux, with KOH (6.5 g), methanol (40 mL), H2O (10 mL), and betulin (1 mL), used as standard. Previous to cooling at room temperature, H2O (100 mL) was added and extracted with diethyl ether (2 × 50 mL). The resulting organic mixture was washed with H2O (3 × 50 mL) and the organic layer was washed with (20 mL) of brine. It was then dried with Na2SO4 and filtrated; the solvent was removed using a rotary evaporator system. The unsaponifiable residue was dissolved in a 1:1 hexane:ether solution that afforded the corresponding sterols, separated by using a silica column as the mobile phase of 1:1 hexane: ether solution. The sterols fraction obtained were concentrated under vacuum and silanized by 30 μL of N, O-Bis (trimethylsilyl) trifluoroacetamide in 60 μL of pyridine solution for 30 min.
Fatty Acids Analysis
The fatty acids composition was determined after methylation.[Citation18] Cheese extract (0.5 g) was vigorously stirred for 5 min with 0.5 g of KOH/methanol solution and n-hexane (10 mL) and left to stabilize. The organic phase was injected and the fatty acid identification was done by comparing its retention time with the standard mixture. The final concentrations (%) of fatty acids were reported as the percentage of total FFAs. All analyses were performed in duplicate, and the analysis was repeated if necessary.
Apparatus
Analysis was performed using a Clarus 500 instrument (PerkinElmerm, Norwalk, CT, USA), with the use of gas chromatography analyses equipped with a flame ionization detector (GC-FID) and an auto sampler system. Data acquisition and processing were performed using the Soft-link and Total-Chrom Software application (PerkinElmer®'s TotalChem® Chromatography Data Systems [CDS] Software). For cholesterol analysis, 3 μL of standard/sample solution was injected, via the autosampler, to a fused silica capillary column (25 m × 0.25 mm i.d.) coated with 5% phenyl and 95% dimethylpolysiloxane (Varian CP-Sil 8CB, Varian, Dallas, TX, USA). The injector operating conditions were as follows: injector temperature was 260°C, hydrogen carrier gas flow maintained at 1 mL/min. The oven temperature was adjusted to 300°C and the detector was at 310°C. For free fatty acids analysis, 1 μL of standard/sample solution was injected, via the autosampler, to a fused silica capillary column (60 m × 0.25 mm i.d.) coated with 90% polybiscyannopropyl and 10% cyanopropylphenylsiloxane (Supelco SP-2380, Sigma-Aldrich Química, Barcelona, Spain). The injector operating conditions were as follows: injector temperature was 250°C, hydrogen gas flow was maintained at 1 mL/min. The oven and the detector temperature were adjusted to 180 and 260°C, respectively. The pH was measured by an X S 100 Eutech (Thermo Scientific, Surrey, UK). The optical images were recorded by means of a Nikon Microscopy Eclipse E 800, connected to a Nikon DXM1200 digital camera, controlled by a PC using the ACT-1 software (Nikon Corporation, Sendai, Japan).
Multivariate Chemometric Analysis
A chemometric classification of the cheeses was done using firstly a non-supervised approach [non-linear mapping (NLM) and hierarchical clustering analysis (HCA)] followed by a brief supervised analysis (linear discriminate analysis). Non-linear mapping (ALSCSAL) and HCA were used as an unsupervised exploratory technique to detect natural similarities among the fatty acids content of the cheeses. Squared Euclidean distance was always used as the similarity index for clustering, preferably using the average linkage and centroid method. Raw data was standardized based on variable Z-scores. The discriminate capacity power of the variables was assessed with linear discriminate analysis. All the statistical multivariate analysis was performed using the SPSS 16.0 for Windows software (SPSS Inc., Chicago, IL, USA).
RESULTS AND DISCUSSION
Figure 1 shows the optical microscopy images of the different dairy preparations according to the selected procedure of incorporating the avocado oil into the milk. These experiments were carried out using different ranges of oil concentration, from 0–10% (v/v), and left for several days to observe the evolution. and (1 and 2% added avocado oil) show that oil is found on the surface of the fat globules (dark spots). In the case of milks with 4% () and 6% () avocado oil is present on the surface of the fat globule as micro-vesicles. These observations show that cheese preparations with a maximum of 2% avocado oil can be prepared and are analysed in the following sections.
Figure 1 Optic microscopy of mixed avocado oil-milk at (a) 1%, (b) 2%, (c) 4%, and (d) 6%. (Color figure available online.)
Fat Content, Fatty Acids, and Cholesterol Composition in the Fresh Cheeses
Fat content is the most quantitatively and qualitatively variable component of cheeses, depending on the lactation stage, season, breed, genotype, and feeding of the animal. This last important factor has been studied in depth and the main findings concerning the impact of feeding, basic roughage, and lipid supplementation, on quantitative and qualitative variation.[Citation19] The fat content obtained for the fresh cheeses were Type I-F: 28.07%; Type II-F: 33.61%; and Type III-F: 27.21%.
The detailed fatty acids composition of fresh cheese dairy products is shown in . Saturated fatty acids (SFAs) were predominant, 71.04% for Type III-F; 60.94 and 64.63% for Type II-F and Type I-F, respectively. Monounsaturated fatty acids (MUFAs) reached a value of 25.35% for Type III-F; 33.61 and 28.25% for Type II-F and Type I-F, respectively. Palmitic and stearic acids were the major SFAs present in both. Oleic acid and palmitoleic acid were the most abundant MUFAs, where the total content in oleic acid was 23.28% for Type III-F; 30.21 and 23.59% for Type II-F and Type I-F, respectively. Previous studies have reported that the concentrations of unsaturated fatty acids increase and those of saturated fatty acids decrease.[Citation20]
The analysis of polyunsaturated fatty acids (PUFAs) showed a composition of 3.60% for Type III-F; 5.46 and 7.12% for Type II-F and Type I-F, respectively. In the present study, the PUFAs' concentration increased with the content obtained for Type III-F at 51.7 and 97.8% for Type II-F and Type I-F, respectively. The cholesterol contents were measured as 69.36 mg per 100 g of sample for the Type III-F cheese; 56.40 and 63.72 mg per 100 mg of sample for Type II-F and Type I-F, respectively. Therefore, the hand-made fresh cheese with an added 2% of avocado oil represents a good source of PUFAs for humans with lower consumption of other lipids. shows a bar illustration of the results previously discussed.
Figure 2 Total content of different fatty acids, fat, and cholesterol of the fresh cheeses.
Fat Content, Fatty Acids, and Cholesterol Composition of the Mature Cheeses
Figure 3 summarises the global composition of fatty acids, fat content, and cholesterol contents of the previous dairy products, after the typical La Mancha cheese variety had been left in the ripening process for 4 months. The fat content in theses cheeses were 43.0%, an increase of 58% in respect to the fresh Type III-F cheese and 45.3 and 45.0% where the fat increases were of 34.8 and 60.3% for Type II-M and Type I-M, respectively.
The percentage values of the (SFAs) were predominantly 70.56% for Type III-M and 63.72 and 56.09% for Type II-M and Type I-M, respectively, where the most abundant fatty acids were palmitic, stearic, and myristic acids. Monounsaturated fatty acids (MUFAs) reached a value of 25.61% for Type III-M; 31.23 and 37.71% for Type II-M and Type I-M, respectively. Oleic acid corresponds to more than 95% of MUFAs in all cheeses. The analysis of PUFAs afforded a composition of 3.83% for Type III-M; 5.05 and 6.19% for Type II-M and Type I-M, respectively. In the present study, the concentration of PUFAs represents an increase when compared to the value obtained for the Type III-M of 31.85 and 61.62 % for Type II-M and Type I-M, respectively.
The cholesterol contents were measured as 91.54 mg/100 mg of sample for Type III-M; 71.07 and 62.41% for Type II-M and Type I-M, respectively. Therefore, the hand-made Manchego type cheese enriched with 2% (v/v) avocado oil represents a good source of PUFAs for humans with lower consumption of other lipids. shows a bar illustration of the results previously discussed.
Figure 3 Total content of different fatty acids, fat, and cholesterol of the ripened cheese samples.
Multivariate Chemometric Analysis
Figure 4 shows the NLM of the cheese data set. The analysis of this figure shows that the Manchego cheese is quite dissimilar from the other cheeses and it shows the existence of some clusters of similar cheeses. However, a detailed analysis of the NLM should be done with caution because it is a bi-dimensional projection of the real hyperspace graph represented by the 20 fatty acid concentrations. Indeed, the error function (Young's S-stress formula) of the NLM of is 0.04832. Although the NLM suggests some similarities among the fatty acids compositions of the cheeses, a more rigorous analysis of the cluster composition should be done with HCA. shows two dendograms of the cheese data set. Dendogram analysis shows that some cheeses cluster at distances close to zero, and at a rescaled distance of about ten, three clusters and two outlier cheeses (Reference and Type I-M-B) are detected. presents the average and standard deviation of the fatty acids of the detected classes. The borders represented in the NLM of highlights the clusters of similar cheeses and outlier cheeses.
Figure 4 Nonlinear map (Stress = 0.04832 and RSQ = 0.98523) of the cheeses with the classes detected by hierarchical cluster analysis highlighted.
Figure 5 Dendograms obtained using the centroid (a) and average linkage (b) methods of the hierarchical cluster analysis of the cheese data set. 4.0 Reference; 4.0 Type I-M-B; 3.0 Type II-M-B*; 3.0 Type II-M-B**; 2.0 Type III-M; 2.0 Type III-F; 1.0 EC; 1.0 Type I-M; 1.0 Type II-F; 1.0 Type I-F.
Table 3 Average and standard deviation of the fatty acids of the classes of cheese detected by hierarchical cluster analysis
The three clusters detected by NLM and HCA correspond to fresh and mature cheeses with added advocado oil (Type II-M, Type-I-M, Type II-F, and Type I-F), fresh and mature batch chesses (Type II-M and Type III-F), and preliminary cheese preparations with whole milk (Type II-M-B* and Type II-M-B**). The detected outlier, Reference and Type I-F-B, correspond respectively, to the traditional cheese and to a preliminary cheese preparation with skimmed milk enriched with 2% avocado oil. This result shows, as expected, that the preliminary cheese preparations are different from the other cheeses. Also, and as expected, the reference cheeses (Type III-F and Type II-M) are different from the cheeses with 2% avocado oil (Type II-M, Type I-M, Type II-F, and Type I-F).
A detailed analysis of the dendograms of shows that by increasing the similarity, for example to a rescaled distance cluster combination of 5, discrimination between the cheeses with 2% avocado oil is achieved. Indeed, now two sub-clusters are observed (Type II-M and Type I-M) and (Type III-F and Type I-F), and they correspond respectively to cheeses prepared from milk with 6.5 and 4.1% fat content. In order to assess the discriminating capacity of the fatty acids among the three classes that compose the data set, the Wilks' Lambda and F-tests were performed (). The smallest value of Wilks' Lamda and the highest value of the F-tests depict the highest discriminating value of the variable.[Citation21] The fatty acids with the higher discriminating capacity are myristoleic, lauroleic, eicosenoic, caproleic, stearic, palmitoleic, and margaroleic. Consequently, classification strategies based on the fatty acids composition for these cheeses should focus on these substances.
Table 4 Wilks' Lambda and F tests of group means for the linear discriminate analysis of cheese classes
CONCLUSIONS
The results presented and discussed in this study showed that cheeses obtained from ewe milk supplemented with avocado oil at 2% (v/v), contained a smaller percentage of SFAs and a higher percentage of MUFAs and PUFAs, compared with the traditional Manchego type cheeses. Also, these new chesses present lower cholesterol contents. Multivariate analysis of the fatty acid composition in the newly prepared cheeses enriched with 2% avocado oils confirms that they are different from the traditional cheeses and that a small set of fatty acids can be used to develop classification rules. Thus, due to the SFAs, MUFAs, PUFAs, and cholesterol composition, these new cheeses can help to prevent excessive fat intake related diseases. Accordingly, the fat content in comparison with other cheeses type varieties, such as Camembert (average of 48 g/100 g cheese),[Citation22] Cheddar (48 g/100 g cheese),[Citation23] Manchego type cheese (average of 54 g/100 g cheese,[Citation24] or 30 g/100 g cheese,[Citation25] Roncal type (average of 48 g/100 g cheese,[Citation26] and Swiss cheese (30 g/100 g cheese),[Citation27] offer a good alternative to traditional and commercials, where the incorporation of avocado oil does not increase this chemical parameter.
ljfp_a_503358_sup_18087278.zip
Download Zip (9.3 KB)ACKNOWLEDGMENTS
The authors would like to thank Dehesa Los Llanos (Albacete, Spain) for the purchase of the ewe milk, industry facilities for making the cheeses, and Salsa Natura (Málaga, Spain) for funding this project. M. A. would like to thank Fundação para a Ciência e a Tecnologia (Lisboa, Portugal), under the frame of the Ciência 2007 program.
REFERENCES
- Hall , J.H. , Moore , S. , Harper , S.B. and Lynch , J.W. 2009 . Global variability in fruit and vegetable consumption . American Journal Prevention Medicine , 36 ( 5 ) : 402 – 409 . e5
- Day , J. , Jones , D.P. , Goldberg , J. , Ziegler , R. , Bostick , R.M. , Wilson , P.W. , Manatuga , A. K. , Shallenberger , L. , Jones , L. and Vaccarino , V. 2008 . Association between adherence to the Mediterranean diet and oxidative stress . American Journal Clinical Nutrition , 88 ( 5 ) : 1364 – 1370 .
- Hu , F.B. , Manson , J.E. and Willet , W.C. 2001 . Types of dietary fat and risk of coronary heart disease: Critical review. Journal of the American Colleague of Nutrition . 20 ( 1 ) : 5 – 19 .
- Esteves da Silva , J.C.G. 2010 . “ Chemometric classification of cultivars of olives: Perspectives on Portuguese olives ” . In Olives and Olive Oil in Health and Disease Prevention , Edited by: Preedy , V.R. and Watson , R.R. 33 – 42 . Academic Press (Elsevier) . Chap. 4
- Saura-Calixto , F. and Goñi , I. 2006 . Antioxidant capacity of the Spanish Mediterranean diet . Food Chemistry , 94 : 442 – 447 .
- Tennant , D.R. and usage , The . 2004 . occurrence and dietary intakes of white mineral oils and waxes in Europe . Food Chemistry Toxicology , 42 : 481 – 492 .
- Cole , E.T. , Cadé , D. and Benameur , H. 2008 . Challenges and opportunities in the encapsulation of liquid and semi-solid formulations into capsules for oral administration . Advanced Drug Delivery Review , 60 : 747 – 756 .
- Boileau , C. , Martel-Pelletier , J. , Caron , J. , Cheng , S. and Pelletier , J.P. 2009 . Protective effects of total fraction avocado/soybean unsaponifiables (ASU) on the structural changes in experimental dog osteoarthritis: Inhibition of nitric oxide synthase and MMP-13 . Arthritis Research Therapeutic , : R41
- Benaiges , A. and Guillén , P. 2007 . Botanical extracts. Actives for skin-care products. Actives for personal hygiene and other toiletry products. Actives with specific claims . Analysis of Cosmetic Products , : 345 – 363 .
- Moreno , A.O. , Dorantes , L. , Galindez , J. and Guzman , R.I. 2003 . Effect of different extraction methods on fatty acids, volatile compounds, and physical and chemical properties of avocado (Persea Americana Mill.) oil . Journal Agriculture Food Chemistry , 51 : 2216 – 2221 .
- Bergh , B. 1992 . Nutritious value of avocado . California Avocado Society Yearbook , 76 : 123 – 135 .
- Gains , A. 1992 . Avocados will lower your cholesterol. Here is Health . : 36 – 37 .
- Lassen , S. , Bacon , K. and Sutherland , J. 2006 . Vitamin D and pro-vitamin D content of the avocado . Journal Food Science , 10 : 1 – 4 .
- Orozco , P.L. , Zwart , M.S. , Garrofa , E.G. and Domínguez , C.O. 2004 . Prediction of the total calcium intake from consumption of milk products in Spain adult population . INDICAD Study. Atención Primaria , 33 : 237 – 243 .
- Tholstrup , T. , Høy , C.E. , Andersen , L.N. , Christensen , R.D.K. and Sandström , B. 2004 . Does fat in milk, butter and cheese affect blood lipids and cholesterol differently?. Journal of the American College of Nutrition . 23 : 169 – 176 .
- Thompson , L.U. , Jenkins , D.J.A. , Amer , M.A.V. , Reichert , R. , Jenkins , A. and Kamulsky , J. 1982 . The effect of fermented and unfermented milks on serum cholesterol . American Journal Clinical Nutrition , 36 : 1106 – 1111 .
- Cabello , R.M. , Vadillo , J.M. , Casado , J. and Jiménez-Jiménez , J. 2008 . Milk preparation based in avocado oil . Dehesa de los Llanos S.L. Patent ES , 2 : 301 – 365 .
- International Dairy Federation . 2002 . “ Milk fat. Preparation of fatty acid methyl esters ” . In IDF Standard ISO 15884-IDF 182. International Dairy Federation , Belgium : Brussels .
- Sanz-Sampelayo , M.R. , Chilliard , Y. , Schmidely , P. and Boza , J. 2007 . Influence of type of diet on the fat constituents of goat and ewe milk . Small Ruminant Research , 68 : 42 – 63 .
- Martinez , N.L. and Moreno , R.V. 1995 . Parameters for determining the maturity of avocados . Industries. Alimentaires et Agricoles , 112 : 200 – 203 .
- Pinheiro , P.B.M. and Esteves da Silva , J.C.G. 2005 . Detection of 2,4,6-trichloroanisole in chlorinated water at nanogram per litre levels by SPME-GC-ECD . Analytical and Bioanalytical Chemistry , 382 : 341 – 346 .
- Guizani , N. , Kasapis , S. , Al-Attabi , Z.H. and Al-Ruzeiki , M.H. 2002 . Microbiological, physicochemical, and biochemical changes during ripening of camembert cheese made of pasteurized cow's milk . International Journal of Food Properties , 5 : 483 – 494 .
- Ereifej , K.I. 2006 . Characteristics of milk, cheddar-like and white cheese obtained from three goat phenotypes during three lactation periods . International Journal of Food Properties , 9 : 803 – 811 .
- Cabezas , L. , Sánchez , I. , Poveda , J.M. , Seseña , S. and Palop , M.L. 2007 . Comparison of microflora, chemical and sensory characteristics of artisanal Manchego cheeses from two dairies . Food Control , 18 : 11 – 17 .
- Illescas-Chavez , E. and Vélez-Ruiz , J.F. 2009 . Effect of the salting process on the mass transfer kinetics of manchego type cheese. International Journal of Food Properties . 12 : 791 – 807 .
- Taborda , G. , Molina , E. , Martínez-Castro , I. , Ramos , M. and Amigo , L. 2003 . Composition of the water-soluble fraction of different cheeses . Journal Agricultural Food Chemistry , 51 : 270 – 276 .
- Kocaoglu-Vurma , N.A. , Harper , W.J. , Drake , M.A. and Courtney , P.D. 2008 . Microbiological, chemical, and sensory characteristics of Swiss cheese manufactured with adjunct lactobacillus strains using a low cooking temperature . Journal Dairy Science , 91 : 2947 – 2959 .