WO2015186021A1

WO2015186021A1 - Food based food, process for the production and use of the food

Info

Publication number: WO2015186021A1
Application number: PCT/IB2015/053959
Authority: WO
Inventors: Antonino MANTI; Franz SCHWEIGKOFLER
Original assignee: Dolomiti Fruits S.R.L.
Priority date: 2014-06-04
Filing date: 2015-05-27
Publication date: 2015-12-10

Abstract

Fruit based food, including an apple and bergamot juice obtained by sorting, washing, grinding and pressing, under cold conditions, of whole fruits of apple and bergamot, containing flavenoids, brutieridin, melitidin and peripolin, which are mainly in the albedo and flavedo of the bergamot, extracted during the joint pressing (P) of about 10.30% of bergamots and about 90-70% of apples. A yield of 75/78% versus 33/35% with the traditional extraction. Finally, from a waste, potentially pollutant product, a tasty, refreshing and health juice has obtained.

Description

FRUIT BASED FOOD, PROCESS FOR THE PRODUCTION AND USE OF THE FOOD

The invention concerns a fruit based food. In particular, the present invention relates to a composition of a juice obtained from apple and bergamot (citrus bergamia) containing flavonoids, in particular brutieridin, melitidin and peripolin present in the flavedo and albedo of the bergamot having a cholesterol-lowering property.

It is known that the main constituents of apple include hydroxycinnamic acid, dihydrochalcones, flavonols, catechins and procyanidins, as the triterpenoids in the peel and the anthocyanins in red apples. These constituents shows a wide spectrum of beneficial biological activities against cardiovascular disease, asthma and pulmonary dysfunction, diabetes, obesity and cancer.

Bergamot (Citrus bergamia Risso), is a citrus fruit characterized by a unique composition of flavonoids and glycosides present in the juice and flavedo and albedo, as neoeriocitrin, neohesperidin, naringin, rutin, neodesmin, roifolin, poncirin, melitidin, bruteridin, and peripolin. This fruit differs from other citrus fruits not only for its composition in flavonoids but also for the high concentrations.

Among the different flavonoids there is the naringin, a glycosidic derivative of the naringenin, which, in turn, like hesperetin, possesses a similar structure to the substrate of the liver enzyme HMG-CoA reductase, by inhibiting through a statin-like action. The result is a reduction in total cholesterol levels and an inhibition of the oxidation of LDL, as well as a reduction of atherosclerotic plaques. These polyphenols reduce the activity of the enzyme acyl CoA:cholesterol acyltransferase (ACAT), by inhibiting the assembling of lipoproteins. At the same time, it was also demonstrated an improvement in insulin resistance as naringenin leads to a better uptake of blood glucose by the GLUT4 receptors, increasing the activity of AMP kinase (AMPK).

Despite the known properties of bergamot, this fruit, having an unpleasant taste, is mainly used for the production of essential oils extracted from the flavedo and used as a flavoring agent, for example in cologne and in the toilet waters in general. Nowadays, essential oils are extracted mechanically using machines called "peelers", which abrade the outside of the fruit, in particular removing only the essential oil in the flavedo, leaving intact the inner underlying white layer called albedo and the pulp of the fruit contained in the segments. The pulp and the waste in flavedo and albedo, in Italian called "pastazzo" (i.e., fruit pulp waste), is used as a food concentrate for farm animals such as pigs or cattle. The very acid and very bitter juice, for the presence of naringin, obtained by squeezing a possibly mature (yellow) bergamot, because it is slightly less acidic and less bitter, without squeezing the flavedo and albedo, is used sometimes in minimum percentages for the preparation of fruit juices to which it gives a bitter note. The presence of citric acid in the bergamot juice is 66 g/L on average, and for this high acidity is possible to obtain from bergamot juice a natural citric acid, instead, the consumption of the juice causes heartburns compromising the bacterial flora. A few beverages on the market use a small part of bergamot juice, obtained without squeezing the flavedo and albedo, mixed with water, sugar, and various preservatives. It is known from WO 2010/041290 A1 (Sindona, Di Donna, Dolce) a process for the extraction of flavonoids contained in bergamot for pharmaceutical or parapharmaceutical use; said process comprises the use of pure solvents (methanol, ethanol, chloroform), mixed in different proportions and added to the mass of the crushed fruit. The extraction lasts 2 hours, the solvents are subsequently removed by filtration at reduced pressure, leaving a residue which is subjected to a drying process by evaporation. Finally, the mixture of flavonoids obtained is separated by means of a chromatographic system (HPLC).

Said process is relatively complex and expensive, so only justified for obtaining the flavonoids suitable for pharmaceutical use or for the preparation of specific pharmacological products.

The object of the invention is making a fruit based food, preferably a juice contains flavonoids, which are inhibitors of cholesterol present in the bergamot, having an acidity well tolerated by the consumer and an innovative refreshing taste, by applying a simple process without the use of specific solvents and/or the addition of water.

The technical task of the present invention is also achieved by the use of the fruit based food to induce actions and/or metabolic reactions in the body to counteract the formation of cholesterol.

To fulfil this task, the invention proposes the use of the full bergamot fruit (of course, after having removed the essential oil), that is, including flavedo and albedo containing flavonoids and with juicy pulp, and the use of the juice extracted from the apple, having an acidity (about 3-5 g/L on average, expressed as tartaric acid at pH7) significantly lower than that of the bergamot juice, and suitable for:

- making possible the processing of bergamot (pulp + flavedo + albedo) acting as a diluent and as a "vehicle" for the flavonoids extracted from flavedo and albedo;

- effectively covering the bitterness caused by the presence of naringin;

- lowering the overall acidity levels (about 5.6 g/L) well tolerated by consumers;

- sweetening overall the juice to a pleasant level without the addition of sugars or sweeteners.

The production of the juice according to the invention takes place essentially by grinding, in a hammer mill known for the production of fruit juices, a mixture containing from 10 to 30% of bergamot and from 90% to 70% of apples, the milled mixture outgoing from the mill is subsequently cold-pressed in a belt-type fruit press. The juice obtained is optionally added with ascorbic acid (about 200-500 mg/L) in order to avoid oxidation. The juice is collected in a tank (about 1500 L) to be subsequently heated to about 60°C, warm-centrifuged (about 7.000 rpm), and subsequently pasteurized (at about 85°C for about 30 sec.) cooling to about 0-5°C. The practice has shown that this process allows obtaining a juice with a total content of flavonoids equal to about 1 .000 mg/L of which about 540 mg of brutieridin, 425 mg of melitidin and 45 mg of peripolin, with a variation of about 1 -2%, deriving largely from the squeezing of bergamot flavedo and albedo. The process according to the invention allows obtaining a yield in flavonoids, especially bruteridin, melitidin and peripolin, about 10 times compared with the extraction from the pulp alone of the bergamot. Flavonoids then are extracted from flavedo, albedo and juice of bergamot, taking advantage from the presence of the apple juice as a vehicle and as a diluent medium to allow the processing and filtering of the mass after the grinding and squeezing processes, given that said mass, if obtained by squeezing of the bergamot alone, would be a mush quite dense and would not be further processable by conventional machinery used for the production of fruit juices.

With the normal extraction of the juice, with the method used up to now, the yield obtained is the 33/35%, while with the novel process we get a yield of 75/78%. There is therefore a higher yield and a higher concentration of flavonoids.

The juice according to the invention has a taste more bitter than the natural juice of apple alone, and so it is more refreshing than that of pure apple. For the presence of specific flavonoids, the juice inhibits the production of cholesterol, lowers the triglycerides, and with benefits also for blood sugar, and it is also free of added sugar. According to the percentage of bergamot used in the production compared to the amount of apple, and then based on the amount or on the "concentration" of the present flavonoids, the beverage produced can be of the type:

- with low content of flavonoids (less than about 1 .000 mg/L), which is healthy, nutritious, refreshing, and with an innovative and pleasant taste, - with a higher content of flavonoids (about 1 ,000 mg/L), which is healthy healing, without loosing the pleasant taste.

It is known that, to obtain the cholesterol-formation inhibiting effect, it is recommended an intake of about 50 mg per day of specific flavonoids, namely bruteridin, melitidin and peripolin.

The invention is explained more closely on the basis of a diagram of the process of juice production according to the invention, illustrated in the accompanying drawing, for purely illustrative and not limiting purposes. Fig. 1 illustrates a block diagram of the process for producing a juice obtained by squeezing the bergamots and apples comprising the treatment steps following the squeezing.

In accordance with a first preferred embodiment, bergamot is used in full, i.e., with the respective flavedo, (after the extraction of the essential oil), albedo and inner pulp. Before grinding and squeezing of bergamot, the same is micro-drilled at the most superficial region (skin) by special machines of the known type. Such micro-drilling allows to remove and extract the essential oils from bergamot that are not used for the production of the food of the present invention.

In accordance with a further embodiment, only the the essential oil can be removed from the bergamot B (the outer part of the peel) by means of known machines called "peelers" PI; while the fruit (devoid of essential oil) (with flavedo, albedo and pulp) PA is delivered, together with apples M, on a conveyor belt N.

The ratio between the amount of pulp of bergamot, regardless of whether it is full or devoid of flavedo, and the amount of apple M according to the invention can vary between 10 - 30% of bergamot and the balance of 90- 70% of apple. The choice of the ratio depends on the desire to produce a healthy, tasty and refreshing beverage, or a salutary beverage inhibiting the formation of cholesterol in the presence of flavonoids derived mainly from albedo and flavedo of bergamot. In fact, the juice of pulp alone of bergamot contains about 600-800 mg/L, while the juice extracted from albedo and flavedo contains about 4200mg/L. The apple juice, being much less acid than that obtained from bergamot, involves a considerable lowering of the acidity (about 5.6 g/L) mixed in the juice, and such lowering will be more pronounced in the juice containing a higher percentage of apple juice. On said conveyor belt N, the apples M and the bergamot, in full and devoid of essential oil Pa, are subjected to a sorting C, for example on a roller tape to eliminate the waste, any foliages, and foreign bodies Sc. Following the sorting, the fruits undergo a washing step with fresh drinking water L at high pressure, and subsequent a drying step with air at room temperature, moved by a fan, and then the grinding Ma is performed in a hammer mill. The milled fruits go into a container, which acts as a pneumatic reservoir, where they are moved to a belt press P, the juice from the pulp and albedo of bergamot and from apples is collected within a tank S (for example, 1 .500 L), where ascorbic acid is added A, if necessary, in amounts of about 200-500 mg/L in order to prevent the oxidation of the juice, while the pomace Sa is eliminated to be used as animal feed. The addition of ascorbic acid at the indicated amounts results in a negligible increase in acidity of this juice produced. Subsequently, the juice is subjected to a filtering step through a stainless steel cartridge with openings of 1 mm, a heating to about 60°C, a warm centrifugation D at about 7,000 rpm in order to avoid the formation of deposits, and a pasteurization Ps to about 85°C for about 30 sec. with subsequent cooling to about 0-5°C to be moved to storage St in aseptic tanks from which it may be collected for packaging or for transport to packaging companies. In accordance with a further embodiment, the mixed mixture of apple and bergamot juice is produced by addition of the juice obtained, via a separate process, from the pulp and albedo and from flavedo of bergamot to the separately produced juice from apples. In this case, the bergamot PA (pulp and albedo) as well as the apples M separately undergo the same treatment, comprising the pressing on separate presses P by combining the juice exiting from the presses, according to the desired ratio, in a single tank S, where ascorbic acid A is added, if desired, and then the mixed juice mixture is subjected to the same processes described above.

A further preferential embodiment comprises the processing of bergamot PA (pulp, albedo and flavedo) and apples M separately until the end of the process, and the two centrifuged and separately pasteurized juices are mixed in storage tanks or just during packaging.

Shown below are the results of a prospective experimental study conducted in healthy volunteers of both sexes and aged between 18 and 65 years. The study had the aim to evaluate the effects of the uptake of 250 imL/day of the fruit based food according to the present invention for a period of 15 days on body composition, fat profile, glucose, inflammatory and oxidative stress parameters, as part of a balanced diet and a healthy lifestyle.

Among the inclusion criteria there were subjects of both sexes and aged between 18 and 65 years; 19 <BMI <25 kg/m2.

Among the exclusion criteria there were endocrine, metabolic, hepatic, renal, inflammatory, cardiovascular, chronic viral diseases, HIV, cancer; steroid therapy, anti-inflammatory therapy, antibiotic therapy.

STUDY PHASES:

At TO (baseline), at time T1 (Acute - after 2 hours of taking 250 imL of berime juice), and at time T2 (Chronic - after 15 days of taking the food) the blood was collected and a full evaluation of the nutritional status of each enrolled subject was performed:

• Far and/o near in time family and individual history on physiological and/or pathological conditions.

· Evaluation of eating habits by completing two different validated questionnaires.

• Assessing body composition by means of:

- Anthropometric measurements (weight, height, calculation of BMI kg/m²)

- Measurements of waist and flank circumference and their ratio

- Assessment of water body by BIA (bioelectrical impedance): determination of the TWB (total body water), intracellular water (ICW) and water extracellular (ECW), determination of the metabolically active cell mass and its corresponding index (BCM, BCMI), determination of the Na/k exchange, and the phase angle as an indicator of the nutritional status · Functional assessment: Blood pressure and heart rate measurements • Handgrip: evaluation test for contractile force of the flexor muscles of the hand used to assess the nutritional status, as an indicator of overall muscle strength, as a prognostic factor in the course of metabolic diseases, as well as a tool to discover a sarcopenia.

· Assessment of biochemical parameters:

^* CBC with formula

^* Transaminase (GOT, GPT)

^* Gycemia (mg/dl)

^* Insulin (U/mL)

* ESR (mm)

^* C-reactive protein (img/dL)

^* Triglycerides (img/dL)

^* HDL cholesterol (mg/dL)

^* LDL cholesterol (mg/dL)

* Total cholesterol (mg/dL)

^* Determination of oxidized LDL (ng/mL) (ELISA, Enzyme-Linked Immunosorbent Assay)

RESULTS

The results shown are for the evaluation of nutritional status in healthy volunteers at the three times of the study, about the data 1 ) of body composition, obtained by anthropoplicometric and impedenziometric methods; 2) of blood chemistry of the lipid and glucose profile; 3) of biochemical analysis of oxidative stress, assessed by enzyme immunoassay (ELISA) for the oxidation levels of low density lipoproteins (LDL). Table 1 shows the comparison of the values of body composition at enrolment (TO) and after 15 days of administration of the food product

(T2).

Table 1

15 days

Baseline (TO) product (T2)

Mean (SD) Mean (SD)

Systolic blood 1 10.91 (13.04) 1 10.00 (10.95)

pressure³ 0.78

Diastolic blood

pressure³ 69.09 (10.75) 69.36 (7.58) 0.90

Weight¹³ 60.74 (10,61 ) 60,21 (10,63) 0.1 1 -0.88

BMI^b 21 .77 (2,23) 21 ,83 (2,25) 0.1 1

Circumf. Waist³ 69.98 (7,01 ) 70.02 (7,18) 0.96

Circumf. Flanks³ 94.19 (5,14) 94.41 (4,30) 0.74

Waist/Flanks ratio^b 0.74 (0,04) 0.74 (0,05) 0.46

FFM^b 47.86 (8.87) 47.13 (9,32) 0.33

TBW^b 34.96 (6,58) 34.48 (6,85) 0.15

ECW³ 15.86 (2,92) 16.07 (3,16) 0.39

BCM^b 25.45 (5,49) 24.34 (5,63) 0.93

FM³ 13.03 (3,65) 13.01 (3,19) 0.97

BP³ 5.81 (0,73) 5.83 (0,69) 0.83

Na/K³ 1 .09 (0,12) 1 .10 (0,10) 0.76

MM^b 26.35 (5,87) 30.67 (6,64) 0.00^* 16.40 1488,23 1471,28

BMRkcal^b (159,25) (160,00) 0.72

BCMI^a 8,85 (1,29) 9,01 (1,32) 0.44

Biceps skinfold¹³ 6.47 (4.14) 4,56 (2,36) 0.06 -23.20

Triceps skinfold" 11.62 (8.22) 10,17 (8,55) 0.23 -12.30

Subscapular

skinfold¹³ 13.27 (5.45) 13,67 (5,84) 0.40

Over-iliac skinfold³ 11.45 (4.67) 12,23 (4,72) 0.51

* Significant difference. ^a t-test. ^b Test Wilcoxon

The results of the bio-impedenziometric assay show values of lean body mass, after administration of the food according to the invention, significantly increased by 16.40%.

It is clear from the values obtained that, after administration of the food, there was a reduction of the subcutaneous triceps (-12.30%) and biceps (-23.20%) fat skinfold.

The average weight of the enrolled subjects after administration of the product is reduced slightly (-0.88%).

Table 2 shows the comparison between the blood chemistry parameters at enrolment (TO) and after 2 hours from the administration of the food product (T1).

Table 2

Baseline (TO) After the

product (T1) P Δ%

MEAN ±SD

Total cholesterol³ 170.18129.35 172.45131.85 0.58

HDL(mgdL)^a 63.55 ± 11.39 63.82 ±11.40 0.70 0.43 LDL(mgdL)^a 94.09 ± 20.08 92.64 ± 19.86 0.29 -3.33

TOTCol/HDL^b 2.71 ±0.35 2.71 ±0.41 0.92

TG(mgdL)^b 60.67±27.46 56.36±21.40 0.05^* -10.92

LDL/HDL³ 1.53 ± 0.36 1.48 ± 0.37 0.07

TOTCol/LDL³ 1.87 ± 0.29 1.91 ± 0.25 0.70

TGL/HDL³ 0.93 ± 0.60 0.84 ± 0.35 0.59 -18.71

GLYCEMIA(mgdL)^§ 78.91 ± 9.04 76.27 ± 5.42 0.21 -3.34

INSULIN(U/mL)^a 5.57 ± 1.31 5.49 ± 1.30 0.85 -1.47

HOMA³ 1.09 ± 0.29 1.04 ± 0.28 0.61 -4.57

Significant difference. ^a t-test. ^b Wilcoxon test

After 2 hours of taking the food, the data shows: a) A significant reduction in triglycerides (TG) by 10% and a positive increase of high-density lipoprotein (HDL) by 0.43%,

b) A reduction by 18.71 % in the TG/HDL ratio c) A positive reduction of the low-density lipoproteins (LDL) by 3.33%.

d) A reduction in blood glucose of 3.34%, a reduction of insulin by 1 .47%, resulting in a reduction of the insulin resistance index (HOMA-IR) by 4.57%.

Table 3 shows the comparison between the blood chemistry parameters at enrolment (TO) and after 15 days of administration of the food product 250 mL/day (T2). Table 3

Basel ine(TO) After the

product (T2)

MEANiSD MEANiSD

TotCol^a 169.45±29.65 162.91 ±25.27 0.20 -3.86 HDL(mg/dL)^a 63.36±1 1.62 65.27±10.36 0.39 3.01

LDL(mg/dL)^a 94.50±19.98 89.00±18.79 0.20 -4.49

TOTCol/HDL^b 2.74±9.35 2.14±2.73 0,00^* -20.91 LDL/HDL³ 1 .53±0.35 1 .40±0.30 0.08 -7.56 TOTCol/LDL³ 1 .80±0.20 1 .84±0.12 0.38

GLYCEMIA(mg/dL)^a 77.45±7.94 78.18±8.34 0.73

INSULIN(U/mL)^a 5.70±1 .53 6.23±1 .62 0.36

HOMA³ 1 .08±0.29 1 .20±0.32 0.34

* Significant difference. ^a t-test. ^b Wilcoxon test

After 15 days of daily intake of 250ml of the food, the data shows:

a) A significant reduction in the total cholesterol/high density lipoproteins

(HDL) ratio (TotCol/HDL) by 20% with a positive, although not

significant, increase in HDL by 3%, resulting in reduction of

cardiovascular risk.

b) A reduction of low density lipoproteins (LDL) by 4.49%.

c) A reduction in total cholesterol by 3.86%.

d) A reduction of the LDL/HDL ratio by 7.56%.

e) Reductions of the carbohydrate profile parameters (glucose, insulin

and insulin resistance index) are not confirmed.

The diagram 1 represents the comparison between the oxidation levels of

low density lipoproteins (Ox-LDL) at enrolment (TO) and after intake of the

food product 250 imL/day at the two analysis times (T1 , T2).

Percentage change (Δ%)

After fifteen days of daily consumption of 250 mL of the food, we have observed a statistically significant reduction in the cardiovascular risk index, given by the total cholesterol/high density lipoproteins (HDL) ratio (TotColl/HDL) by 21 %. After 2 hours from the assumption of 250 mL of the food product, we have observed a significant reduction in triglycerides (1 1 %, p <0.05). There was not any other statistically significant change to all other analysed bioclinical and body composition parameters (total cholesterol, LDL cholesterol, oxidized LDL, glucose, insulin, pressure, body weight, body circumference, body skinfold, water compartments).

Claims

1 . Fruit based food, comprising 10 to 30% of bergamot and 90 to 70% of apples;

characterized in that said bergamot is composed of flavedo, albedo and endocarp containing flavonoids, brutieridin, melitidin and peripolina.

2. A fruit based food according to the previous claim, characterized in that said bergamot further comprises the flavedo.

3. Product according to claim 1 , characterized in that it comprises an acidity of about 5.6 g/L of the product, which is significantly lower respect to the acidity (about 66 mg/L) of the juice obtained from the bergamot alone.

4. Product according to claim 1 , characterized in that it comprises from 20 to 30% of bergamot and from 80 to 70% of apples.

5. Product according to claim 3, characterized in that it comprises from 21 to 30% of bergamot and from 79 to 70% of apples.

6. Product according to any one of the preceding claims, characterized in that said said bergamot and said apple are in liquid form, obtained by the squeezing thereof, to obtain a base juice of apple and bergamot.

7. Use of a food product in accordance with one or more of the preceding claims, for inducing in an organism actions and/or metabolic reactions to counteract and/or slow down the formation of cholesterol.

8. Use according to the preceding claim, to reduce the cardiovascular risk index, given by the total cholesterol/high density lipoprotein (HDL) ratio (TotColl/HDL).

9. Use according to claim 7 or 8, to reduce the percentage value of triglycerides (1 1 %, p <0.05).

10. Process for the production of the food in accordance with any one of claims 1 to 6, comprising the steps of sorting (C), washing and drying (L), grinding (But) and pressing (P) the fruits, and subsequently adding of ascorbic acid (A) to the resulting juice, centrifuging (D) and pasteurisation (P) of the juice, characterized in that both the fruits undergo jointly a sorting (C) with removal of leaves and foreign bodies by passing it on tape roller, a washing with drinking water at high pressure, an air-drying (L) at ambient temperature, a grinding (But) through hammer mill, a pressing through a belt press, a possible addition of ascorbic acid (A), a filtering with a stainless steel cartridge with openings of about 1 mm, a heating at about 60°C, a centrifugation (D) at about 7.000 rpm, a pasteurization (P) at about 85°C for about 30 sec. with cooling to about 0-5°C, with subsequent storage in aseptic tanks.

1 1 . Process according to the preceding claim, characterized in that all the processes on the pulp with albedo (PA) of the bergamot and of the apples are performed simultaneously on both the fruits passing simultaneously into a single plant.

12. Process according to the preceding claim, characterized in that the two fruits (PA, M) are processed (N, C, L, M) and pressed (P) separately, the juices obtained separately, downstream of the pressing (P), are mixed in the ratio 10 to 30% of bergamot and 90 to 70% of apple, and processed later (D, P) at a mixed state.

13. Process according to claim 1 1 , characterized in that the two fruits (PA, M) are processed (N, C, L, M) and pressed (P) separately, and also the juices obtained separately, downstream of the pressing (P), are processed and treated (D, P) separately performing the mixing in the ratio 10 to 30% of bergamot juice and 90 to 70% of apple juice during or upstream of the packaging.

14. Process according to any one of claims 10 to 13, wherein the bergamot is ground and pressed in full, with the respective flavedo.

15. Process according to the preceding claim, characterized in that it further comprises the step of extraction of essential oils from the flavedo of bergamot; this extraction being implemented through micro-drilling and oil extraction.