FIELD OF THE DISCLOSURE

The present disclosure relates to a process for manufacturing Nata-de-coco.

BACKGROUND

Nata-de-Coco is a chewy translucent, jelly-like traditional food product produced by bacterial fermentation of coconut milk and coconut water or both. It is served in sweetened form in ice creams, halo-halo and other fruit preparations like cocktails and salads. Nata-de-coco is native to Philippines and produced by using Gluconoacetobacter xylinum bacterium. The production of Nata-de-Coco is a cottage scale industry in coconut growing nations of South-east Asia and is produced in the small scale by rural households. Despite its name, Nata-de-Coco is not a cream, but rather a clear solid.
Traditionally, Nata-de-coco is produced by breaking a mature coconut to expose the meat. The coconut meat is either grated or directly pressed to extract coconut milk. A little quantity of water is added during pressing so as to ensure better extraction. To the coconut cream a medium containing the Gluconoacetobacter species is added and left for about a week under room conditions for fermentation. After fermentation a Nata-de-coco layer is formed. Nata-de-coco formed after fermentation is strained, rinsed, and pressed into a mass and cut into cubes.

Nata-de-Coco is enjoyed as a dessert and as a garnish to many regional Asian dishes.

Nata de coco has high fiber content and near-zero cholesterol count and hence is nutritional to health.

Japanese Patent No. 61149055 discloses a method to produce Nata de coco by i) inoculating acetic bacteria to a mixture of coconut fruit juice and a sugar; ii) culturing the inoculated mixture in acidic state to obtain Nata-de-coco; and iii) removing the acid component from the produced Nata-de-coco.

WO 2000/023516 discloses a process for the production of an extruded cellulose product (Nata-de-coco); the process comprising following steps: i) forming a cellulose solution in an amine-oxide solvent containing non-bacterial cellulose and bacterial cellulose in an amount of 0.01 to 5 % by weight of the total cellulose weight; and ii) extruding the cellulose solution to form an extruded cellulose product.

The prior art disclosures use processes having relative low yield for the production of Nata-de-coco. Accordingly, it is desirable to develop a cost-efficient and high yielding process for manufacturing Nata-de-coco.

OBJECTS

Some of the objects of the present disclosure, which at least one embodiment herein satisfies, are as follows:

It is an object of the present disclosure to provide a process for manufacturing Nata-de-coco.

It is another object of the present disclosure to provide a process using microbial consortium for manufacturing Nata-de-coco.

It is still another object of the present disclosure to provide a process using nutrients for manufacturing Nata-de-coco.

It is still another object of the present disclosure to provide a cost-efficient and high yielding process for manufacturing Nata-de-coco.

Other objects and advantages of the present invention will be more apparent from the following description when read in conjunction with the accompanying figures, which are not intended to limit the scope of the present invention.

SUMMARY

These and other objects of the present disclosure are to a great extent dealt in the disclosure herein after.

In one aspect of the present disclosure there is provided a process for manufacturing Nata-de-coco, said process comprising the following steps:

i) treating a coconut substance with at least one nutrient;

ii) inoculating the treated coconut substance with a microbial consortium containing Gluconoacetobacter Xylinum and yeast to obtain inoculated coconut substance;

iii) fermenting the inoculated coconut substance to produce Nata-de-coco; and

iv) harvesting Nata-de-coco therefrom.

Typically, the coconut substance is at least one selected from the group consisting of coconut meat, grated coconut meat, coconut water and coconut milk.

Typically, the nutrient is at least one selected from the group consisting of sugar carbon sources, organic acid carbon sources, alcohol carbon sources, inorganic nitrogen sources, organic nitrogen sources, vitamins and enrichment components.

Typically, the sugar carbon source as a nutrient is at least one selected from the group consisting of fructose, glucose, sucrose and lactose.

Typically, the proportion of sugar carbon source as a nutrient ranges between 1.0% and 15.0 % with respect to the total mass of the inoculated coconut substance.

Preferably, the sugar carbon source as a nutrient is sucrose in an amount 1.0 to 10.0 % with respect to the total mass of the inoculated coconut substance.

Typically, the organic acid carbon source as a nutrient is at least one selected from the group consisting of acetic acid, succinic acid, lactic acid and gluconic acid.

Typically, the proportion of organic acid carbon source as a nutrient ranges between 1.0 % and 5.0 % with respect to the total mass of the inoculated coconut substance.

Preferably, the organic acid carbon source as a nutrient is acetic acid in an amount 1.0 to 2.0 % with respect to the total mass of the inoculated coconut substance.

Typically, the alcohol carbon source as a nutrient is at least one selected from the group consisting of glycerol and ethanol.

Typically, the proportion of the alcohol carbon source as a nutrient ranges between 1.0 % and 5.0 % with respect to the total mass of the inoculated coconut substance.

Typically, the inorganic nitrogen source as a nutrient is at least one selected from the group consisting of ammonium sulphate, potassium nitrate and ammonium phosphate.

Typically, the proportion of inorganic nitrogen source as a nutrient ranges between 0.1 % and 2.0 % with respect to the total mass of the inoculated coconut substance.

Typically, the organic nitrogen source as a nutrient is at least one selected from the group consisting of tryptone, peptone and yeast extract.

Preferably, the organic nitrogen source as a nutrient is a combination of yeast extract and peptone.

Typically, the proportion of organic nitrogen source as a nutrient ranges between 0.1 % and 2.0 % with respect to the total mass of the inoculated coconut substance.

Preferably, the organic nitrogen source as a nutrient is in an amount ranging between 0.1 and 1.0 % with respect to the total mass of the inoculated coconut substance.

Typically, the vitamin as a nutrient is at least one selected from the group consisting of thiamine mononitrate, vitamin B2, vitamin B12, nicotinamide, calcium pentothenate, folic acid, vitamin B6 and vitamin C.

Typically, the proportion of vitamin as a nutrient ranges between 0.01 % and 0.05 % with respect to the total mass of the inoculated coconut substance.

Typically, the enrichment component is pineapple juice, and the proportion of enrichment component ranges between 1.0 % and 5.0 % with respect to the total mass of the inoculated coconut substance.

Typically, the proportion of microbial consortium containing Gluconoacetobacter Xylinum and yeast ranges between 5 % and 20 % with respect to the total mass of the inoculated coconut substance.

Preferably, the proportion of Gluconoacetobacter Xylinum: yeast used for inoculation is 1:1.

Typically, the method step of fermenting is carried out at a temperature ranging between 20 °C and 40 °C for a time period ranging between 6 days and 15 days.

Typically, the method step of fermenting is carried out at a pH ranging between 3.5 and 5.5. Preferably, the pH is 4.0 and 5.0.

Typically, the method step of harvesting is carried out by separating Nata-de-coco from the fermented coconut substance followed by pressing and cutting into cubes.

Typically, the method step of harvesting further comprises the following steps;

i) boiling Nata-de-coco cubes in water at least once to obtain Nata-de-coco cubes with a predetermined pH; and

ii) poaching Nata-de-coco cubes in sugar solution followed by packaging under aseptic conditions.

DETAILED DESCRIPTION OF THE DISCLOSURE

The embodiments herein and the various features and advantageous details thereof are explained with reference to the non-limiting embodiments in the following description. Descriptions of well-known components and processing techniques are omitted so as to not unnecessarily obscure the embodiments herein. The examples used herein are intended merely to facilitate an understanding of ways in which the embodiments herein may be practiced and to further enable those of skill in the art to practice the embodiments herein. Accordingly, the examples should not be construed as limiting the
scope of the embodiments herein.

The description herein after, of the specific embodiments will so fully reveal the general nature of the embodiments herein that others can, by applying current knowledge, readily modify and/or adapt for various applications such specific embodiments without departing from the generic concept, and, therefore, such adaptations and modifications should and are intended to be comprehended within the meaning and range of equivalents of the disclosed embodiments. It is to be understood that the phraseology or terminology employed herein is for the purpose of description and not of limitation. Therefore, while the embodiments herein have been described in terms of preferred embodiments, those skilled in the art will recognize that the embodiments herein can be practiced with modification within the spirit and scope of the embodiments as described herein.

The processes for the preparation of Nata-de-coco disclosed in the prior art documents suffer with various drawbacks such as these processes produce relatively low yield of Nata-de-coco. In view of this, the inventor of the present disclosure envisaged a simple process which results in increased production of Nata-de-coco.

In order to improve the yield of Nata-de-coco, the inventor of the present disclosure tried incorporation of different types of nutrients such as sugar carbon sources, organic acid carbon sources, alcohol carbon sources, inorganic nitrogen sources, organic nitrogen
sources, vitamins, enrichment components into the nutrient broth. The inventor found that the presence of nutrients in specific proportions enhances the yield of Nata-de-coco.

The inventor also found that the yield of Nata-de-coco is further enhanced when a microbial consortium (combination of bacterium and yeast) is employed during manufacturing of Nata-de-coco.

The process of the present disclosure is described herein below:

In the first step, coconut substance is selected from various sources such as coconut meat, grated coconut meat, coconut water and coconut milk. The selected coconut medium is sterilized at 121°C and 15 lb/inch2 for 15 minutes and is treated with at least one nutrient.

The nutrient used for treating the coconut substance includes but is not limited to sugar carbon sources, organic acid carbon sources, alcohol carbon sources, inorganic nitrogen sources, organic nitrogen sources, vitamins and enrichment components (pineapple juice). The sugar carbon source which is used as a nutrient is selected from the group consisting of fructose, glucose, sucrose and lactose. The proportion of sugar carbon source as a nutrient ranges between 1.0 % and 15.0 % with respect to the total mass of the inoculated coconut substance. The preferred sugar carbon source as a nutrient is sucrose and it is used in a proportion ranging between 1.0 and 10.0 % with respect to the total mass of the inoculated coconut substance. Alternatively, the organic acid carbon source is used as a nutrient which is selected from the group consisting of acetic acid, succinic acid, lactic acid and gluconic acid. The proportion of organic acid carbon source used ranges between 1.0 % and 5.0 % with respect to the total mass of the inoculated coconut substance. Acetic acid is preferred amongst various organic acid carbon sources. It is used in an amount of about 1.0 to 2.0 % with respect to the total mass of the inoculated coconut substance. Further, alcohol carbon source such as glycerol and ethanol can be used as a nutrient. The proportion of alcohol carbon source used ranges between 1.0 % and 5.0 % with respect to the total mass of the inoculated coconut substance. Still further, the inventor of the present disclosure used inorganic nitrogen source as a nutrient. The
inorganic nitrogen source includes ammonium sulphate, potassium nitrate and ammonium phosphate. The amount of inorganic nitrogen source used is about 0.1 % and 2.0 % with respect to the total mass of the inoculated coconut substance. Furthermore, organic nitrogen source such as tryptone, peptone and yeast extract in the amount ranging between 0.1 % and 2.0 % with respect to the total mass of the inoculated coconut substance is used as a nutrient. The combination of yeast extract and peptone is a preferred organic nitrogen source. The preferred amount of organic nitrogen is ranging between 0.1 and 1.0 % with respect to the total mass of the inoculated coconut substance.

Apart from these nutrients, the inventor also tried various vitamins as a source of nutrients such as thiamine mononitrate, vitamin B2, vitamin Bl2, nicotinamide, calcium pentothenate, folic acid, vitamin B6 and vitamin C. The vitamins are used in an amount of about 0.01 % and 0.05 % with respect to the total mass of the inoculated coconut substance. The inventor also found that the enrichment component such as pineapple juice can be used as nutrient in the proportion of about 1.0 % and 5.0 % with respect to the total mass of the inoculated coconut substance.

In second step, the treated coconut substance is inoculated with a microbial consortium containing Gluconoacetobacter Xylinum and yeast to obtain inoculated coconut substance. The proportion of microbial consortium containing Gluconoacetobacter Xylinum and yeast ranges between 5 % and 20 % with respect to the total mass of the inoculated coconut substance. In the microbial consortium, 1:1 proportion of Gluconoacetobacter Xylinum: yeast is maintained.

In third step, the inoculated coconut substance is fermented at a temperature ranging between 20°C and 40°C for a time period ranging between 6 days and 15 days to produce Nata-de-coco. The fermentation is carried out at a pH of about 3.5 to about 5.5. Preferably, the fermentation is carried out at a pH of about 4.0 and about 5.0.

Furthermore, the fermentation is carried out at a surface area ranging between 1cm x 13 cm and 25 cm x 2.5 cm. The preferred surface area is 25 cm x 2.5 cm.

Finally, the Nata-de-coco is harvested by separating Nata-de-coco from the fermented coconut substance followed by pressing and cutting into cubes. Then, the Nata-de-coco cubes are boiled in water at least once to obtain Nata-de-coco cubes with a predetermined pH and then poached in sugar solution followed by packaging under aseptic conditions.

The present disclosure is further described in light of the following examples which are set forth for illustration purpose only and not to be construed for limiting the scope of the disclosure.

Example I: A process for manufacturing Nata-de-coco using Gluconoacetobacter xylinum (without nutrient and yeast)

1 kg of freshly grated coconut was mixed with 10 to 30 liter of water (coconut water and/or tap water) and strained through cheese cloth to obtain coconut water/milk broth.

The coconut water broth was boiled at a temperature 121°C and allowed to cool. The cooled coconut water/milk broth was then inoculated with Gluconoacetobacter xylinum at a proportion ranging between 5 % and 20 % with respect to the total mass of the inoculated coconut substance. The inoculated coconut substance was then poured into a vessel. The vessel top was covered to allow inoculated coconut substance to ferment at 3.5 to 5.5 pH and temperature ranging between 20 °C and 40 °C for a time period ranging between 6 days and 15 days. Nata-de-coco was produced in the vessel which was harvested therefrom and washed.

To make Nata-de-coco edible, the harvested Nata-de-coco was cut into cubes which were subjected to a series of boiling with fresh water to remove acidity. The cubes were then poached in sugar solution and mixed thoroughly. The cubes were boiled in sugar solution in the ratio of 1:1 till the cubes became transparent to obtain edible Nata-de-coco which was packed in glass jars or pouches, sterilized and sealed.

The yield is 16.40%.

Example II: A process for manufacturing Nata-de-coco using Gluconoacetobacter xylinum and nutrient (without yeast).

1 kg of freshly grated coconut was mixed with 10 to 30 liter of water (coconut water and/or tap water) and strained through cheese cloth to obtain coconut water/milk. To the obtained coconut water/milk, nutrient was added;

wherein the nutrient is at least one selected from the group consisting of sugar carbon sources such as fructose, glucose, sucrose and lactose (1.0 % to 15.0 % with respect to the total mass of the inoculated coconut substance), organic acid carbon sources such as acetic acid, succinic acid, lactic acid and gluconic acid (1.0 % to 5.0 % with respect to the total mass of the inoculated coconut substance), alcohol carbon sources such as glycerol and ethanol (1.0 % to 5.0 % with respect to the total mass of the inoculated coconut substance), inorganic nitrogen sources such as ammonium sulphate, potassium nitrate and ammonium phosphate (0.1 % and 2.0 % with respect to the total mass of the inoculated coconut substance), organic nitrogen sources such as tryptone, peptone and yeast extract (0.1 % and 2.0 % with respect to the total mass of the inoculated coconut substance), vitamins such as thiamine mononitrate, vitamin B2, vitamin B^, nicotinamide, calcium pentothenate, folic acid, vitamin B6 and vitamin C (0.01 % to 0.05 % with respect to the total mass of the inoculated coconut substance) and enrichment component such as pineapple juice (1.0 % to 5.0 % with respect to the total mass of the inoculated coconut substance) to obtain a coconut water broth.

The coconut water broth was boiled at a temperature 121°C and allowed to cool. The cooled coconut water broth was then inoculated with microbial consortium containing Gluconoacetobacter xylinum at a proportion ranging between 5 % and 20 % with respect to the total mass of the inoculated coconut substance. The inoculated coconut substance was then poured into a big mouthed vessel having large surface area and ample space at top of the inoculated coconut substance. The vessel top was covered to allow inoculated coconut substance to ferment at 3.5 to 5.5 pH and temperature ranging between 20 °C

and 40 °C for a time period ranging between 6 days and 15 days. Nata-de-coco was produced in the vessel which was harvested therefrom and washed.

To make Nata-de-coco edible, the harvested Nata-de-coco was cut into cubes which were subjected to a series of boiling with fresh water to remove acidity. The cubes were then poached in sugar solution and mixed thoroughly. The cubes were boiled in sugar solution in the ratio of 1:1 till the cubes became transparent to obtain edible Nata-de-coco which was packed in glass jars or pouches, sterilized and sealed.

Example 1: Effect of carbon source on Nata-de-coco yield.

Example 1.1: Effect of sugar carbon source on Nata-de-coco yield

The process of Example II was followed except that the nutrient is sugar and fermentation is carried out for a time period of 16 days at 28 °C.

The results are provided in the Table No. 1.1:

Table 1.1: Effect of sugar carbon source on Nata-de-coco yield

Example 1.2: Effect of organic acid carbon source on Nata-de-coco yield.

The process of Example II was followed except that the nutrient is organic acid and fermentation is carried out for a time period of 16 days at 28 °C.

The results are provided in the Table No. 1.2:

Table 1.2: Effect of organic acid carbon source on Nata-de-coco yield

Example 1.3: Effect of alcohol carbon source on Nata-de-coco yield.

The process of Example II was followed except that the nutrient is alcohol and fermentation is carried out for a time period of 16 days at 25 °C to 30 °C.

The results are provided in the Table No. 1.3:

Table 1.3: Effect of alcohol carbon source on Nata-de-coco yield

Conclusion:

1. Among the different sugars used as a carbon source, sucrose yielded maximum
Nata-de-coco. It was found that increase in the concentration of sucrose beyond
10% is not economical for manufacturing Nata-de-coco.

2. The texture of Nata-de-coco varied from soft to tough depending on type of sugar
carbon source in the inoculated coconut substance.

3. Among different organic acids used as carbon source, acetic acid and lactic acid
were found to give high Nata-de-coco yield. However, acetic acid preferred over
lactic acid because of availability and cost.

4. Among different alcohols as carbon source, glycerol (1%) and ethanol (1%) served as effective carbon sources and yielded maximum Nata-de-coco.

Example 2: Effect of nitrogen source on Nata-de-coco yield.

Example 2.1: Effect of inorganic nitrogen source on Nata-de-coco yield

The process of Example II was followed except that the nutrient is inorganic nitrogen source and fermentation is carried out for a time period of 16 days at 25 °C to 30 °C.

The results are provided in the Table No. 2.1:

Table 2.1: Effect of inorganic nitrogen source on Nata-de-coco yield

Example 2.2: Effect of organic nitrogen source on Nata-de-coco yield

The process of Example II was followed except that the nutrient is organic nitrogen source and fermentation is carried out for a time period of 16 days at 25 °C to 30 °C.

The results are provided in the Table No.:

Table 2.2: Effect of organic nitrogen source on Nata-de-coco yield

Conclusion:

Organic source of nitrogen was found to be effective as compared to inorganic source in the production of Nata-de-coco. However, inorganic source can be preferred over organic due to its availability and handling.

Example 3: Effect of vitamin concentration on Nata-de-coco yield

The process of Example II was followed except that the nutrient is vitamin stock solution and fermentation is carried out for a time period of 16 days at 25 °C to 30 °C.
100 ml vitamin stock solution was prepared using following vitamins:
10.0 mg of thiamine mononitrate, 100.0 mg of vitamin B2, 3.0 mg of vitamin B]2, 100.0 mg of nicotinamide, 50 mg of calcium pentothenate, 1500.0 ^g of folic acid, 15 ug of vitamin B6 and 150 mg of vitamin C.

The results are provided in the Table No. 3:

Table 3: Effect of vitamin concentration on Nata-de-coco yield

Conclusion:

There was no significant increase in the yield of Nata-de-coco when vitamins were added as a nutrient.

Example 4: Effect of enrichment component on Nata-de-coco yield

The process of Example II was followed except that the nutrient is pineapple juice as enrichment component and fermentation is carried out for a time period of 16 days at 28 °C.

The results are provided in the Table No. 4:

Table 4: Effect of enrichment component on Nata-de-coco yield

Conclusion:

It was found that the addition of pineapple juice result in increased Nata-de-coco yield.

Further, it was found that the yield of Nata-de-coco is directly proportional to the concentration of pineapple juice.

Example 5: Effect of pH on Nata-de-coco yield

The process of Example II was followed to study the effect of pH on Nata-de-coco yield.

The effect of pH on the yield of Nata-de-coco by different isolates of Gluconoacetobacter xylinum was studied with varied pH levels of 3.5, 4.0, 4.5, 5.0 and 5.5 in 100 ml of coconut water broth. The pH of the medium was maintained by adding suitable acid (as per the procedure followed by Embuscado et al., (1994a)). The pH of the medium before and after Nata production was determined by using Beckmarm model H-4 glass electrode pH model. Coconut medium contains sucrose 10% and NH4H2PO4-0.5%.

The results are provided in the Table No. 5: Table 5: Effect of pH on Nata-de-coco yield
Conclusion:

It was found that the maximum yield of Nata-de-coco is achieved at a pH of about 4.5 Further, increase or decrease in pH result in lowering of Nata-de-coco yield.

Example 6: Effect of fermentation temperature on Nata-de-coco yield

The process of Example II was followed to study the effect of fermentation temperature on Nata-de-coco yield. The effect of temperature on the production of Nata-de-coco by different isolates of Gluconoacetobacter xylinum was studied by inoculating all the isolates to a coconut water broth and exposed to different temperature conditions continuously. The varied temperatures of 25°C, 30°C and 35°C were maintained at different sugar levels of 2.5%, 5% and 10% in coconut water inoculum for Nata-de-coco production from different isolates and the final fresh weight was recorded after 16 days of incubation.

The results are provided in the Table No. 6:

Table 6: Effect of fermentation temperature on Nata-de-coco yield

Conclusion:

It was found that when fermentation is carried out at 30 °C maximum yield of Nata-de-coco was achieved. Further, it was observed that any deviation from 30 °C result in decrease in Nata-de-coco yield.

Example 7: Effect of surface area on Nata-de-coco yield

The process of Example II was followed to study the effect of surface area on Nata-de-coco yield during fermentation. The effect of surface area on the yield of Nata-de-coco by different isolates of Gluconoacetobacter xylinum was studied by fermenting all the isolates in 100 ml broth in different vessels of varying diameter. Coconut medium contains Sucrose 10% and NH4H2PO4-0.5% at 30°C and 4.5 pH for 15 days.

The results are provided in the Table No. 7:

Table 7: Effect of surface area on Nata-de-coco yield

Conclusion:

It was found that the maximum surface area result in maximum yield of Nata-de-coco under static culture condition. Further, It was observed that yield of Nata-de-coco is directly proportional to the surface area.

Example III: A process for manufacturing Nata-de-coco using Gluconoacetobacter xylinum, nutrient and yeast.

1 kg of freshly grated coconut was mixed with 10 to 30 liter of water (coconut water and/or tap water) and strained through cheese cloth to obtain coconut water/milk. To the obtained coconut water/milk, nutrient was added;

wherein the nutrient is at least one selected from the group consisting of sugar carbon sources such as fructose, glucose, sucrose and lactose (1.0 % to 15.0 % with respect to the total mass of the inoculated coconut substance), organic acid carbon sources such as acetic acid, succinic acid, lactic acid and gluconic acid (1.0 % to 5.0 % with respect to the total mass of the inoculated coconut substance), alcohol carbon sources such as glycerol and ethanol (1.0 % to 5.0 % with respect to the total mass of the inoculated coconut substance), inorganic nitrogen sources such as ammonium sulphate, potassium nitrate and ammonium phosphate (0.1 % and 2.0 % with respect to the total mass of the inoculated coconut substance), organic nitrogen sources such as tryptone, peptone and yeast extract (0.1 % and 2.0 % with respect to the total mass of the inoculated coconut substance), vitamins such as thiamine mononitrate, vitamin B2, vitamin B12, nicotinamide, calcium pentothenate, folic acid, vitamin B6 and vitamin C (0.01 % to 0.05 % with respect to the total mass of the inoculated coconut substance) and enrichment component such as pineapple juice (1.0 % to 5.0 % with respect to the total mass of the inoculated coconut substance) to obtain a coconut water broth.

The coconut water broth was boiled at a temperature 121°C and allowed to cool. The cooled coconut water broth was then inoculated with microbial consortium containing Gluconoacetobacter xylinum and yeast at a proportion ranging between 5 % and 20 % with respect to the total mass of the inoculated coconut substance. The inoculated coconut substance was then poured into a big mouthed vessel having large surface area and ample space at top of the inoculated coconut substance. The vessel top was covered to allow inoculated coconut substance to ferment at 3.5 to 5.5 pH and temperature ranging between 20 °C and 40 °C for a time period ranging between 6 days and 15 days. Nata-de-coco was produced in the vessel which was harvested therefrom and washed.

To make Nata-de-coco edible, the harvested Nata-de-coco was cut into cubes which were subjected to a series of boiling with fresh water to remove acidity. The cubes were then poached in sugar solution and mixed thoroughly. The cubes were boiled in sugar solution in the ratio of 1:1 till the cubes became transparent to obtain edible Nata-de-coco which was packed in glass jars or pouches, sterilized and sealed.

The rate of production of Nata-de-coco was maximum (80%) in initial 8 days of fermentation. Later it reached stationary phase but still it is economical to harvest Nata-de-coco after 16 days of fermentation. The population of Gluconoacetobacter xylinum was also co-related with the production of Nata.

The concentration of total sugars in the inoculum decreased as the fermentation period progressed but there was no significant change in the concentration of reducing sugar during incubation. The total titrable acidity decreased while non volatile acidity increased during fermentation period.

Example 8: Effect of concentration of microbes on Nata-de-coco yield

The process of Example III was followed to study the effect of concentration of microbes on Nata-de-coco yield. The effect of different concentration of microbes with respect to the total mass of the inoculated coconut substance on the yield of Nata-de-coco by different isolates of Gluconoacetobacter xylinum was studied at 10% sucrose concentration with respect to the total mass of the inoculated coconut substance for different isolates. Final fresh weight was recorded as per the method followed by Gollarado, et. al, (1971). The growing medium was inoculated with different concentration of microbes (48 hour culture) which was maintained in coconut water broth. The concentrations of microbial inoculum were 5 %, 10 %, 15 % and 20 % with respect to the total mass of the inoculated coconut substance at 30 °C and 4.5 pH.

The results are provided in the Table No. 8:

Table 8: Effect of concentration of microbial inoculum on Nata-de-coco yield

Conclusion:

It was found that 10 % concentration of microbial inoculum with respect to the total mass of the inoculated coconut substance is optimum for higher Nata-de-coco yield.

Example 9: Synergistic effect of microbial consortium on Nata-de-coco yield

The process of Example III was followed to study the effect of microbial consortium on Nata-de-coco yield. The synergistic effect of Gluconoacetobacter xylinum on Nata-de-coco yield was studied. The microorganisms which were isolated from the same source and were non-Nata producing isolates were co-inoculated with Gluconoacetobacter xylinum. The different combinations were studied for their effect on Nata-de-coco yield at 5% and 10% sucrose (using the method of Collado (1988)) in 100 ml of coconut water broth. Finally, the fresh weight of Nata was recorded at 16th day and the best combination was used for further studies.

The results are provided in the Table No. 9:

Table 9: Synergistic effect of microbial consortium on Nata-de-coco yield

Conclusion:

It was found that the yeast No. 5 isolated along with Gluconoacetobacter xylinum grown on pineapple source cause significant increase in Nata-de-coco yield.

The texture of Nata-de-coco varied from soft to tough depending on sucrose concentration in the inoculated coconut substance. Softness of Nata-de-coco increases with increased sucrose concentration in the coconut water medium.

Example 10: Effect of combination of Gluconoacetobacter xylinum and yeast with sugar carbon source on Nata-de-coco yield

The process of Example III was followed to study the effect of combination of sugar carbon source at same concentration on Nata-de-coco yield. The combination of Gluconoacetobacter xylinum and yeast which yielded maximum Nata-de-coco was used to the study the effect of sugar carbon source on Nata-de-coco yield. The combinations of different sugar carbon sources were used in 100 ml coconut water broth at the same concentration of 5% to study their effect on Nata-de-coco yield.

The results are provided in the Table No. 10:

Table 10: Effect of combination of Gluconoacetobacter xylinum and yeast with sugar carbon source on Nata-de-coco yield

Conclusion:

It is found that the combination of lactose and sucrose provided the highest yield of Nata-de-coco.

Example 11: Effect of combination of sugar carbon source at different concentration on Nata-de-coco yield

The process of Example III was followed to study the effect of combination of sugar carbon source at different concentration on Nata-de-coco yield. The combination of Gluconoacetobacter xylinum and yeast which yielded maximum Nata was used to study the effect of sugar carbon source. The different combinations of sugars were used in 100 ml coconut water broth at different concentration to study their effect on Nata-de-coco yield.

The results are provided in the Table No. 11:

Table 11: Effect of combination of sugar carbon source at different concentration on Nata-de-coco yield

Conclusion:

The texture of Nata-de-coco varied from soft to tough depending on sugar carbon source concentration in the inoculated coconut substance.

Example 12: Effect of vitamin concentration and broth composition on Nata-de-coco yield

The production of Nata-de-coco by the combination of Gluconoacetobacter xylinum and yeast with different vitamin concentration and broth composition was studied using the process of Example III. Final fresh weight of Nata-de-coco was recorded. The 100ml vitamin stock solution was prepared using following vitamins;

10.0 mg of thiamine mononitrate, 100.0 mg of vitamin B2, 3.0 mg of vitamin B12, 100.0 mg of nicotinamide, 50 mg of calcium pentothenate, 1500.0 ug of folic acid, 15 ug of vitamin B6 and 150 mg of vitamin C.

The yield of Nata-de-coco by was significantly enhanced when vitamins were used in the
presence of Gluconoacetobacter xylinum mid yeast (70/100ml).

The use of vitamins for Nata-de-coco production reduced the sugar requirement from
2.5% to 1.0%.

The results are provided in the Table No. 12:

Table 12: Effect of vitamin concentration and broth composition on Nata-de-coco yield

Conclusion:

From the results as shown in table no. 12, it is found that:-

- The yield of Nata-de-coco is ranging between 11% and 16 % when only Gluconoacetobacter xylinum is used for fermentation along with vitamin.

- The yield of Nata-de-coco is ranging between 47% and 71 % when combination of Gluconoacetobacter xylinum and yeast is used for fermentation along with vitamin.

Example 13: Effect of inoculum of microbial consortium on Nata-de-coco yield

The process of Example III was followed to study the effect of concentration of microbial consortium on Nata-de-coco yield. The effect of different inoculated coconut substance concentration on Nata-de-coco yield were tested at 2.5 % sucrose concentration in coconut water broth for different isolates of Gluconoacetobacter xylinum in combination with yeast and final fresh weight of Nata-de-coco was recorded as per the method followed by Gallarado, et. al., (1971). The experiment was conducted in containers of 10cm diameter. Broth was supplemented with 0.02ml vitamin stock solution per 100ml.

The growing medium was inoculated with different concentrations of inoculum (48 hour culture) which was maintained in optimized coconut water broth. The different concentrations of inoculum were 5%, 10%, 15% and 20% with respect to the total mass of the inoculated coconut substance.

The results are provided in the Table No. 13:

Table 13: Effect of concentration of microbial consortium on Nata-de-coco yield

Conclusion:

The yield of Nata-de-coco was enhanced at 10% concentration of microbial consortium containing Gluconoacetobacter xylinum and yeast with respect to the total mass of the inoculated coconut substance when used with 0.02ml vitamin and 2.5% sucrose.

In general, it was observed that the Nata-de-coco produced by different isolates under different conditions varied in their textural and organoleptic qualities.

Throughout this specification the word "comprise", or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated element, integer or step, or group of elements, integers or steps, but not the exclusion of any other element, integer or step, or group of elements, integers or steps.

The use of the expression "at least" or "at least one" suggests the use of one or more elements or ingredients or quantities, as the use may be in the embodiment of the disclosure to achieve one or more of the desired objects or results.

The numerical values mentioned for the various physical parameters, dimensions or quantities are only approximations and it is envisaged that the values higher/lower than the numerical values assigned to the parameters, dimensions or quantities fall within the scope of the disclosure, unless there is a statement in the specification specific to the contrary. Wherever a range of values is specified, a value up to 10% below and above the lowest and highest numerical value respectively, of the specified range, is included in the scope of the disclosure.

In view of the wide variety of embodiments to which the principles of the present disclosure can be applied, it should be understood that the illustrated embodiments are exemplary only.

While considerable emphasis has been placed herein on the particular features of this disclosure, it will be appreciated that various modifications can be made, and that many changes can be made in the preferred embodiments without departing from the principle of the disclosure. These and other modifications in the nature of the disclosure or the preferred embodiments will be apparent to those skilled in the art from the disclosure herein, whereby it is to be distinctly understood that the foregoing descriptive matter is to be interpreted merely as illustrative of the disclosure and not as a limitation.

We Claim:

1. A process for manufacturing Nata-de-coco, said process comprising the following
steps:

i) treating a coconut substance with at least one nutrient;

ii) inoculating the treated coconut substance with a microbial consortium containing Gluconoacetobacter Xylinum and yeast to obtain inoculated coconut substance;

iii) fermenting the inoculated coconut substance to produce Nata-de-coco; and

iv) harvesting Nata-de-coco therefrom.

2. The process for manufacturing Nata-de-coco as claimed in claim 1, wherein the coconut substance is at least one selected from the group consisting of coconut meat, grated coconut meat, coconut water and coconut milk.

3. The process for manufacturing Nata-de-coco as claimed in claim 1, wherein the nutrient is at least one selected from the group consisting of sugar carbon sources, organic acid carbon sources, alcohol carbon sources, inorganic nitrogen sources, organic nitrogen sources, vitamins and enrichment components.

4. The process for manufacturing Nata-de-coco as claimed in claim 3, wherein the sugar carbon source as a nutrient is at least one selected from the group consisting of fructose, glucose, sucrose and lactose, and the proportion of sugar carbon source ranges between 1.0 % and 15.0 % with respect to the total mass of the inoculated coconut substance.

5. The process for manufacturing Nata-de-coco as claimed in claim 4, wherein the sugar carbon source as a nutrient is sucrose in an amount 1.0 to 10.0 % with respect to the total mass of the inoculated coconut substance.

6. The process for manufacturing Nata-de-coco as claimed in claim 3, wherein the organic acid carbon source as a nutrient is at least one selected from the group consisting of acetic acid, succinic acid, lactic acid and gluconic acid, and the proportion of the organic acid carbon source ranges between 1.0 % and 5.0 % with respect to the total mass of the inoculated coconut substance.

7. The process for manufacturing Nata-de-coco as claimed in claim 6, wherein the organic acid carbon source as a nutrient is acetic acid in an amount 1.0 to 2.0 % with respect to the total mass of the inoculated coconut substance.

8. The process for manufacturing Nata-de-coco as claimed in claim 3, wherein the alcohol carbon source as a nutrient is at least one selected from the group consisting of glycerol and ethanol, and the proportion of the alcohol carbon source ranges between 1.0 % and 5.0 % with respect to the total mass of the inoculated coconut substance.

9. The process for manufacturing Nata-de-coco as claimed in claim 3, wherein the inorganic nitrogen source as a nutrient is at least one selected from the group consisting of ammonium sulphate, potassium nitrate and ammonium phosphate, and the proportion of inorganic nitrogen source ranges between 0.1 % and 2.0 % with respect to the total mass of the inoculated coconut substance.

10. The process for manufacturing Nata-de-coco as claimed in claim 3, wherein the organic nitrogen source as a nutrient is at least one selected from the group consisting of tryptone, peptone and yeast extract, and the proportion of organic nitrogen source ranges between 0.1 % and 2.0 % with respect to the total mass of the inoculated coconut substance.e process for manufacturing Nata-de-coco as claimed in claim 10, wherein the organic nitrogen source as a nutrient is combination of yeast extract and peptone in an amount 0.1 to 1.0 % with respect to the total mass of the inoculated coconut substance.

11. The process for manufacturing Nata-de-coco as claimed 10, wherein the organic nitrogen source as a nutrient is combination of yeast extract and peptone in an amount 0.1 to 1.0% with respect to the total mass of the inoculated coconut substance.

12. The process for manufacturing Nata-de-coco as claimed in claim 3, wherein the vitamin as a nutrient is at least one selected from the group consisting of thiamine mononitrate, vitamin B2, vitamin B12, nicotinamide, calcium pentothenate, folic acid, vitamin B6 and vitamin C, and the proportion of vitamin ranges between 0.01 % and 0.05 % with respect to the total mass of the inoculated coconut substance.

13.The process for manufacturing Nata-de-coco as claimed in claim 3, wherein the enrichment component is pineapple juice, and the proportion of enrichment component ranges between 1.0 % and 5.0 % with respect to the total mass of the inoculated coconut substance.

14. The process for manufacturing Nata-de-coco as claimed in claim 1, wherein the method step of treating is carried at a temperature 121°C.

15. The process for manufacturing Nata-de-coco as claimed in claim 1, wherein the proportion of microbial consortium containing Gluconoacetobacter Xylinum and yeast ranges between 5 % and 20 % with respect to the total mass of the inoculated coconut substance.

16. The process for manufacturing Nata-de-coco as claimed in claim 1, wherein the proportion of Gluconoacetobacter Xylinum: yeast is 1:1.

17. The process for manufacturing Nata-de-coco as claimed in claim 1, wherein the method step of fermentation is carried out at a temperature ranging between 20 °C and 40 °C for a time period ranging between 6 days and 15 days.

18. The process for manufacturing Nata-de-coco as claimed in claim 1, wherein the method step of fermenting is carried out at a pH ranging between 3.5 and 5.5, preferably 4.0 and 5.0.

19. The process for manufacturing Nata-de-coco as claimed in claim 1, wherein the method step of harvesting is carried out by separating Nata-de-coco from the fermented coconut substance followed by pressing and cutting into cubes.

20. The process for manufacturing Nata-de-coco as claimed in claim 1, further comprising the following steps;

i) Boiling Nata-de-coco cubes in water at least once to obtain Nata-de-coco cubes with a predetermined pH; and

ii) Poaching Nata-de-coco cubes in sugar solution followed by packaging under aseptic conditions.