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Aloe vera: Development of gel extraction process for Aloe vera leaves

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Aloe vera is a succulent belongs to the Liliaceae family. It is a native, chiefly to the warm dry areas of South Africa, but cannot survive freezing temperatures. The cultivation of Aloe vera has acquired great commercial importance for medicinal products and cosmetics processing but information is scarce about processing of this crop. This investigation was aimed at standardizing important process parameters with the specific objective of developing an appropriate process technology for extraction of gel from Aloe vera barbadensis. The principle of centrifugation was employed for the extraction of gel from Aloe vera leaves The experiment was planned using a 4 factor Completely Randomised Design. The treatments consisted of two levels of acetone i.e. without addition of acetone and 10 % acetone, three level of centrifuge temperature i. e. 5, 10 and 32 0C (Ambient), three level of centrifuge speed i.e. 2000, 5000 and 10000 rpm and three level of centrifuge duration i. e. 10, 20 and 30 min with 3 replications. The optimum proportion of acetone, temperature, speed and duration was decided on the basis of quality parameters of gel. The results obtained are analysed statistically. The effect of acetone, temperature, speed and duration on quality parameters like gel recovery (%), viscosity of gel, (Stokes), refractive index of gel, optical density of gel, and TSS content of gel, (Brix) were studied. The combined effect of the different extraction parameters such as proportion of acetone addition to pulp, centrifuge temperature, speed and duration were also studied. Physical properties of Aloe vera leaf such as length, width, thickness, weight and pulp weight were measured. The average value of length, width, leaf weight, pulp weight, apparent volume, and pulp recovery was ranged from 480 to 655 mm, 55 to 115 mm, 326 to 658 g, 156 – 331 g, 127.32 – 485.73 cc and 42.82 – 57.14 % respectively for Aloe vera leaf. The moisture content, fiber content, pH and sugar content was found to be 99.8 %, 0.2 %, 6.389, 1.9125 (total sugar) and 0.0259 (reducing sugar) from Aloe vera gel respectively. It may be recommended that the extraction of gel from Aloe vera by the method of centrifuge should be carried out at 5 0C temperatures, 10000 rpm speed and 30 min duration of centrifuge, without addition of acetone to pulp so as to obtain higher gel recovery (50.17 %) and good quality of gel i.e. viscosities: 0.675 (Stokes), refractive index: 1.33550, optical density: 0.218 (abs) and TSS content: 0.93 (Brix).
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CONTENTS
Chapter Title Page
No. No.
Contents 1
Acknowledgement 2
List of Abbreviations 3
List of Nomenclature 5
I Introduction 7
II Review of Literature 11
III Materials and Methods 37
IV Results and Discussion 50
V Summary and Conclusion 95
References 101
Appendices 105
2
ACKNOWLEDGEMENT
The author wishes to express profound gratitude and esteemed reverence to
major guide Dr. A. K. Varshney, Professor & Head, Department of Agricultural
Process and Food Engineering, College of Agricultural Engineering and Technology,
Junagadh Agricultural University, Junagadh, minor guide Dr. A. H. Raval and
advisory committee members Prof. D. M. Vyas, Dr. V. K. Poshia and Prof. V. M.
Bhatt for providing guidance and timely suggestions during the course of
investigation. The author also grateful to Dr. V. P. Chovatiya, Professor and Head and
Mr. K.H. Dabhi Department of Botany, JAU, Junagadh for procurement of Aloe vera
raw material and utilization of Cooling type Centrifuge.
The author is sincerely grateful to Prof. J. B. Savani, Principal and Dean,
College of Agricultural Engineering and Technology, Junagadh Agricultural
University, Junagadh for providing necessary facilities during course of studies
I record my deep sense of gratitude to Dr. N. C. Patel, Hon’ble Vice
Chancellor, Junagadh Agricultural University, Junagadh for providing me an
opportunity to conduct research project. I am also grateful to Dr. C. J. Dangaria,
Director of Research & Dean, PG Studies, Junagadh Agricultural University,
Junagadh and Dr. R. L. Shiyani, Registrar, Junagadh Agricultural University,
Junagadh for their keen interest in the academic values.
The author is extremely grateful to his beloved parents and brothers for their
everlasting love, during the study. Last but not the least; the author extends deep
compliments to his wife Usha, and son Prashant and Harshit for relieving from home
responsibilities and allowing him to complete the study successfully.
Date : August 15, 2012 (V. K. Chandegara)
Juangadh
3
ABBREVIATIONS
Anon. Anonymous
Cal. Calculated
CEG Cold- extracted gel
Co. Company
Com. Commercial
FAC Fat adsorption capacity
HEG Hot extracted gel
HTST High temperature short time
http Hyper text transfer protocol
IASC International Aloe Science Council
Inc. Incorporation
ISI Indian Standard Institution
J. Journal
M/s Messer’s
MPS Methanol Precipitable Solids
POD Peroxidase
PPO Polyphenol oxidase
RCF Relative centrifugal force
SW Swelling
Syn. Synonymous
TTS Time, temperature and sanitation
TSS Total soluble solid
Vol. Volume
UK United Kingdom
US United States
WRC Water retention capacity
WHO World Health Organization
www World wide web
4
NOMENCLATURE
ANOVA Analysis of Variance
Av. Average
Abs absolute
CaCl
2
Calcium Chloride
cc cubic centimetre
cm Centimeter
CO
2
Carbon Dioxide
C.D. Critical Difference
C.R.D. Completely Randomised Design
C.V. Coefficient of Variance
ed. Edition
d.f. Degree of freedom
dl Deci litre
et al. And others
etc. Etcetera
F Centrifugal force
f Function
Fig. Figure
g Gram
gm Gram mole
gr Gram
h Hour
HCL Hydrochloric Acid
ha Hectare
hp Horse power
H
2
SO
4
Sulphuric acid
i.e. That is
kcal Kilo calorie
5
kg Kilogram
kw Kilowatt
Kwh Kilowatt per hour
Kcal Kilocalories
kg Kilogram
L Length
m Meter
meq Mili equivalent
mOsm Mili osmo
min Minute
mm Millimetre
M.S.S. Mean sum of squares
mm
3
Cubic milimeter
mg Milligram
min Minute
mg/L Mili gram per litre
ng Nena gram
nm Nano meter
No. Number
Para Paragraph
pg Pico gram
pH Negative logarithm (base 10) of the hydrogen ions (H+)
ppm Parts per million
pp Pages
r Radius of rotation
rpm Revolution per minute
Rs. Rupees
Sd Standard deviation
S.Em. Standard error of mean
Sig. Significant
6
S.S. Sum of squares
Sr. Serial
T Thickness
t Time
V Volume of the leaf
vs. Versis
viz., Videlicet (that is to say)
wb Wet basis
W Width
µg Micro gram
µm Micro meter
µmole/g Micro mole per gram
LIST OF SYMBOLS
% Per cent
@ At the rate of
0
A Degree Angstrum
0
C Degree Celsius
µ Kinematic Viscosity
ω Angular velocity
π Pie
β Beta
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CHAPTER I
INTRODUCTION
Aloe vera is a succulent, belongs to the Liliaceae family. It is one of the 250
known species of aloes, referred to by the scientific terms of Aloe vera and Aloe
barbadensis. The semi-tropical plant, Aloe vera barbadensis Miller from the Lily
(Liliaceae) family has a long and illustrious history dating from biblical times. It has
been mentioned throughout recorded history and given a high ranking as an all-
purpose herbal plant. Aloe's thick, tapered, spiny leaves grow from a short stalk near
ground level.
Aloe Juice is colourless, transparent water like juice obtained from fresh aloe
leaves. It is tasteless and odourless. Aloe vera Latex commonly known as aloin
having a bitter taste and purgative quality. Because of its bitter taste, it is also known
as bitter aloes. Aloin is destructive to healthy tissue and cells. It is obtained from
specialized cells known as pericyclictubules that occur just beneath the epidermis or
rind of the same leaves from which the juice is derived. Aloe can be used in raw form
or in processed forms; it can be used both externally and internally.
Dried Juice is commercially known as aloe or musavvar. This is the solidified
juice coming spontaneously when the leaf is cut out of the cells in pericycle and
adjacent leaf parenchyma. This juice is then dried with or without heat to give a
strongly bitter substance having a characteristic disagreeable odour. This is known as
aloe in the market. Three types of aloe are available in market depending upon the
source plant viz. (i) Curacao aloe: a dark brown coloured substance sourced from
Aloe vera (ii) Cape aloe: greenish brown coloured aloe sourced from Aloe ferox and
(iii) Socotrine aloe: reddish black coloured aloe sourced from Aloe peyrii baker. The
original commercial use of the Aloe plant was in the production of a latex substance
called Aloin, a yellow sap used for many years as a laxative ingredient.
8
Aloe vera gel is the commercial name given to the fiber free mucilaginous
exudate extracted from the hydroparenchyma of the succulent leaves of Aloe vera
(Aloe barbadensis Miller). Aloe vera Gel a clear colorless, jelly-like material is
derived from tissue that comprises the inner portion of the leaves. It is slightly bitter
and odourless. The gel loses its transparency if extracted after 3 hours of plucking the
leaves. This Aloe vera Gel, beginning in the 50's, has gained respect as a commodity
used as a base for nutritional drinks, as a moisturizer, and a healing agent in
cosmetics. The exudate of Aloe vera is used for numerous medical and cosmetic
applications since ancient times (Morton, 1961). Commercially available aloe gel is
stabilized for maintenance of its potency.
The Aloe gel is used as a moisturizer for skin care, hair care products and a
healing agent in cosmetics. Medicinally it is used as antiseptic agents, natural
antibiotic agent, calming agent, detoxifier and dilator to increases circulation and
blood flow to the skin. It is also used as insect repellent, and a transparent pigment
used in miniature painting. The cords and nets are made from the leaf fiber.
Gels, concentrates and powders are suitable for cosmetic, hair care, personal
care, pharmaceutical, beverage, food, functional food, nutraceutical and dietary
supplement formulations. Products are available in regular and decolorized, cosmetic
and food grade preservative systems. The International Aloe Science Council has
solidified its dedication to providing the world with the highest quality Aloe. The
IASC has a dedicated group of professionals committed to the further growth,
research and marketing of quality Aloe vera Gel and Aloe products made from this
Gel. Hand filleting and whole leaf processing, the two types of Aloe vera gel
extraction methods are prevalent. Gel is extracted either cold process or hot process.
Combination of hand filleting and the entire whole leaf processing are used to avoid
the undesirable elements, while maximizing the desired constituents. Among the
9
desirable constituents are the polysaccharides (glucomannans), glycoproteins and
associated growth factors.
However, over 95 % of the Aloes on the market today still use only the inner
gel and stabilize the Aloe in a high-heat process that degrades some of the enzymes,
polysaccharides and mucopolysaccharides. High heat (pasteurization and/or autoclave
methods) breaks down the constituents in Aloe that are the most valuable for healing.
Heat also kills the live enzymes necessary for digestion. Most Aloes are heat
processed (Grindlay et al., 1986).
Commercial exploitation of Aloe vera gel has been carried out for at least 50
years. Various companies in the US act as primary growers and processors of the
plant and manufacture bulk supplies of the gel for domestic and export market. Many
other companies are secondary processors of Aloe vera products, and cosmetics firms
and chain store often buy the gel for incorporation into their own brand name
products (Grindlay and Reynolds, 1986).
The cultivation of Aloe vera has acquired great commercial importance for
medicinal products and cosmetics processing but information is scarce about
processing of this crop. As such there is no scientific information is available in
India, regarding processing of Aloe vera, its importance in nutrition, cosmetics and
pharmaceutics.
Looking to the importance of the Aloe products such as creams, ointments,
juices, and shampoo containing the Aloe gel, it is very much essential to study the
post harvest technology of the Aloe vera plant. The expanding Aloe industry
urgently needs to develop test procedures and a reliable database so that a product
claiming to have Aloe could be tested and certified. According to Reynolds and
Dweck, (1999), there is no scientific literature available for the processing of leaf gel.
What so ever processes either gel extraction or gel stabilization are patented, so it is
10
essential to develop the gel extraction process, which may useful for industrial
community. Despite the ideal climatic conditions for the cultivation of Aloe vera, we
have not been able to exploit the excellent potential of the miraculous medicinal
plant. The reasons are simple: lack of cultivation and processing know-how.
Keeping above importance in view a research project was undertaken with the
following specific objectives:
Objectives:
1. To study the physical and chemical properties of Aloe vera leaves.
2. To develop gel extraction process.
3. To evaluate the gel extraction process
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CHAPTER II
REVIEW OF LITERATURE
This chapter deals with the review of literature on the origin and description
of Aloe vera plant, production practices of Aloe vera, physical and chemical
composition of Aloe vera leaf and gel, Aloe vera gel extraction, purification and
stabilization technology and quality parameters used for Aloe vera gel. Very little
information is available on the extraction of gel from Aloe vera leaves. However, a
brief review on the available information has been presented here as under.
2.1 Aloe vera
Several species of the genus aloe has been in use under the common name of
aloe viz. Aloe vera, Aloe barbadensis, Aloe ferox, Aloe chinensis, Aloe indica, Aloe
peyrii, etc. Amongst these Aloe vera Linn syn. Aloe barbadensis Miller is accepted
unanimously as the correct botanical source of aloe. In most reference books Aloe
barbadensis Miller is regarded as the correct name but as per the WHO monograph
Aloe vera (L) Burm f. is accepted as the legitimate name for this species. The genus
aloe is placed taxonomically in Liliaceae family. It has also been known as Aloe
vulgaris ("common aloe") and Aloe barbadensis.
2.1.1 Origin of Aloe vera
Aloe vera barbadensis a member of the lily (Liliaceae) family is a spiky,
succulent, perennial plant and a native to warm dry regions, especially southern
Europe, Asia, and Africa. It is indigenous to eastern and southern Africa and has been
spread throughout many of the warmer regions of the world. It grows best in full
sunshine and does not require much water (Denk, 2000 http://www.bio.gasou.edu/). It
is popularly grown as indoors plant and cultivated almost everywhere in the world,
12
both as a houseplant and for its medicinal qualities. It is commercially cultivated in
Aruba, Bonaire, Haiti, India, South Africa, the United States of America, and
Venezuela. There are about 300 identified species.
2.1.2 Physical description of Aloe vera plant
Aloe vera barbadensis can grow up to 1000 mm height, although most
specimens are 300 to 600 mm in height (Gilman, 1999 http://www.hort.ifas.ufl.edu).
It has thick leaves that grow in a rosette shape (Denk, 2000
http://www.bio.gasou.edu/). The parenchyma cells of the leaves contain large
quantities of pulp. It is an evergreen perennial succulent plant that looks like a cactus
grown throughout the world. The fleshy leaves with serrated edges that arise from a
central base and grow to nearly 300 500 mm long and 100 mm width at the base.
Usually each plant is having 12-16 leaves, when mature and weigh about 1500 g. The
Aloe leaf consists of three layers i.e. (a) the outer thick rind, (b) a viscous, jelly like
mucilage layer into which the vascular bundles are attached to the inner surface of the
rind, protrude and (c) the fillet proper, which has structural integrity consisting of
hexagonal, structures containing the fillet fluid. This is the water storage area for the
plant. (www.bonasana.com) The fibrous outer part of the leaf serves a protective
function.
The plant has yellow flowers having 250 350 mm in length arranged in a
slender loose spike; stamens frequently project beyond the perianth tube. A clear gel
is colorless mucilaginous gel obtained from the parenchymatous cells in the fresh
leaves of Aloe vera (Bruneton, 1995 and Grindlay et al., 1986). Gel is a part of the
plant used for topical application. Anthraquinones, which exert a marked laxative
effect, are contained in the bitter yellow sap of the middle leaf layer. Agarwala,
(1997) studied the pharmaceutical properties of aloes and suggested that a clear
distinction between substances in the colorless, tasteless parenchyma cells, is called
13
aloe gel and substances in the bitter exudate from cells associated with vascular
bundles in the outer green rind of the leaf is known as the aloin.
2.1.3 Production practices of Aloe vera
Aloe vera requires tropical, sub-tropical climatic conditions with a very sunny
position and a well-drained loamy soil. Plants are tolerant of poor soils (Farooqi and
Sreeramu, 2001). Though Aloe vera can be cultivated on any soil for 'dry land
management', sandy loamy soil is the best suited for it. It does not grow well at
temperatures below 32 degree
Fahrenheit or zero degree
Celsius ( 0
0
C).
Aloe vera is generally propagated by root suckers and planting them to the
main field or it can also be propagated through rhizome cuttings. Aloe is a perennial
and takes 4-5 years to mature. Plants can live and reproduce for up to 25 years (Denk,
2000). The plants can be harvested every 6 to 8 weeks by removing 3 to 4 leaves per
plant. Aloe vera leaves are normally sensitive to subfreezing temperatures The
weather conditions are highly affect the Aloe vera processing schedule. Pulling back
on the green leaf and cutting at the white base carry out the harvesting of the Aloe
vera plant. (http://www. aloecorp.com). Sachedina and Bodeker, (1999) reported the
harvesting of Aloe vera leaves by hand pulling.
The potential for commercial cultivation of Aloe vera has also been
considered in Tanzania by Sachedina, (1998), who recommends the encouragement
of village medicinal plant gardens; followed by the establishment of Aloe growing
co-operatives supplying a central processing plant for local production; and
eventually a plantation, nursery and processing plant for export.
14
2.2 Handling of Aloe vera leaves
Biological activity loss is due to the microbial decay of the gel. The first
exposure of the inner gel to microbes is when the leaves are harvested from the plant.
Leaves in which the base is not intact and sealed will greatly increase the microbial
counts in the finished product. Higher level of microbial counts significantly reduces
the biological activity in the product. The other major source of microbial
contamination comes from the rind of the leaf. To prevent contamination of the gel,
the leaves are handled very carefully and soaked in a food grade sanitizer, which
effectively reduces the microbial counts in the leaf exterior to acceptable levels.
(http://www. aloecorp.com)
2.3 Physico-chemical properties of Aloe vera leaves
Wang and Strong, (1993) have studied the physical properties of Aloe vera
leaves and reported that, the average weight of the individual leaves was ranging
from 387 to 704 g, length 48 to 60 cm, width 8.9 to 11.5 cm and optical density 1.020
to 1.437 (abs) (Table 2.1).
Table 2.1 Characteristics of fresh Aloe vera leaves from producers averaged
over a 6-month period (Wang and Strong, 1993)
Grower Weight
(g)
Leaf length
(cm)
Width
(cm)
Optical density
(abs)
1 478 53 9.7 1.095
2 611 56 11.4 1.356
3 704 60 11.5 1.020
4 387 48 8.9 1.437
15
Shih-Jen Chiou, (2003, http://ethesys.lib.pu.edu.tw/ETD-db/ETD-search/)
studied the physicochemical properties of Aloe vera including vitamins, viscosity,
molecular weight distribution, and monosaccharide composition of Aloe vera. The
contents of vitamin C and B1 were significantly decreased after blanching treatment
but those of niacin and vitamin B2 differed significantly. Heating treatment, resulting
in the decomposition of Aloe vera polysaccharide, decreased the average molecule
weight of Aloe vera. The viscosity of Aloe vera also declined with the increase of
heating time. Changing the pH value and adding thio-compounds could efficiently
inhibit the degree of nonenzymatic browning of blanched Aloe vera during storage.
2.4 Chemical composition of Aloe vera leaves
Joshi, (1998) had given the chemical constituents of Aloe vera barbadensis.
The aloe plant contains 99 and 99.5 per cent water, with an average pH of 4.5. The
remaining solid material contains over 75 different ingredients including vitamins,
minerals, enzymes, sugars, anthraquinones or phenolic compounds, lignin, saponins,
sterols, amino acids and salicylic acid. Table 2.2 present the analytical profile of Aloe
vera leaves.
The main constituents of the Aloe vera leaves are: (i) Aloin: It is an irritant
laxative contained in the yellow sap of Aloe, which is a constituent of the
Anthraquinone complex, (ii) Methanol-Precipitable Solids (MPS): When alcohols are
added to Aloe solutions about 20-25 % of the total solids come out of solution or
'precipitate'. It consists mainly Polysaccharides, Glycoprotein and salts of Organic
Acids. The Polysaccharides represent about one-half to two-thirds of the MPS or
about 10-15% of the total solids. (iii) Polysaccharides: There are over 200
constituents in Aloe vera, the single most important constituent being the
polysaccharides. Polysaccharides consist of simple sugar molecules and are called
hexoses.
16
Table 2.2 Analytical profiles of Aloe vera leaves
Tests Units Minimum Maximum Aeverage
Solids % 0.75 1.50 0.92
Water % 98.5 99.25 99.1
Glucose mg/dl 28.0 103.0 77.8
Purine mg/dl 0.1 5.6 0.8
Ueea-Nitrogen mg/dl 1.0 1.0 1.0
Creatinine mg/dl 0.1 1.5 0.4
Sodium meq/l 4.0 13.0 8.7
Potassium meq/l 10.0 22.5 13.4
Chloride meq/l 1.0 11.0 3.0
CO2 meq/l 1.0 7.0 1.7
Calcium mg/dl 19.4 48.5 30.0
Cal. Calcium mg/dl 23.3 52.3 33.8
Magnesium mg/dl 3.2 4.7 3.9
Zinc mg/dl 14.0 77.0 31.0
Phosphorus mg/dl 0.6 1.3 1.0
Total Protein gm/dl 0.1 0.4 0.2
Albumin gm/dl 0.1 0.5 0.14
Globulin gm/dl 0.0 2.0 0.06
Alkaline
Phosphatase mg/dl 1.0 50.0 18.0
Sgotransaminase mg/dl 6.0 49.0 21.0
Sgotransaminase mg/dl 8.0 85.0 24.0
Lactic
Dehydrogenase mg/dl 0.0 9.0 3.0
Amylase mg/dl 0.0 2.0 1.0
Lipase units/dl 0.0 1.6 0.5
Cholestrol mg/dl 4.0 12.0 8.0
Triglycerides mg/dl 1.0 12.0 2.4
Iron mg/dl 3.0 30.0 15.0
B12 pg/ml 141.0 403.0 265.0
Folic Acid ng/ml 2.7 20.0 13.2
Osmolarity m0sm/kg 43.0 67.0 60.0
HPLC Ratio 0.51 1.1 0.67
Source: http://www.drwolfe.com
17
2.5 Physico-chemical properties of Aloe vera Gel
Following companies had reported the physico-chemical properties of Aloe
vera gel and shown in Table 2.3. This meets or exceeds the standards established by
the International Aloe Science Council's (IASC) certification program for the
determination of purity.
Table 2.3 Physico-chemical properties of Aloe vera gel
Test http://www.aloecorp.com M/s Delta International
(http://www.shantidatta.com)
Appearance Clear Yellow / Green
Liquid
*Absorbance @
400nm NMT 0.500
Refractive Index 1.3340-1.3355 1.33789 - 1.34390
Specific Gravity 1.0030-1.0070 1.0221 - 1.0339
pH Value 3.5-4.7 3.5 - 4.7
Total Solids NLT 0.46% by weight
*Reported as reconstituted
Table 2.4 Chemical properties of Aloe vera gel (Wang and Strong, 1993)
Grow
pH Aloin
(mg/L)
Soluble solids
(%)
Sugars
(mg/L)
*Fibre
(%)
1 4.47 31 0.708 2555 0.077
2 4.49 39 0.610 1441 0.088
3 4.51 29 0.675 2530 0.088
4 4.54 41 0.586 1361 0.074
* Percentage of fresh weight
18
2.6 Chemical composition of Aloe vera gel
The Chemical composition of Aloe vera gel as reported by Wang and Strong,
(1993) is presented in Table 2.4. Waller et al., (1978) had worked on sugar analysis
of Aloe vera gel and given the sugar composition (Table 2.5). The other scientists
namely Pierce (1983), Rowe and Park, (1941), Bruneton, (1995) and Devis et al.,
(1994) had worked time to time and reported following chemical composition of
Aloe vera gel which is shown in Table 2.6.
Table 2.5 Sugar content of Aloe vera gel (Waller et al., 1978)
Sugar Sugar in whole gel
mole/g) Total Sugar in lyophilized
residue (%)
Arabinose 4.23
a
4.7
a
Galactose 3.60 4.3
Glucose 31.3 37.7
Mannose 39.4 47.5
Rhamnose 1.27 1.5
Xylose 4.44 4.4
a
Arabinose could not be distinguished from fructose
19
Table 2.6 Aloe gel composition
Mono and polysaccharides (50-60% of solids)
(Specific concentrations have not yet been determined)
Ployhexanoses
Hexans Xylose Arabinose Galactose Glucose
Amino acids (ppm)
Lysine 5-6 Histadine 2.8-3.3 Arginine 4.5-5.5
Threonine 5-6 Aspartic
Acid
13-15 Serine 6-7
Glutamic Acid
13.5-15.5 Proline 8-9 Analine 1.0-1.3
Glycine 7-8 Valine 6.5-7.0 Methionine 1.5-2.0
Isoleucine 3.5-4.0 Leucine 8.5-9.0 Tyrosine 2.8-3.3
Pheylalanine 4.3-4.7
Vitamins (mg per 100ml)
B-1 6-7 B-2 6-7 C 47-61
Niacinamide 30-37 B-6 3.0-3.7 Choline 9.5-11.2
Enzymes (per 100 ml)
Amylase 1100-1600 units Lipase 600-800 units
Protein 0.11g / 100 gr Fat 0.09g / 100 gr
Ash 0.25% Crude fiber 0.10%
Calories 3.3/100 gr
Source: http://www.garudaint.com
20
2.7 Quality parameters of Aloe vera gel
The quality parameters such as fiber content, viscosity, refractive index,
optical density and total soluble solid plays an important role in judging the quality
and purity of extracted gel from Aloe vera leaf.
2.7.1 Fiber content
The fiber content is directly related to the purity of gel and become the criteria
of efficiency of gel filtration unit. More fiber content, suggests poor filtration
operation. For getting pure gel fiber should be minimum. The difference between
crude gel recovery and pure gel recovery gives the amount of fiber in crude gel.
Wang and Strong, (1993) had found the fibre content 0.074 to 0.088 % of fresh
weight of pulp.
2.7.2 Viscosity
Viscosity of gel is very important factor for deciding quality in terms of
activities of biological compounds. The viscosity decreases as the time passes. After
some duration viscosity of gel becomes equal to the viscosity of water, the gel
becomes water. Gowda et al., (1979) reported that after harvest of Aloe vera leaf, the
viscous pseudoplastic nature of Aloe vera gel, mainly due to the presence of
polysaccharides composed of a mixture of acetylated glucomannans was lost shortly
after extraction, apparently due to enzymatic degradation. This shows there are some
biological activities, which related to the viscoelastic behavior of gel.
2.7.3 Refractive index
Refractive index is the physical property of gel determines the purity of gel as
compared to double distilled water. Gel with lowest refractive index, is the best
treatment for extraction process. More refractive index indicates the impurities in the
extracted gel.
21
2.7.4 Optical density
Optical density is the physical property of gel determines the purity of gel as
compared to double distilled water. Gel with lowest optical density, is the best
treatment for extraction process. More optical density indicates the impurities in the
extracted gel. Wang and Strong, (1993) have reported optical density of 1.020 to
1.437 (abs) for Aloe vera leaves.
2.8 Method of processing Aloe leaves
2.8.1 Traditional hand filleted Aloe vera
In order to avoid contaminating the internal fillet with the yellow sap, the
traditional hand- filleting method of processing Aloe leaves was developed. In this
method, the lower portion i.e. 25 mm of the leaf base (the white part attached to the
large rosette stem of the plant), the tapering point (50 –100 mm) of the leaf top, and
the short, sharp spines located along the leaf margins are removed by a sharp knife.
The mucilage layer below the green rind avoiding the vascular bundles, and the top
rind is also removed with the help of knife. The bottom rind is similarly removed.
The materials of the mucilage layer, subsequent to their synthesis, are
distributed to the storage cells (cellulose-reinforced hexagons) of the fillet, a process
that is accompanied by dilution owing to the water (the major fillet constituent),
which is stored in the fillet cells. The fillet consists of more than 99% water. The
hand filleting method is very labour intensive. Owing to this fact, machines have
been designed and employed which attempt to simulate the hand filleted techniques,
but generally the product contains higher amounts of the anthraquinones laxatives
than the traditional hand filleted approach (www.bonasana.com).
22
2.8.2 Whole leaf Aloe process
In this process, the base and tip are removed as previously delineated, and then
the leaf is cut into sections and ground into particulate slurry. The material is then
treated with chemicals which break down the hexagonal structure of the fillet
releasing the constituents, by means of a series of coarse and screening filters, or
passage through a juice press, the rind particles are removed, the expressed juice is
then passed through various filtering columns which remove the undesirable laxative
agents. This process, performed properly, can produce a constituent-rich juice
(generally containing 3 times or more constituents than hand filleted juice), which
should be virtually free of the laxative anthraquinones; this process was developed in
the 1980’s (www.bonasana.com).
The present method of processing of Aloe vera is using the whole leaf, from
which the undesirable elements can be selectively removed, while maximizing the
desired constituents. The desirable constituents are polysaccharides (glucomannans),
glycoproteins that are associated with growth factors. Table 2.7 shows the data of
hand and whole leaf filleting which reveals that the quantity of desirable
polysaccharides is 2.5 to 3 times higher as compare to hand filleting methods.
Table 2.7 Yields and Aloe leaf processing
Process Fraction Hand Filleting
(%)
Whole Leaf
(%)
Total solids (Without preservatives or
additives)
0.45 – 0.65 1.30 – 3.50
Polysaccharides 0.12 0.16
Source: http://www.wholeleaf.com
23
Table 2.8 data compares various processing methods and the effect on yield
(total solids), aloin concentration, and the distribution of sizes of constituents. The
whole leaf method can produce an Aloe juice which is high in total solids, high in
retained high dalton (molecular weight) polysaccharides with their scientifically
demonstrated benefits, while the aloin concentration is at a very acceptable low level.
Table 2.8 Methods of leaf preparation and constituents
Method of preparation pH Aloin
(ppm) H
2
O
(%) Total solids
(%)
(1) (2) (3) (4) (5)
Hand -filleting 4.27 6 99.25 0.48
Roller 4.30 32 99.61 0.39
Leaf Splitter 4.24 18 99.61 0.42
Whole Leaf 4.09 1 99.62 1.38
Source: http://www.wholeleaf.com
2.8.3 Total process Aloe
The total process of Aloe vera is combination of hand filleting and whole leaf
processing. In this process, aloe leaves are hand filleted by the traditional method.
Then the green rinds and the mucilage pulp are processed separately. A combination
of the products obtained by these two procedures, produces a product called Total
Process Aloe. Total Process Aloe contains considerably higher concentrations of total
solids, calcium, magnesium, and malic acid, which are virtually free from undesirable
laxative anthraquinones. The International Aloe Science Council for certification
recommends using the total process Aloe, which retains major portion of desirable
constituents (www.bonasana.com).
24
2.8.4 ALOECORP process
The M/s ALOECORP, an American company has suggested their own method
for the processing of Aloe vera leaf. In this method, three of the outermost mature
leaves are cut from each plant. Leaves are gathered in boxes, which are transported
immediately to the production facility. The harvested leaves are primarily washed
with hand and then conveyed through a stainless steel conveyer to the mechanical
leaf washer. The leaves are cut and finally pass through the gel expulsion machines.
The production room is kept in an ultra-sanitary state, even when not in use.
Employees are required to go through a process of sanitation every time they enter
the room. The entire area is thoroughly cleaned after each production run. Once the
gel is expelled from the leaves, it is pumped through a de-pulping machine. The pure
Aloe gel is then moved through a chilling system designed to bring the temperature
of the gel below to 37 degrees Fahrenheit or 2.5 - 3 degrees Celsius. The chilled gel is
stored in an insulated tank, ready to be pumped into a transport tanker for delivery to
the Harlingen processing facility (Aloecorp De Mexico), (http://www. aloecorp.com).
2.8.5 American Quality Aloe, Aloe vera processing
The M/s American Quality Aloe Company suggested their own methods for
processing of Aloe vera leaves. The method consists of harvesting of matured Aloe
vera leaves and processing within an hour to avoid biological contamination and
degradation of the leaves. The mature leaves are scrubbed and rinsed several times in
a special solution. The leaves are ground up and filtered to remove the large pieces
and carbared filtered to remove the chlorophyll. Aloe Liquid is pumped through state
of the art "high temperature/short term" heat exchange equipment for a rapid
pasteurization and quick cooling to prevent degradation from a slow heat and cool
down (batch method) stabilization procedure (http://www.aqaloe.com).
2.8.6 Steam distillation
Waller et al., (1978) had reported the steam distillation of Aloe vera leaves. In
this process, the tough outer portion consisting of the cuticle, epidermis, and
mesophyll was removed from the leaves of mature plants. The green outer portion
25
and the colorless inner part as well as the "stalk" i.e. portion of the plant above the
ground remaining after the leaves had been removed and the roots were steam
distilled, each separately. The distillation was allowed to proceed as long as the
condensate had a definite odor; this process takes approximately 5 h. The condensate
from the steam distillation was saturated with sodium chloride, extracted with diethyl
ether, dried with anhydrous sodium sulfate and the ether was reduced in volume
using a stream of nitrogen.
2.9 Commercial Processing
One of the earliest gel producers was Carrington Laboratories,
(http://www.carringtonlabs.com) has a range of products and patented the preparation
in the name of Acemannan or CarrisynTM. The well established (1973) firm is Terry
Laboratories (http://www. terrylabs. com.) and is the major supplier of gel to many
multinational companies. They are also the supporters of aloe research and quality
control. Aloe vera Company U.K. (Forever Living Products)
(http://www.aloevera.co.uk/) many companies like CRH International
(http;//www.aloealoe.com.) and Valley Aloe vera Inc. (http://www.quikpage.com),
Bonnsana Co. (www.bonasana.com) and Aloecorp USA (http://www. aloecorp.com)
have developed process for gel extraction and stabilization of gel and concentrated
gel either by air-drying or freeze-drying.
Basically, the ‘aloe dessert’s processing process is simple which involved
sorting, grading, washing, peeling, cutting, cooking in syrup, adding flavor, packing,
and pasteurizing. The most difficult part of processing is the removal of aloe and
retains its original taste and its marketability. Aloe is more popular as a material for
cosmetics produce purposes not as an edible produce. The process flow diagram of
Aloe vera dessert as suggested by Herlina, (2001) and later developed and modified
by PT. Niramas Utama Indonesia is shown in Figure 2.1. The flow chart of
commercialized processing of Aloe vera is shown in Figure 2.2.
26
Aloe vera leaves
Aloe vera meat flesh
Water
Sugar
Malic acid
Citric acid
Flavour agent
‘Aloe Vera’ Dessert
Figure 2.1 Processing process of ‘Aloe vera’ Dessert developed and modified
by PT. Niramas Utama Indonesia
Grading, trimming and washing
Peeling and separating
Cutting and grading
Cooking
Mixing
Packaging
Pasteurization
27
Fresh Aloe vera leaves
Tips and butts
Aloe oil
Cellulous pulp
Figure 2.2 Commercial Aloe vera processing flow chart (Aloe Vera -CRH
International, Inc http://www.aloealoe.com)
Whole leaf processing
Bulk Aloe pulp
Gel extraction
Aloe pulp Processing
Separation
Liquefaction
Concentration of Aloe juice to
32%
Solids
Reconstitution of 32 % concentrate into
lower %
Hand filleting
Filleting leaf
Decolorization with
charcoal
Washing
Soaking & Rinsing
Trimming
28
2.10 Processing of Aloe vera gel
During the past several decades several basic methods of processing Aloe vera
gel have been developed.
2.10.1 Gel extraction from Aloe vera pulp
Shafi et al., (2000) developed a commercially viable process for preparing a
stable and pharmacological active crystalline substance from the fresh whole leaf
meal and tested the product on experimental animals and volunteers for wound
healing remedy for all kinds of all damaged skin conditions.
Anon., (1967) had carried out the gel extraction from Aloe vera leaves,
remaining after the removal of its exudates were cut upon and its mucilage was
scraped out with blunt edged knife. This mucilage was stirred vigorously in a blender
to make it uniform. This solution was strained through a muslin cloth and filtered.
This uniform solution was extracted for cold- extracted gel (CEG) and hot extracted
gel (HEG).
Cold extracted gel (CEG)
This solution was acidified with Hydro chloric acid (HCL) having pH 3.50 and
the crude gel were precipitated out from the extract by adding slowly 95 % alcohol
while stirring. The gel was obtained by centrifugation.
Hot extracted gel (HEG)
Material left after passing the blended solution through muslin cloth, was
repeatedly treated with hot water until the complete extractions of gel was affected.
The crude gel (HEG) was prepared as described as above.
29
Waller et al., (1978) had reported the gel extraction process. As Aloe leaves
rapidly loose their medicinal properties, the material used was either fresh or
lyophilized and stored at –15
0
C. A. barbadensis leaves (35 g equivalent of dry
material) were macerated and extracted with water-acetone (1:1) and then acetone at
room temperature. The combined extracts (2:1) were concentrated in a rotary
evaporator (35
0
C) and the acetone free residue was extracted three times with
diethyl ether (250 m/each time).
Yaron, (1993) have extracted gel from full sized mature leaves and half size
young leaves picked from the same shrub. After removal of the ‘peel’ the colourless
hydroparenchyma was ground in a blender and centrifuged at 10,000 x g for 30 min
at 4
0
C to remove the fibers. Leaves weighing 800 g produced about 300 ml of gel.
Microbial growth was inhibited by addition of a preservative, 0.05 % sodium azide or
0.1 % Girgard (Givaudan).
2.10.2 Purification of Aloe vera gel
Anon., (1967) had reported the purification of crude gel. The crude gel
obtained by above method, was washed several times with ethanol till free from
chlorideions. It is also stirred with absolute acetone, ether and dried over anhydrous
CaCl
2
in vacuum desiccators. The dried gel was also greenish tinge of chlorophyll.
The gel was also de-proteinated by shaking the aqueous solution with chloroform.
Dry aloe gel was white amorphous powder when dried under reduced pressure in
vacuum desiccators.
2.10.3 Stabilization of Aloe vera gel
Many of the greatest benefits of Aloe vera can be lost in the processing unless
great care is taken to stabilize the gel. M/s Forever Living Products has developed a
stabilization method (http://www.aloevera.co.uk).
30
Yaron, (1992) had studied the fresh gel stabilization. Aloe vera gel, like most
natural juices, both fruit and vegetable, is an unstable product when extracted and is
subject to discoloration and spoilage from contamination by microorganisms.
Sulphated polysaccharides of the red microalga Porphyridium aerugineum were
obtained from the algal laboratories. An aqueous solution containing 2 % algal
polysaccharide solution and 50 % Aloe vera gel was prepared and its shear stress vs.
shear rate curve was generated. The solution was then stored at room temperature for
6 months for observations of the structure and homogeneity of the polysaccharides.
Addition of the algal sulphated polysaccharide resulted in a homogeneous stable
product: the algal polysaccharide may inhibit degradation and also browning of the
aloe polysaccharide.
2.11 Quality parameters for processing of Aloe vera
Time, temperature and sanitation (TTS) are necessary to preserve these
biological activities. The TTS Aloe Process not only preserves the natural biological
activities of Aloe vera but also enhances the physical stability of the finished
products. The TTS Aloe Process was developed with the idea of preserving the
natural state of the Aloe plant throughout each stage of production by maintaining
beneficial properties inherent in the plant. Data generated from these studies showed
that minor compositional changes, while not significant in the chemical makeup of
the product, resulted in great degrees of biological activity losses. These losses in
activity were not limited to one area, but different activities were affected at different
stages of production (http://www. aloecorp.com).
2.11.1 Timing of leaf process
Leaves show losses of biological activity beginning at six hours following
harvest when the leaves are stored at ambient temperature. Most biological activities
are completely lost after 24 hours at ambient temperatures. A decrease in activity is
31
also evident when the leaves are stored refrigerated even though the rate of activity
loss is greatly reduced. The losses of activity appear to be the result of enzymatic
activity after the leaf is removed from the plant. In fact, it has been shown that the
gel, once extracted from the leaf, has greater stability than gel, which is left in the
leaf. This means that shipping of leaves, even at refrigerated temperatures, will result
in loss of biological activity. The overall timing of TTS production phases is
extremely critical. The processing must be completed within 36 hours of harvesting
the leaves. (http://www. aloecorp.com)
2.11.2 Process Temperature
The Aloe gel processing temperature plays very important role for gel quality
for cosmetic and medicinal use.
A. Flash Cooling
As a crucial step to preserve biological activity, the gel is cooled to below 5
0
C
in ten to fifteen seconds following the gel extraction. Rapid cooling not only slows
enzymatic and microbial deterioration of the gel, but also aids in reducing the
microbial counts in the product.
B. Pasteurization
Biological activity remains essentially intact when the gel is heated at 65
0
C
for periods of less than fifteen minutes. Extended periods or higher temperatures will
result in greatly reduced activity levels. The best method of pasteurization is HTST
(high temperature short time), which exposes the gel to elevated temperatures for
periods of one to three minutes. Once heated, the gel is flashing cooled to 5
0
C or
below.
Ashleye, (1983) had revealed that heat during pasteurization was one of the
stresses imposed on the gel and there were advantages in using high temperatures for
short times preferably with addition of an antioxidant such as ascorbic acid.
32
C. Concentration
The gels obtained using the pasteurization and flash-cooling methods can be
concentrated under vacuum without the loss of biological activity. The concentration
operation must be conducted under 125 mm mercury vacuum at temperatures below
50
0
C and must not exceed two minutes. Higher vacuums and temperatures will cause
activity loss as will extend concentration times.
D. Freeze or Spray Drying
The concentrated product can then be freeze dried at temperatures between -
45
0
C and 30
0
C or can be spray dried with product temperatures below 60
0
C without
losses in biological activity.
E. Dehydration
Simal et al., (2000) had studied the effect of air-drying temperature (from 30
to 80
0
C) on dehydration curves and functional properties (water retention capacity,
WRC; swelling, SW; fat adsorption capacity, FAC) of Aloe vera cubes. The effective
diffusivities estimated with the proposed model varied with the air-drying
temperature according to the Arrhenius law except for the experiment carried out at
80
0
C, where casehardening took place. The three studied functional properties
exhibited a maximum when drying temperature was 40
0
C decreasing either at higher
or lower temperatures.
F. Blanching
Shih-Jen Chiou, (2003) in his investigation reported the nutrient contents of
Aloe vera and effect of additives on nonenzymatic browning of blanched Aloe vera
and studied the nutritive compositions of the Aloe vera and the effects of pH and
additives, on nonenzymatic browning of blanched Aloe vera during storage. In order
to establish an optimal blanching processing of Aloe vera, the activity changes of
peroxidase (POD) and polyphenol oxidase (PPO) of Aloe vera were also assayed
with different heating treatment.
33
2.12 Aloe vera Product Preparations
Choosing effective Aloe vera products can be challenging. Once a leaf is cut,
enzymes start to break down some of the long chain sugars which make Aloe vera gel
an effective healing product, so it is important for the plant to have been properly
handled and stabilized. Commercial, stabilized gel products may not work as well as
the fresh gel, but cold processing is thought to best retain the beneficial properties.
Aloe vera juice is most often the form of the gel that is used internally. The
nutritional composition of Aloe vera drink is shown in Table 2.9. A product that is
made from the whole leaf does not necessarily contain anthraquinones from the latex
layer, as those are water-soluble and can be separated out during processing.
Capsules and tinctures of the gel are also available. Oral forms of the latex extract are
generally capsules, as it is extremely bitter.
Salve: Remove the thin outer skin and process the leaves in a blender, add 500 units
of vitamin C powder to each cup and store in refrigerator.
Dried Juice: Aloe vera juice containing the equivalent of 360 - 900 mg of dried sap
is recommended by most herbalists per day.
Aloe tea and fibre tablets: Dry aloe leaves are harvested and crushed to form tea
leaves. A delicious herbal tea can be brewed. Tea leaves are pressed to form fibre
tablets.
34
Table 2.9 Aloe vera Drink's Nutrition fact
Nutrients Unit Thai
RDI* Composition of
Aloe vera
N (c) . 2 (11) 4 (15)
Ash Gram 0.5 0.2
Moisture (Water) Gram 84.4 88.3
Energy (Enerc) Kilo Calories 61 49
Protein (Procnt) Gram 50** 0
Fat Gram 65** 0.6
Total available CHO (Chocdf)
include FIBTG
Gram 300** 10.9
Dietary Fiber (Fibtg) Gram 25 0.2
Calcium (Ca) Milligram 800 31
Phosphorus (P) Milligram 800 3
Iron (Fe) Milligram 15 -
Sodium (Na) Milligram 2400 22
Potassium (K) Milligram 3500 12
Copper (Cu) Milligram 2 -
Zinc (Zn) Milligram 15 0.1
Vitamin A (Retinol) µgram . 0
β-Carotene (Cartb) µgram . -
Total vitamin A (Retinol-
Equivalent, RE)
µgram 800 -
Vitamin B1 (ThiA) Milligram 1.5 -
Vitamin B2 (Ribf) Milligram 1.7 -
Niacin (NIA) Milligram 20** -
Vitamin C (VitC) Milligram 60 -
* Percentage of Thai Recommended Daily Intake is based on a 2,000 kcal diet.
** %Energy distribution from protein, total fat and carbohydrate = 10:30:60, Total Saturated fat = 10% of
total energy.
Source: Thai Food Composition Tables (1999), Institute of Nutrition, Mahidol University (INMU).
35
2.13 Use of Aloe vera gel
Grindlay and Reynolds, (1986) in his review cited that the mucilaginous gel
from the parenchymatous cells in the leaf pulp of Aloe vera has been used since early
times for a host of curative purposes. This gel should be distinguished clearly from
the bitter yellow exudates originating from the bundle sheath cells, which is used for
its purgative effects. Aloe vera gel has come to play a prominent role as a
contemporary folk remedy. Modern clinical use of the gel began in the 1930s, with
reports of successful treatment of X-ray and radium burns, which led to further
experimental studies using laboratory animals in the following decades
However, over 95 % of the Aloes on the market today still use only the inner
gel and stabilize the Aloe in a high-heat process that degrades some of the enzymes,
polysaccharides and mucopolysaccharides. High heat (pasteurization and/or autoclave
methods) breaks down the constituents in Aloe vera that are the most valuable for
healing. Heat also kills the live enzymes necessary for digestion. Most Aloes are heat
processed.
Davis et al., (1989) Aloe vera preparations were evaluated for topical anti-
inflammatory activity using the croton oil-induced edema assay. Throughout
recorded history, it has been used to keep skin beautiful and restore it to health. A
frequent moisturizing ingredient in cosmetics and hair care products, it also promotes
the healing of burns and superficial wounds, but should not be used on deep or
surgical wounds of punctures. Topical application has been successful in treatment of
sunburn, frostbite, radiation injuries, some types of dermatitis, psoriasis, cuts, insect
stings, poison ivy, ulcerations, abrasions, and other dermatologic problems.
It also exerts antifungal and antibacterial effects, and thus helps to prevent
wound
infections
. One study showed it to have a little more activity than the antiseptic
silver sulfadiazine against a number of common bacteria that can infect the skin. It
36
has moisturizing and
pain
relieving properties for the skin lesions, in addition to
healing effects.
Aloe vera gel products may also be used internally. They should not contain
the laxative chemicals found in the latex layer. There is some evidence that Aloe vera
juice has a beneficial effect on peptic ulcers, perhaps inhibiting the causative bacteria,
Helicobacter pylori. It appears to have a soothing effect on the ulcer, and interferes
with the release of hydrochloric acid by the stomach. Colitis and other conditions of
the intestinal tract may also respond favorably to the internal use of gel products.
Aloe vera has been shown to exert a stabilizing effect on blood sugar in studies done
on mice, indicating a possible place for it in the treatment of diabetes. One study
suggested that giving Aloe vera extract orally to patients with asthma, oral Aloe vera
gel include prevention of kidney stones and relief of arthritis pain.
2.14 Organoleptic properties of Aloe vera gel
Aloe vera gel is viscous, colourless, odorless, taste slightly bitter. Gorloff,
(1983) had reported that in gel preparation processes, organoleptic properties are
important when the gel is intended for internal use.
It is evident from the review of various aspects of processing of aloe vera that
many stages subjective criteria during processing are followed. Therefore, there is a
scope to carry out the scientific investigations on processing of aloe vera and to
develop process and machineries for efficient aloe vera processing operations.
37
CHAPTER III
MATERIALS AND METHODS
This chapter deals with the selection of raw material, procedures followed for
determination of physical and chemical properties, experimental set-up, experimental
and analytical technique and quality evaluation used for optimisation of process
parameters for the extraction of gel from Aloe vera leaves.
3.1 Selection and procurement of Aloe vera leaves
The two years old and matured Aloe vera (Aloe barbadensis Miller) leaves
were selected for the experiment. The leaves were obtained from Department of
Botany, College of Agriculture, Junagadh Agricultural University, Junagadh. The
leaves of Aloe barbadensis variety were cut in the early morning every day for
experimentation. To avoid bio-degradation of Aloe vera leaves, each leaf is harvested
by hand with knife and pulled carefully from the mother plant so as not to break the
rind. The leaves were transported to the working place in a covered polyethylene bag
to avoid oxidation or contamination.
3.2 Physical properties and characteristics of Aloe vera leaf
The physical properties of Aloe vera leaf namely size, shape, test weight and
pulp weight were studied. The moisture content of the Aloe vera pulps and gel was
measured by hot oven drying method. The 10 g of Aloe vera pulp and gel sample
was kept in glass petri dish, dried at 105
0
C ± 2
0
C temperatures for 24 hours and its
weight determined for calculating of total solid (Wang and Strong, 1993). The
difference between initial weight and final oven dried weight of sample gives the
moisture content of the pulp and gel.
38
3.2.1 Size and shape
The maximum tri-axial dimension as shown in Figure 3.1 which were obtained
as length, width and thickness of Aloe vera leaves, were measured from 23 randomly
selected leaves using metallic tape having a least count of 1 mm. The average of
readings was taken as its length, width and thickness. The shape was determined by
comparing the longitudinal and lateral cross-section of the Aloe vera leaves.
Figure 3.1 Diagram of the approximate geometry of Aloe vera leaves and definition
of parameters for volume calculation.
(L = length; W = width; T = Thickness)
3.2.2 Apparent leaf volume
The apparent volume was calculated by considering the geometry of the object
similar to the geometrical shape. Knowing the values of length (L), width (W) and
thickness (T), the volume of the Aloe vera leaf was calculated. The leaf volume was
also calculated by considering the geometry as to be a cone with elliptical rather than
circular cross section as shown in Figure 3.1.
39
The volume was calculated by the formula given by Hernandez-Cruz et al.,
(2002) as below:
V = (L/12) π W T. -------------------------------(3.1)
Where;
V = Volume of the leaf, mm
3
L = Length of the leaf, mm
W = Width of the leaf, mm
T = Thickness of the leaf, mm
3.2.3 Leaf Weight
For determination of leaf weight, 23 nos. of randomly selected leaves were
weighed in a precision balance (Sartorious make, 0.01 g least count) and their
weights were recorded. The measurements of weights were replicated three times and
the average of the readings were recorded in g.
3.2.4 Fibre content
Crude gel is defined as the gel obtained after the centrifuge operation of Aloe
vera pulp, while pure gel is the gel obtained after purification of the crude gel. The
fibre content is defined as the difference between the dry weight of the crude gel and
that of the filtered gel (pure gel). It was measured by filtering the homogenate
through a 2.0 µm muslin cloth followed by Whatman No. 4 filter paper under
vacuum. Ten grams of the filtrate was placed in a dry glass petridish and dried at 105
0
C ± 2
0
C for 24 hours and its dry weight determined and the difference gives the
fibre content (Wang et al. 1993).
40
3.3 Laboratory scale gel extraction set up
The laboratory scale gel extraction experimental set up consists of Blender
used for making the fillets into pulp homogenate, test tubes with caps for keeping
extracted gel and, Buchner flask and Buchner funnel was attached to the vacuum
pump for filtration. The refrigerate box was used for handling the samples at low
temperature i.e. 4 to 5
0
C.
3.3.1 Centrifuge
Centrifuge is equipment, used to separate solid particles from liquid media,
based on size or shape with the action of centrifugal force. Basically, it consists of a
container fixed on the central axis, is rotated with the help of 1 hp electric motor. The
cooling type centrifuge (Remi Make C-24 Model) , having 31mm top radius and 37
mm bottom radius was used with maximum centrifugal force of 33500 g, at 20000
rpm, maximum rotor speed. The tube size was 100 ml and the maximum capacity of
the centrifuge was 400 ml.
Centrifugation is based on the fact that when an object is moving in a circle at a
steady angular velocity, is subject to an outward directed force F. The magnitude of
force, depends upon the angular velocity ω, radius of rotation r. Therefore the
centrifugal force is given by the following formula:
F = ω
2
r ......................(3.2)
Where;
F = Centrifugal force in
ω = Angular velocity, radians
r = Radius of rotation centimetres
41
The centrifugal force F is also expressed in terms of earth’s gravitational force
and it is then referred as the relative centrifugal force i.e. RCF, and commonly
expressed as the number “g”. The RCF is also expressed in form following
formula:(Thimmaiah, 1999).
RCF = ω
2
r/980 .....................(3.3)
ω
= π (rpm)/30
g = RCF = (1.119 X 10
-5
) ( rpm)
2
( r )
Where;
RCF = Relative centrifugal force
r = average radius of rotor, cm
3.3.2 Filtration unit
The filtration unit was consisted of vacuum pump, Buchner flask and Buchner
funnel. The crude gel, which was obtained after centrifuge operation, was mixed with
Charcoal for gel purification. The gel then was filtered in the filtration unit with the
help of Whatman No. 4 filter paper for further analysis.
3.3.3 Photospectrometer
Photospectrometer was used for measuring the optical density of extracted gel.
The Photospectrometer was calibrated before it was used for the experiment. The
procedure consists of keeping filter slot on number 6 and then zero transmittance was
set by lower knob. The Photospectrometer lower knob was kept remain in the same
position till all experiments were over. Initially the Photospectrometer was calibrated
with the help of distilled water by taking the optical density of distilled water as one.
The filter slot was then fixed according to wavelength of material i.e. for Aloe gel
400 nm and was set at 100% transmittance for blank sample or distilled water by
adjusting the side knobs.
42
3.4 Design of experiments
The experiments were planned using Four Factor Completely Randomised
Design. The treatments consisted of 2 levels of Acetone, 3 levels of centrifuge
temperature, 3 levels of centrifuge speed and 3 levels of centrifuge duration. The
independent and dependent variables considered in this study are given below.
Independent variables
a. Acetone : 2 i.e. Without addition of Acetone, and
10 % addition of Acetone
b. Centrifuge temperature : 3 i.e. 5, 10, and 32 (Ambient)
0
C
c. Centrifuge speed : 3 i.e. 2000, 5000 and 10,000 rpm
d. Centrifuge duration : 3 i.e. 10, 20 and 30 min
Dependent variables
a. Gel recovery, %
b. Viscosity of gel, Stokes
c. Refractive index of gel
d. Optical density of gel, abs
e. TSS content of gel, Brix
Experimental details
1. Treatments : Fifty four
2. Replications : Three
3. Total number of tests: Fifty four
4. Experimental design : Factorial Completely Randomised Design
In all 54 experiments were conducted and results obtained were analysed
statistically. The analysis of variance along with the level of significance was also
determined. Subsequently, the principal components analysis was carried out to get
the single optimum values of independent variables for the extraction gel from Aloe
vera leaves. The results are presented in Chapter IV.
43
3.5 Experimental procedure
After standardizing process parameter of the extraction of gel, following
experimental procedure was adopted to develop an appropriate process technology
for production of gel from Aloe vera barbadensis.
The freshly harvested leaves of Aloe barbadensis variety were cut manually in
the early morning for experimentation. To avoid bio-degradation the Aloe vera leaf
is harvested and pulled carefully from the mother plant so as not to break the rind and
immediately after cutting the leaves were kept in the icebox at 4 -5
0
C and
transported to the laboratory. The leaves were thoroughly washed with fresh water.
The outer skin and the exudates of the leaves were removed manually with the help
of knife to form fillet (Plate 3.6). The domestic blander (Boss Make) was used to
ground the fillets for one min at low speed (700 rpm) to obtain the crushed pulp
(Plate 3.7). The crushed pulp was again grounded at higher speed (1000 rpm) to
obtained homogenised pulp. The 60 ml pulp on volume basis was centrifuged in
cooling type centrifuge for separation of crude gel and fibre. The charcoal was mixed
with crude gel for purification. The vacuum filtration method was used to obtain pure
gel from crude gel. The pure gel was collected in the test tubes for further analysis.
3.6 Chemical properties of Aloe vera leaf
The pure gel was used to study the chemical properties of gel namely: pH,
TSS content, sugar content etc. The chemical composition of gel ultimately affects
the quality of gel. The quality of gel also affected by cultivation practices like number
of irrigation applied. The method of determination of the chemical properties is given
in the subsequent paragraphs.
44
3.6.1 Sugar content
For estimation of sugar content the sample was prepared by taking 0.1 g Aloe
vera pure gel in a 100 ml beaker, in which 5 ml of hot, 80 % ethanol was added. The
mixture was filtered (What-man No. 1 filter paper) into a beaker. The sample was
kept in hot water bath at 100
o
C so that sample became dry and to evaporate the
excess ethanol. Further 10 ml of distil water was added to each sample for
determination of sugar content. (Sadasivam and Manickam, 1996). The methods of
their estimation are given below.
3.6.1.1 Reducing sugar
Nelson-Somogiy method was used for estimation of reducing sugar. From the
collected aliquot, 0.2 ml sample was taken in a test tube by pipeting and final volume
was made up to 1 ml by adding distilled water. The test tubes were placed in hot
water bath for 10 minutes in boiling water after adding 1 ml Alkaline Copper Tartrate
reagent. The sample was kept for cooling at room temperature. 1 ml of
Arsenomolybdate reagent was added to the solution. The volume of the solution was
made up to 10 ml by adding 6 ml distilled water. The intensity of blue colour was
read at 620 nm on spectrophotometer. Standard graph was prepared using Glucose
(0-500 µg). The reducing sugar estimation was replicated twice and the average is
taken as the reducing sugar content of Aloe vera gel and it was calculated by equation
3.4.
Sample x Glucose x Volume
reading equivalent made up
Reducing sugar (%) = -------------------------------------------- x 100 x 10
-6
Weight of Aloe vera gel x aliquote taken
................(3.4)
45
3.6.1.2 Total sugar
The phenol sulphuric acid method given by Sadasivan et al., (1996) was used
to estimate total sugar. The filtrate obtained in the estimation of reducing sugar was
used in total sugar estimation. 0.2 ml aliquot was taken into test tube by pipeting and
final volume was made up to 2 ml by adding distilled water. 1 ml of 5 % phenol
solution and 5 ml of 96 % H
2
SO
4
was added quickly and allowed to stand for 10
minutes after mixing. The colour of solution was read at 490 nm on the
spectrophotometer. The estimation of total sugar was carried out using standard
graph prepared with glucose (0-150 µg) using following expression. The total sugar
estimation was replicated twice and the average is taken as the total sugar content of
Aloe vera gel and it was calculated by equation 3.5 given below.
Sample x Glucose x Volume
reading equivalent made up
Total sugar (%) = ------------------------------------------------ x 100 x 10
-6
Weight of Aloe vera gel x aliquote taken
................(3.5)
3.6.2 Total soluble solid test
The total soluble solids content is the summation of all the solids dissolved in
the water, beginning with sugar, salts, protein, acids, etc. The total solid contents in
the extracted gel affect its quality. More will be the total solids in the gel; poor will
be the quality of gel. Hand Refractometer was used for the measurement of total
soluble solid content. Hand Refractometer is a simple devise used for measuring
concentrations of aqueous solutions and it gives direct reading of total soluble solid
content in degree Brix. The % Brix scale expresses the concentration percentage of
the soluble solids content of a sample with water solution taken as reference
materials. The total soluble solid content was measured by placing two drops of the
Aloe vera gel on the scale of hand refractometer (0-32
0
C). The averages of three
readings were taken as TSS content of extracted gel and expressed in degree Brix in
percent.
46
3.6.3 Determination of pH of gel
The pH of a solution is the negative logarithm (base 10) of the activity or the
product of the molar concentration and the activity coefficient of the hydrogen ions
(H+) in the solution. pH of Aloe vera gel was measured by a recording type pH
meter. 30 ml of each sample was taken a test tube then probe was inserted in the test
tube and reading was taken as the pH of the each sample.
pH = - log [H
+
] ----------- (3.6)
3.7 Evaluation of gel quality parameters
Aloe gel quality was judged by its purity. Purity of gel was determined by the
viscosity, optical density, and refractive index.
3.7.1 Viscosity
The viscosity of a fluid is a very important property in the analysis of liquid
behavior and fluid motion near a solid boundary. Often it is defined as the resistance
to flow of a fluid. The resistance is caused by intermolecular friction exerted when
layers of fluids attempts to slide by another layer. It is measured in units of poises
(dyne-seconds per square centimeter), stokes or a subdivision of poises. The Oswald
viscometer and Redwood viscometer were used for the measurement of viscosity of
the Aloe vera gel.
3.7.1.1 Oswald type glass viscometer
Viscosity of the liquid materials is measured in a calibrated glass vessel known
as a viscometer. After calibration the Oswald viscometer was filled to the lower
calibration mark by applying suction with a rubber bulb and drawing the liquid
analyte into the apparatus. The time required for the volume of liquid between the
two marks to drain from the bulb is measured. The tube at the lower end of the upper
bulb has a fixed length and radius, which is used along with the pressure differential
column between the upper and lower ends of the apparatus to measure the viscosity.
47
3.7.1.2 Redwood viscometer
Redwood viscometer is based on the principle of laminar flow through
capillary tube of standard dimension under falling head. The Redwood No. 1
viscometer having capillary diameter.1.62 mm and length. 10.0 mm. The kinematic
viscosity ) of liquid and the time (t) required to pass 50cc of liquid are correlated
by the expression
µ = 0.0026 t – 1.175 / t --------( 3.7)
Where,
µ = Kinematic Viscosity in Stokes
t = time in seconds to collect 50 cc of oil.
The standard method was used in the measurement of viscosity of Aloe vera
gel as given by equation 3.7.
3.7.2 Optical density
Optical density is the measure of transparency of liquid and also a measure of
quality for Aloe vera gel. Photospectrometer was used for the measurement optical
density, which gives the direct reading of Absorbance. The filter slot No.6 of the
Photospectrometer was set on zero transmittance by lower knob and left as such with
out disturbing lower knob. The filter slot was also set according to wavelength (for
Aloe gel 400 nm) and set 100% transmittance for blank sample or distilled water by
side knobs.
48
3.7.3 Refractive index
Refractive Index is the ratio of the Sine of the angle of incidence to the Sine of
the angle of refraction, when a ray of light of monochromatic sodium light of
wavelength 589.3
0
A passes from (defined wavelength passes from) air into the
material keeping temperature as constant i.e. 27
0
C. The wavelength is 589.3 + nm
corresponding with D1 and D2 lines of sodium spectrum. The Refractometer used for
measurement of refractive index having range of Refractive Indices between 1.3000
and 1.7000 with an accuracy of + 0.0002. It was Calibrated with known refractive
indices i.e. doubled distilled water (1.3323) at 27
0
C ± .2
0
C.
Two drops of Aloe vera gel were placed on the Refractometer prism surface
and closed carefully. The mirror was adjusted until the reading was sharp. The
instrument was allowed to stand for a few minutes before the reading was taken so
that the sample and instrument came to equilibrium. The reading was taken when the
blue and yellow shade crossed the cross mark. The results were expressed in four
decimal places (Sangani, 1997).
Refractive index is the physical property of gel determines the purity of gel as
compared to double distilled water. Gel with lowest refractive index, is the best
treatment for extraction process. More refractive index indicates the impurities in the
extracted gel.
3.8 Development of Process
In development of process each step of Aloe vera gel extraction was
considered and standardised. Proportion of chemicals for addition determined. The
process time for each method is fixed. After standardising the process parameters, the
final gel extraction process was developed. The final process was evaluated for
quality parameters of extracted gel.
49
3.9 Statistical Analysis
The statistical analysis of experiment was carried out at Statistics Department,
College of Agriculture, Junagadh Agricultural University, Junagadh, with 4 Factors
Completely Randomized Design.
The data of results were interpreted. The main effect, interaction and combine
effect were studied. The F-test was carried out to determine whether the effect is
significant or not. Critical difference and Coefficient variation were considered for
the interpretation of data.
The test data of the studies conducted on different levels of acetone, centrifuge
temperature, centrifuge speed and centrifuge duration for different parameters such as
crude and pure gel recovery, viscosity, refractive index, optical density and TSS
content are presented in Appendix A F. The data of physical characteristics like
length, width, thickness, apparent volume, weight, pulp recovery of aloe vera leaves
and properties of Aloe vera gel such as m.c., fiber content, pH, sugar content were
analysed and the results are presented and discussed in Chapter IV.
50
CHAPTER IV
RESULTS AND DISCUSSION
This chapter deals with the results of the experiments conducted on physical
and chemical properties of Aloe vera leaf and Aloe vera gel, standardization of
process parameters, combine effect of process parameters on dependent variables like
recovery of gel, viscosity, refractive index, optical density and TSS etc. The
observations were analysed using Factorial CRD (Completely Randomised Design)
technique and the variation in the parameters was depicted graphically. The results of
the studies conducted are presented and discussed in the subsequent paragraphs.
4.1 Moisture content
The moisture content of the Aloe vera pulp and gel was measured by hot oven
drying method as described in Chapter III para No.3.2 The measurement of moisture
content was replicated 5 times and their average moisture content is presented in
Table 4.1. The average moisture content for pulp and gel were found to be 98.88 and
99.80 % respectively.
Table 4.1 Moisture content of Aloe vera leaf pulp
Sr. No. Moisture content (%)
Pulp Gel
Average 98.88 99.80
4.2 Physical characteristics and property of Aloe vera leaf
4.2.1 Size and shape
The size was determined from the measurement of length, width and thickness
of Aloe vera leaf. The results of variation in size of Aloe vera leaf are presented in
Table 4.2. The length, width and thickness are expressed in mm. From the Table 4.2,
51
it is clear that the length width and thickness ranges from 480 to 655, 55 to 115 and
19 to 32 mm, whereas, their averages are 55.98, 93.9 and 26.8 mm respectively.
Wang and Strong, (1993), has given similar results for length, width and thickness of
Aloe vera leaf. The average values and range of length, width and thickness also
decides size which taken as the apparent volume of Aloe vera leaf. The average size
is 371.75 cc. It can be noted from the table that the length is about 21 times to the
thickness. It is said to be, that, more is the thickness better will be the pulp recovery.
Therefore the thickness could be taken as one of selection criteria of Aloe vera leaf
for the extraction of gel for commercial use. The shape of Aloe vera leaf may be
classified as conic- tapered towards apex as per classification given by Mohsenin
(1980).
4.2.2 Apparent volume
Table 4.3 gives the variation in the apparent volume of Aloe vera leaf. The
average apparent volume is found to be 371.75 cc, whereas, it ranges from 127.32 to
485.73 cc. The relationship between apparent volume and fresh weight of leaves is
shown in Figure 4.1. Apparent volume can be taken as a parameter for the estimation
of leaf fresh weight. Taking apparent volume as a function of Aloe vera leaf weight
the following linear equation was developed.
Apparent volume = f (Aloe vera leaf weight)
y = 0.6527x + 215.87 -------------(4.1)
The response of the predicted values is within the experimental range.
4.2.3 Leaf weight
The leaf weight was measured with the help of pan balance having least count
0.01 g (Sartorious make). The leaf weight of 23 leaves were recorded and expressed
in g. Table 4.3 shows variation in leaf weight of Aloe vera leaf. The maximum
52
weight of the Aloe vera leaf was recorded to be 658 g, whereas, minimum was 326 g
with their standard deviation of 0.07. The average weight of the leaf was 459 g. It
was also observed during the experiment that the Aloe vera leaves having higher
weight, had recorded better pulp recovery.
Table 4.2 Variation in length, width, thickness and size of Aloe vera leaf
Average moisture content = 98.88 percent (wb)
Variety = Aloe barbadensis
Sr.
No Length
(a)
mm
Width
(b)
mm
Thickness
(c)
mm
Size
π/12(a x b x c)
cc
1 560 85 26.7 332.73
2 520 100 26.0 353.95
3 590 97.5 27.0 406.62
4 620 100 27.0 438.25
5 655 55.0 27.8 262.19
6 530 65.0 19.0 171.36
7 520 60.0 21.0 171.53
8 620 100.0 26.2 425.27
9 570 95.0 26.5 375.68
10 590 110.0 32.0 543.71
11 520 105.0 26.6 380.23
12 530 115.0 28.8 459.55
13 510 110.0 26.7 392.14
14 480 105.0 25.8 340.42
15 520 115.0 27.3 427.40
16 590 97.5 27.7 417.16
17 600 95.0 28.0 417.83
18 550 102.5 27.2 401.44
19 490 100.0 26.4 338.66
20 500 97.5 29.0 370.12
21 615 85.0 30.0 410.57
22 565 70.0 22.0 227.79
23 630 95.0 31.0 485.73
Average 55.98 93.9 26.8 371.75
Range 480 to 655 55 to 115 19 to 32 127.32 to 485.73
SD 4.97 1.67 0.29 91.91
53
4.2.4 Pulp weight
The pulp from Aloe vera leaves was extracted as per procedure given in Chapter
III section 3.5. The weight of pulp was measured after removing the skin; with the
help of pan balance-having least counts 0.01 g. Pulp was extracted from single leaf
each day and their pulp weight was recorded in g. Table 4.3 also presents the
variation in pulp weight of Aloe vera leaf.
Table 4.3 Variation in apparent volume, leaf weigh, pulp weight and pulp
recovery of Aloe vera leaf
Average moisture content = 98.88 percent (wb)
Variety = Aloe barbadensis
Sr.
No Apparent volume
cc Leaf weight
g Pulp weight
g Pulp recovery
%
1 332.73 431 192 44.55
2 353.95 397 170 42.82
3 406.62 458 205 44.76
4 438.25 445 194 43.60
5 262.19 481 232 48.23
6 171.36 326 156 47.85
7 171.53 336 156 46.43
8 425.27 424 186 43.87
9 375.68 454 226 49.78
10 543.71 658 331 50.30
11 380.23 454 232 51.10
12 459.55 523 266 50.86
13 392.14 452 243 53.76
14 340.42 397 217 54.66
15 427.40 456 232 50.88
16 417.16 476 241 50.63
17 417.83 515 275 53.40
18 401.44 448 256 57.14
19 338.66 440 229 52.05
20 370.12 509 267 52.46
21 410.57 560 312 55.71
22 227.79 360 192 53.33
23 485.73 546 288 52.75
Average
371.75 459 230 50.04
Range 127.32 to 485.73 326 to 658 156 to 331 42.82 to 57.14
SD 91.91 0.07 0.05 4.11
54
Figure 4. 1 Relationship between apparent volume and fresh weight of
Aloe vera leaves.
The maximum pulp weight of the Aloe vera leaf was recorded to be 331 g,
whereas, minimum was 156 g with their standard deviation of 0.046. The average
pulp weight of the leaf was 230 g. It was observed during the experiment that the
Aloe vera leaves having higher pulp weight, had recorded better gel recovery. Leaf
fresh weight can be taken as a parameter for the estimation of pulp recovery. The
y = 0.652x + 215.8
R² = 0.654
0
100
200
300
400
500
600
700
0.00 200.00 400.00 600.00
Leaf weight, g
Leaf volume, cc
55
relationship between leaf weight and pulp recovery is shown in Figure 4.2. Taking
leaf fresh weight as a function of pulp recovery the following linear equation was
developed.
Aloe vera leaf fresh weight = f (Aloe vera pulp recovery)
y = 0.0172x + 42.158 -------------(4.2)
The response of the predicted values is within the experimental range.
Figure 4.2 Relationship between fresh weights of Aloe vera leaves and pulp
recovery.
y = 0.017x + 42.15
R² = 0.096
30
35
40
45
50
55
60
300 400 500 600 700
Pulp recovery, %
Leaf weight,g
56
4.2.5 Pulp recovery
The variation in pulp recovery of Aloe vera leaf is presented in Table 4.3. The
maximum pulp recovery of the Aloe vera leaf was recorded to be 57.14 %, whereas,
minimum was 42.82 % with their standard deviation of 4.11 %. The average pulp
recovery from the leaf was 50. 04 %. It was observed during the experiment that the
Aloe vera leaves having higher pulp weight, had recorded better pulp recovery.
The relationship between apparent volume and pulp recovery is shown in
Figure 4.3. Apparent Volume can be taken as a parameter for the estimation of leaf
pulp recovery. Taking apparent volume as a function of Aloe vera leaf pulp recovery
the following linear equation was developed.
Apparent volume = f (Aloe vera pulp recovery)
y = 0.0067x + 47.561 -------------(4.3)
The response of the predicted values is within the experimental range.
4.3 Chemical property of Aloe vera gel
The chemical properties such as pH, total soluble solid, sugar content and fiber
content of extracted gel were determined. These chemical properties are required for
product preparation from gel. All these parameters are discussed in the subsequent
paragraphs.
Table 4.4 presents the chemical properties of Aloe vera gel. From the table it is
clear that the fiber content and pH of gel was found to be 0.2 % and 6.389 which is
closer to the findings of Wang et al., (1993) and sugar content 1.9125 % (Total sugar)
and 0.0259 % (Reducing sugar) from Aloe vera gel. M/s Delta International has
found Aloe vera gel pH 3.5 - 4.7 (http://www.shantidatta.com).
57
Figure 4.3 Relationship between apparent volume and pulp recovery of Aloe
vera leaf
Table 4.4 Chemical properties of Aloe vera gel
Sr.
No.
TSS
(Brix)
pH Fiber content
(%)
Sugar
(%)
Pulp Gel Total Reducing
Average 1.366 6.398 1.117 0.2 1.9125 0.0259
Range 0.77
to
1.97
5.88
to
7.02
1.114
to
1.120
0.18
to
0.22
1.53
to
2.295
0.024
to
0.0278
y = 0.0067x + 47.561
R2= 0.0223
0.00
10.00
20.00
30.00
40.00
50.00
60.00
0.00 200.00 400.00 600.00
Pulp recovery, %
Leaf volume, cc
58
4.4 Standardization of process parameters for extraction of gel from Aloe
vera leaves
The process parameters like acetone proportion, centrifuge temperature;
centrifuge speed and centrifuge duration for extraction process was studied. The
effect of process parameters on different quality parameters such as crude and pure
gel recovery, viscosity, refractive index, optical density and TSS content of gel
extracted from Aloe vera leaf were recorded.
4.4.1 Effect of acetone on gel extraction process
The effect of acetone on different quality parameters such as crude and pure
gel recovery, viscosity, refractive index, optical density and TSS content of gel
extracted from Aloe vera leaf is presented in Table 4.5 and also shown in form of bar
diagram in Figures 4.4 and 4.5. The statistical analysis is given in Appendix A F.
From the table it is clear that all the parameters increase with 10 % addition of
acetone to Aloe vera pulp. The refractive index was found to be little higher in case
of 10 % of acetone as compared to, without acetone in Aloe vera pulp. The distilled
water was taken as reference material with the consideration that, it is one of the
purest forms of liquid. The both treatment i. e, without and with 10 % acetone shows
the little higher values of refractive index i.e. 1.33601 and 1.33610 respectively when
compared to distilled water which is having only 1.3323 at 27
0
C ± 0.2
0
C. It may be
said that, the values of the refractive index closer to water is considered to be the best
in terms of their purity. The refractive index of the extracted gel, without acetone had
found to be closer to distilled water. It may be concluded that refractive index of the
Aloe vera gel without acetone is to be considered as purest forms of gel.
It is also clear from the Figure 4.5 that, addition of 10 % acetone to pulp
increases the crude gel recovery by 2.8 %, pure gel recovery by 2.3 % and viscosity
by 136.5 % (Figure 4.5). It indicates favorable results because more will be the
59
viscosity of the gel active will be the biological material, but, at the same time the
percent increase in refractive index by 0.0067 %, optical density 12.93 % and TSS
93.55 % (Figure 4.5) which shows un-favorable results for the extraction process
because the values of the indices is little higher as compared to distilled water which
has been taken as measurement of the purity of gel.
Table 4.5 Effect of acetone on gel extraction process
Treatments
Dependent Variables
Acetone
Proportion
(%)
Crude gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes)
Refractive
index Optical
density
(abs)
TSS
(Brix)
0 57.32 40.93 0.591 1.33601 0.232 0.93
10 59.24 43.23 1.398 1.33610 0.252 1.80
S.Em. 0.171 0.131 0.004 2.15 x 10
-
5
2.8 x 10
-
4
0.01
CD @ 5 % 0.479 0.368 0.011 6.0 x 10
-
5
0.01 0.027
Test Sig. Sig. Sig. Sig. Sig. Sig.
The statistical analysis shows the significant effect of acetone on different
quality parameters such as crude and pure gel recovery, viscosity, refractive index,
optical density and TSS at 5 % Cd.
Based on the results it may be concluded that the effect of 10 % acetone
addition to pulp for gel extraction, results in increase in the gel recovery and
viscosity, but at the same time it deteriorate the purity of gel. Hence the use of
addition of acetone may be avoided.
60
Figure 4.4 Effect of Acetone on Aloe gel recovery
Figure 4.5 Effect of acetone on quality parameters of Aloe gel
57.32 59.24
40.93 43.23
0
10
20
30
40
50
60
70
0 % acetone
10 % acetone
Gel recovery (%)
Proportion of Acetone (%)
Crude gel recovery (%)
Pure gel recovery (%)
0.591
1.398
0.232 0.252
0.93
1.8
1.33601 1.3361
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
1.8
2
0 % acetone
10 % acetone
Values of quality parameters
Proportion of Acetone (%)
Viscosity (Stokes)
Optical density (abs)
TSS (Brix)
Refractive index
61
4.4.2 Effect of centrifuge temperature on gel extraction process
Temperature is considered, as one of the major factor, which ultimately
affects the viscosity, optical density, refractive index and TSS content of gel
extracted from Aloe vera leaves. Danhof, (2000) has reported that the sun or heat
exposure and also the time of processing of Aloe vera leaves should be minimum.
The effect of centrifuge temperature on different quality parameters such as crude
and pure gel recovery, viscosity, refractive index, optical density and TSS content on
gel extraction from Aloe vera leaf at different temperatures are presented in Table
4.6, and also given in form of bar diagram in Figure 4.6 and 4.7. The statistical
analysis is given in Appendix A to F. The percentage recovery of crude and pure gel
varies from 57.91 to 58.89 and 42.04 to 42.14 % respectively with the variation in
temperature, which is also represented graphically in Figure 4.6. It is seen from the
table that the percent recovery of gel is more or less same in both the case with the
increase of temperature. This shows that there is no effect of temperature on the
recovery of gel either crude or pure.
The viscosity of the extracted gel (Table 4.6) is largely affected with the
changes of centrifuge temperature. The maximum viscosity (1.369 Stokes) was
recorded at 5
0
C and minimum (0.696 Stokes) at 32
0
C (Ambient temperature).
Viscosity was recorded 96.69 and 49.13 % higher at 5
0
C as compared to 32
0
C and
10
0
C temperature. This statement is satisfied with the study conducted by Shih-Jen
Chiou, (2003) that the viscosity of Aloe vera gel decreases with the increase of
heating time. It is obvious that higher is the value of viscosity of Aloe vera gel, better
will be the product.
62
Figure 4.6 Effect of centrifuge temperature on Aloe gel recovery
Figure 4.7 Effect of centrifuge temperature on quality parameters of Aloe
gel
57.91 58.04 58.89
42.14 42.04 42.06
0
10
20
30
40
50
60
70
5 10 32
Values of quality parameters
Centrifuge temperature (C)
Crude gel recovery (%)
Pure gel recovery (%)
1.369
0.918
0.696
0.238 0.243 0.244
1.4 1.39 1.31
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
5
10
32
Values of quality parameters
Centrifuge temperature (C)
Viscosity(Stokes)
Optical density (abs)
TSS (Brix)
Refractive index
63
The Refractive index and TSS increases with the increase of temperature
(Table 4.6). The minimum and maximum refractive index was found to 1.33603 and
1.33610, while TSS was found 1.31 and 1.40 Brix respectively. The refractive index
of the pure gel is found to be closer to distilled water at all the temperatures. It may
be said that the temperature has non-significant effect on the purities of the gel. The
TSS content decreases with the increase of temperature. The decrease in TSS content
with temperature may be due to enzymatic degradation of Aloe vera gel at higher
temperatures.
Table 4.6 Effect of centrifuge temperatures on gel extraction process
Treatments Dependent Variables
Centrifuge
Temperature
(
0
C)
Crude gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes)
Refractive
index Optical
density
(abs)
TSS
(Brix)
5 57.91 42.14 1.369 1.33610 0.238 1.40
10 58.04 42.04 0.918 1.33604 0.243 1.39
32 58.89 42.06 0.696 1.33603 0.244 1.31
S.Em. 0.213 0.164 0.005 2.68x10
-
5
3.51x10
-
4
0.012
CD @ 5 % 0.597 0.459 0.014 7.5x10
-
5
0.001 0.034
Test Sig. NS Sig. NS Sig. Sig.
From the Table 4.6 it is seen that the optical density of the Aloe vera gel
increases with the increase of temperature. The maximum value was found to be
0.244 at 32
0
C and minimum at 5
0
C temperatures. The increase in the value of
optical density may be due to enzymatic degradation of Aloe vera gel at higher
temperatures. Gel extracted at different centrifuge temperatures is shown in Plate 4.1.
The statistical analysis shows the significant effect of centrifuge temperature
on different quality parameters such as crude gel recovery, viscosity, refractive index,
64
optical density and TSS, whereas for pure gel recovery it shows non-significant effect
at 5 % Cd.
It is concluded that 5
0
C temperature yield in higher viscosity, lower refractive
index and optical density with optimum recovery of gel. It may be suggested that the
gel extraction may be carried out by Centrifuge at 5
0
C temperature to obtained
better quality gel.
4.4.3 Effect of centrifuge speed on gel extraction process
The centrifuge process is carried out to separate solid particles from pulp for
getting crude gel. The Aloe vera gel also contains sugar molecules surrounded by gel
molecules. Centrifugal force is required to break this chain of sugar molecules. Table
4.7 shows the results of effect of centrifuge speed on different quality parameters
such as gel recovery, viscosity, optical density, refractive index and TSS content of
gel extracted from Aloe vera leaves. Also it is represented graphically in Figure 4.8
and 4.9 and statistical analysis is given in Appendix A to F.
It is seen from the table that, the crude and pure gel recovery, viscosity, and
TSS content increases with the increase of centrifuge speed, where as other parameter
i.e. optical density, refractive index decreases with the increase of centrifugal speed.
The maximum recovery of crude gel as well as pure gel is found to be 67.05 and
48.72 % at 10000 rpm respectively. The recovery for crude gel as well as pure gel at
2000 and 5000 centrifuge speed is 17.98 and 26.74 % and 8.33 and 14.14 %
respectively lower, when compared to 10,000 rpm centrifuge speed. The higher
recovery at higher speed i.e.10,000 rpm may be due the separation of solid particles
from pulp or break down of the chain of sugar molecules.
The viscosity and TSS are found maximum at 10,000 rpm i.e. 1.040 strokes
and 1.40 Brix respectively where as minimum values at 2000 centrifuge speed for
65
both the cases. It is said that, higher is the viscosity better will be the quality of the
product. At the same time the product is considered to be biologically active. (Gowda
et al., 1979). The maximum value of TSS at higher speed is found to be 1.40 Brix.
This TSS value is much closer to the finding of Wang et al., (1993). Similarly the
value of refractive index (1.33789 - 1.34390) at 10,000 rpm is match with the finding
of the M/s. Delta International (http://www.shantidatta.com). The lower is the optical
density of gel better will be the product. The 10,000 rpm centrifuge speed shows the
lowest value i.e., 0.234 abs and highest at 2000 rpm speed 0.250 abs. It may be
concluded that after considering all the quality parameters the 10,000 rpm centrifuge
speed are found better for the gel extraction process. Extracted gel sample is shown in
Plate 4.2 for different centrifuge speed.
Table 4.7 Effect of centrifuge speed on gel extraction process
Treatments Dependent Variables
Centrifuge
speed
(rpm)
Crude
gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes)
Refractive
index Optical
density
(abs)
TSS
(Brix)
2000 49.07 35.69 0.954 1.33689 0.250 1.33
5000 58.72 41.83 0.989 1.33599 0.242 1.37
10,000 67.05 48.72 1.040 1.33530 0.234 1.40
S.Em. 0.213 0.164 0.005 2.68x10
-
5
3.51x10
-
4
0.012
CD @ 5 % 0.597 0.459 0.014 7.5x10
-
5
0.001 0.034
Test Sig. Sig. Sig. Sig. Sig. Sig.
66
Figure 4.8 Effect of Centrifuge speed on Aloe gel recovery
Figure 4.9 Effect of Centrifuge speed on quality parameters of Aloe gel
49.07
58.72
67.05
35.69 41.83 48.72
0
10
20
30
40
50
60
70
80
2000
5000
10000
Gel recovery (%)
Centrifuge speed (rpm)
Crude gel recovery (%)
Pure gel recovery(%)
0.954 0.989 1.04
0.25 0.242 0.234
1.33 1.37 1.4
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
2000
5000
10000
Values of quality parameters
Centrifuge speed, (rpm)
Viscosity (Stokes)
Optical density (abs)
TSS (Brix)
Refractive index
67
The statistical analysis shows that all the quality parameters are found to be
significant at 5 % Cd.
From the above results it is suggested that higher centrifuge speed is found
suitable for all quality parameters of gel extraction except viscosity. Hence the gel
extraction by centrifuge may be carried out at 10,000 rpm speed.
4.4.4 Effect of centrifuge duration on gel extraction process
In gel extraction process, the duration of centrifuge plays an important role in
the separation of solid particles from the crude pulp. The results of effect of
centrifuge duration on different quality parameters such as gel recovery, viscosity,
optical density, refractive index and TSS content of gel extracted from Aloe vera
leaves are shown in Figure 4.10 and 4.11 and presented in Table 4.8. The statistical
analysis is given in Appendix A to F. It is seen from the Table 4.8 that, the crude and
pure gel recovery increases with the increase of centrifuge duration, where as optical
density decreases with the increase of centrifuge duration. It is also seen from the
table that centrifuge duration has very minor effect on the remaining parameters such
as viscosity, refractive index and TSS.
The maximum recovery of crude gel as well as pure gel is found to be 61.91
and 45.13 % at 30 min respectively. The recovery for crude gel as well as pure gel at
10 and 20 min centrifuge duration is 12.21 and 5.37 % and 13.12 and 7.16 %
respectively lower, when compared to 30 min centrifuge duration. Higher recovery of
the gel at 30 min duration may be due the fact that all the particles are getting
sufficient residual time during the extraction process. Therefore it may be concluded
that sufficient time should be given to obtain optimum gel recovery from the crude
pulp.
68
Viscosity as well as refractive index is higher at 10 min duration and minimum
at 30 min duration. Their values are of 1.008 and 0.987 Stokes and, 1.33634 and
1.33576 at 10 and 30 min duration respectively. M/s Aloecorp, (2003) has also
reported values for refractive index. Optical density of gel decreases as the duration
of centrifuge increase. It was found lowest with 30 min duration i.e. 0.239 and
highest with 10 min duration i.e. 0.245 abs. Lower values of optical density indicate
that the product content less amount of fibrous materials. The TSS content increases
with increase in centrifuge duration. It was found 1.35, 1.38 and 1.36 Brix with 10,
20 and 30 min duration respectively. Looking to the recovery of crude and pure gel
extraction and other quality parameters, the 30 min centrifuge duration is found to be
the best for gel extraction from pulp. Plate 4.3 shows the effect of different centrifuge
duration on gel extraction process.
Table 4.8 Effect of centrifuge duration on gel extraction process
Treatments
Dependent Variables
Centrifuge
duration
(min)
Crude gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes)
Refractive
index Optical
density
(abs)
TSS
(Brix)
10 54.35 39.21 1.008 1.33634 0.245 1.35
20 58.58 41.90 0.988 1.33607 0.242 1.38
30 61.91 45.13 0.987 1.33576 0.239 1.36
S.Em. 0.213 0.164 0.005 2.68x10
-
5
3.51x 10
-
4
0.012
CD @ 5 % 0.597 0.459 0.014 7.5x10
-
5
0.001 0.034
Test Sig. Sig. Sig. Sig. Sig. NS
69
Figure 4.10 Effect of Centrifuge duration of Aloe gel recovery
Figure 4.11 Effect of Centrifuge duration on quality parameters of Aloe gel
54.35 58.58 61.91
39.21 41.9 45.13
0
10
20
30
40
50
60
70
10
20
30
Gel recovery (%)
Centrifuge duration (min)
Crude gel
recovery (%)
pure gel recovery
(%)
1.008 0.988 0.987
0.245 0.242 0.239
1.35 1.38 1.36
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
10
20
30
Values of quality parameters
Centrifuge duration (min)
Viscosity (Stokes)
Optical density (abs)
TSS (Brix)
Refractive index
70
The statistical analysis shows that the effect of Centrifuge duration on
different quality parameters such as crude and pure gel recovery, viscosity, refractive
index and optical density are found to be significant, whereas for TSS it is non-
significant. It is concluded that 30 min centrifuge duration may be considered
optimum for gel extraction process to obtained better quality gel. The quality of gel in
terms of viscosity is slightly affected by higher residual time.
4.5 Combine effect of two process parameters on gel extraction process
4.5.1 Effect of acetone and centrifuge temperature on gel extraction process
Table 4.9 presents the combine effect of acetone and centrifuge temperature on
different quality parameters and their statistical analysis are given in Appendix A to
F. The combine effect of acetone and centrifuge temperature on the crude and pure
gel recovery, viscosity, optical density and TSS are found to be significant whereas
refractive index is found to be non significant at 5 % Cd.
4.5.2 Combine effect of acetone and centrifuge speed on gel extraction process
The combine effect of acetone and centrifuge speed on various dependent
variables was studied and given in Table 4.10 and also given in Appendix A to F. The
statistical analysis shows significant effect of acetone and centrifuge speed on crude
and pure gel recovery, viscosity, optical density and TSS and non significant on
refractive index at 5 % Cd.
71
Table 4.9 Combine effect of acetone and centrifuge temperature on gel
extraction process
Treatments Dependent Variables
Acetone
CFT
(
0
C)
Crude
gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes) Refractive
index Optical
density
(abs)
TSS
(Brix)
Without
Acetone
5 56.19 40.43 0.646 1.33603 0.225 0.92
10 57.18 40.86 0.586 1.33600 0.233 0.92
32
58.58 41.50 0.542 1.33602 0.237 0.96
With
10 %
Acetone
5 59.63 43.84 2.092 1.33618 0.251 1.89
10 58.90 43.23 1.250 1.33608 0.253 1.86
32
59.20 42.63 0.850 1.33604 0.252 1.66
Sem 0.301 0.231 0.007 3.79 x 10
-
5
4.96 x 10
-
4
0.017
CD at 5 % 0.844 0.649 0.020 1.02 x 10
-
4
0.001 0.048
Test Sig. Sig. Sig. NS Sig. Sig.
CFT = Centrifuge temperature
Table 4.10 Combine effect of acetone and centrifuge speed on gel extraction process
Treatments Dependent Variables
Acetone
CFS
(rpm) Crude
gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes) Refractive
index Optical
density
(abs)
TSS
(Brix)
Without
Acetone
2000 47.56 35.15 0.584 1.33648 0.241 0.91
5000 57.99 40.40 0.590 1.33596 0.231 0.93
10000 66.41 47.25 0.600 1.33560 0.223 0.95
With
Acetone
(10 %)
2000 50.59 36.23 1.324 1.33729 0.259 1.75
5000 59.45 43.27 1.389 1.33601 0.252 1.81
10,000 67.70 50.19 1.480 1.33500 0.245 1.84
S.Em. 0.301 0.231 0.007 3.79x10
-
5
4.96x10
-
4
0.017
CD @ 5 % 0.844 0.649 0.020 1.02x10
-
4
0.001 0.048
Test Sig. Sig. Sig. NS Sig. Sig.
CFS = Centrifuge speed
72
4.5.3 Combine effect of acetone and centrifuge duration on gel extraction
process
Table 4.11 present the combine effect of acetone and centrifuge duration on
different quality parameters and their statistical analysis are given in Appendix A to
F. The interaction of acetone and centrifuge duration on refractive index is found to
be non significant and for other parameters it is found to be significant at 5 % Cd.
Table 4.11 Combine effect of acetone and centrifuge duration on gel extraction
process
Treatments Dependent Variables
Acetone
CFD
(min)
Crude
gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes)
Refractive
index
Optical
density
(abs)
TSS
(Brix)
Without
Acetone
10 52.83 37.57 0.589 1.33627 0.236 0.92
20 58.15 41.09 0.593 1.33598 0.232 0.92
30 60.98 44.14 0.592 1.33579 0.228 0.96
With
Acetone
(10 %)
10 55.88 40.86 1.427 1.33641 0.255 1.89
20 59.01 42.72 1.382 1.33616 0.252 1.86
30 62.85 46.12 1.384 1.33573 0.249 1.66
S.Em. 0.301 0.231 0.007 3.79 x 10
-
5
4.96 x 10
-
4
0.017
CD @ 5 % 0.844 0.649 0.020 1.02 x 10
-
4
0.001 0.048
Test Sig. Sig. Sig. NS Sig. Sig.
CFD = Centrifuge duration
4.5.4 Combine effect of centrifuge temperature and speed on gel extraction
process
For gel extraction from Aloe vera leaf, different centrifuge temperature and
speed combinations were studied and are given in Table 4.12 and Appendix A to F.
73
The interaction of centrifuge temperature and speed on all quality parameters are
found to be significant except TSS, which shows non-significant effect.
Table 4.12 Combine effect of centrifuge temperature and speed on gel extraction
process
Treatments Dependent Variables
CFT
(
0
C)
CFS
(rpm)
Crude
gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes)
Refractive
index
Optical
density
(abs)
TSS
(Brix)
5 2000 49.11 35.26 1.309 1.33685 0.246 1.36
5000 57.34 41.93 1.346 1.33614 0.238 1.42
10,000 67.28 49.22 1.452 1.33531 0.231 1.43
10 2000 48.64 35.83 0.893 1.33698 0.250 1.36
5000 59.03 41.73 0.927 1.33591 0.243 1.37
10,000 66.45 48.56 0.934 1.33523 0.237 1.43
32 2000 49.46 35.98 0.660 1.33683 0.254 1.28
5000 59.79 41.84 0.694 1.33591 0.245 1.32
10,000 67.43 48.37 0.734 1.33535 0.234 1.32
S.Em. 0.368 0.283 0.009 4.6 x 10
-
5
6.1 x10
-
5
0.215
CD @ 5 % 1.034 0..795 0.025 1.3 x10
-
4
0.002 0.059
Test Sig. Sig. Sig. Sig. Sig. Ns
CFT = Centrifuge temperature CFS = Centrifuge speed
4.5.5 Combine effect of centrifuge temperature and duration on gel extraction
process
The statistical analysis shows that the interaction between centrifuge
temperature and duration on the crude and pure gel recovery, viscosity and refractive
index and are found to be significant whereas on optical density and TSS it is non
significant (Table 4.13 and Appendix A to F).
74
Table 4.13 Combine effect of centrifuge temperature and centrifuge duration on gel
extraction process
Treatments Dependent Variables
CFT
(
0
C)
CFD
(min)
Crude gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes) Refractive
index Optical
density
(abs)
TSS
(Brix)
5 10 53.23 38.94 1.410 1.33647 0.242 1.37
20 58.15 42.41 1.350 1.33609 0.238 1.41
30 62.35 45.06 1.347 1.33574 0.234 1.43
10 10 54.37 39.38 0.927 1.33622 0.247 1.38
20 58.09 41.51 0.922 1.33609 0.243 1.40
30 61.66 45.24 0.906 1.33581 0.240 1.38
32 10 55.45 39.31 0.688 1.33633 0.247 1.31
20 59.50 41.79 0.690 1.33602 0.245 1.34
30 61.72 45.09 0.710 1.33573 0.242 1.28
S.Em. 0.368 0.283 0.009 4.6 x 10-5 6.1 x10-5 0.215
CD @ 5 % 1.034 0..795 0.025 1.3 x10-4 0.002 0.059
Test Sig. Sig. Sig. Sig. NS Ns
CFT= Centrifuge temperature, CFD= Centrifuge duration
4.5.6 Combine effect of centrifuge speed and duration on gel extraction
process
The combine effect of centrifuge speed and duration on gel extraction process
was studied on various dependent variables and found to significant except viscosity
which shows non significant interaction between two (Table 4.14 and Appendix A to
F).
75
Table 4.14 Combine effect of centrifuge speed and duration on gel extraction
process
Treatments Dependent Variables
CFS
(rpm)
CFD
(min)
Crude
gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes)
Refractive
index
Optical
density
(abs)
TSS
(Brix)
2000 10 45.84 32.88 0.958 1.33736 0.254 1.37
20 48.73 35.32 0.959 1.33679 0.250 1.33
30 52.64 38.87 0.946 1.33651 0.247 1.30
5000 10 53.06 38.45 1.007 1.33612 0.246 1.31
20 59.82 42.07 0.976 1.33603 0.242 1.40
30 63.28 44.97 0.984 1.33581 0.238 1.40
10000
10 64.16 46.31 1.059 1.33554 0.236 1.38
20 67.19 48.31 1.027 1.33538 0.234 1.42
30 69.81 51.55 1.033 1.33497 0.231 1.39
S.Em. 0.368 0.283 0.009 4.6 x 10-5 6.1 x10-5 0.215
CD @ 5 % 1.034 0.795 0.025 1.3 x10-4 0.002 0.059
Test Sig. Sig. NS Sig. Sig. Sig.
CFS= Centrifuge speed, CFD= Centrifuge duration
4.6 Combine effect of three process parameters on gel extraction process
4.6.1 Combine effect of acetone, centrifuge temperature and speed on gel
extraction process
The three factors as acetone, centrifuge temperature and speed on varying
proportions were studied and the resultant effect was recorded. Table 4.15 presents
the combine effect of acetone; centrifuge speed and duration on different quality
76
parameters and their statistical analysis are given in Appendix A to F. The statistical
analysis shows that the combine effect of acetone; centrifuge speed and duration for
all quality parameters are found to be significant.
4.6.2 Combine effect of acetone, centrifuge temperature and duration on gel
extraction process
The three factors as acetone, centrifuge temperature and duration on varying
proportions were studied and the resultant effect was recorded. Table 4.16 presents
the combine effect of acetone; centrifuge temperature and centrifuge duration on
different quality parameters and their statistical analysis are given in Appendix A to
F. The statistical analysis shows that the combine effect of acetone; centrifuge
temperature and duration on the pure gel recovery, viscosity and refractive index, are
found to be significant, whereas on crude gel optical density and TSS the combine
effect was non significant.
4.6.3 Combine effect of Acetone, centrifuge speed and duration on gel
extraction process
The three factors as acetone, centrifuge speed and duration on varying
proportions were studied and the resultant effect was recorded. Table 4.17 presents
the combine effect of acetone; centrifuge speed and centrifuge duration on different
quality parameters and their statistical analysis are given in Appendix A to F. The
statistical analysis shows that the combine effect of acetone; centrifuge temperature
and centrifuge duration on, crude gel recovery, refractive index and optical density
are found to be significant, whereas on the pure gel recovery, viscosity and TSS the
combine effect was non significant.
77
Table 4.15 Combine effect of acetone, centrifuge temperature and speed on gel
extraction process.
CFT= Centrifuge temperature, CFS= Centrifuge speed
CFT= Centrifuge temperature, CFS= Centrifuge speed
Treatments Dependent Variables
Acetone CFT
(
0
C)
CFS
(rpm)
Crude gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes)
Refractive
index
Optical
density
(abs)
TSS
(Brix)
Without
Acetone
5 2000 46.96 33.96 0.633 1.33653 0.233 0.84
5000 54.78 39.78 0.648 1.33594 0.224 0.96
10,000 66.83 47.56 0.656 1.33560 0.219 0.96
10 2000 46.70 34.74 0.586 1.33640 0.240 0.96
5000 59.15 40.63 0.581 1.33603 0.232 0.88
10,000 65.69 47.20 0.591 1.33556 0.228 0.93
32 2000 49.00 36.74 0.533 1.33651 0.249 0.94
5000 60.04 40.78 0.540 1.33590 0.237 0.97
10,000 66.70 46.98 0.552 1.33564 0.223 0.96
With
10 %
Acetone
5 2000 51.26 36.56 1.985 1.33717 0.259 1.87
5000 59.91 44.07 2.045 1.33634 0.251 1.88
10000 67.72 50.89 2.247 1.33502 0.243 1.91
10 2000 50.57 36.93 1.200 1.33756 0.260 1.77
5000 58.91 42.83 1.274 1.33578 0.254 1.87
10,000 67.22 49.93 1.277 1.33491 0.245 1.93
32 2000 49.93 35.22 0.788 1.33714 0.259 1.61
5000 59.54 42.91 0.847 1.33592 0.252 1.68
10,000 68.15 49.76 0.916 1.33506 0.245 1.69
S.Em. 0.521 0.401 0.013 6.6x10
-
5
8.6x10
-
5
0.030
CD @ 5 % 1.462 1.124 0.035 1.8x10
-
4
0.002 0.084
Test Sig. Sig. Sig. Sig. Sig. Sig.
78
Table 4.16 Combine effect of acetone, centrifuge temperature and duration on gel
extraction process
Treatments Dependent Variables
Acetone
CFT
(
0
C)
CFD
(min)
Crude gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes)
Refractive
index
Optical
density
(abs)
TSS
(Brix)
Without
acetone
5 10 50.70 36.22 0.640 1.33639 0.231 0.89
20 56.72 41.33 0.650 1.33591 0.225 0.92
30 61.15 43.74 0.647 1.33578 0.220 0.94
10 10 53.13 38.02 0.590 1.33617 0.238 0.92
20 57.94 40.78 0.586 1.33600 0.233 0.94
30 60.46 43.78 0.582 1.33582 0.229 0.90
32 10 54.65 38.46 0.536 1.33624 0.239 0.97
20 59.78 41.15 0.543 1.33603 0.237 0.99
30 61.31 44.89 0.546 1.33578 0.234 0.91
With
10 %
acetone
5 10 55.76 41.67 2.179 1.33656 0.254 1.84
20 59.57 43.48 2.051 1.33628 0.251 1.89
30 63.56 46.37 2.047 1.33570 0.248 1.92
10 10 55.61 40.74 1.263 1.33627 0.256 1.84
20 58.24 42.24 1.257 1.33618 0.252 1.86
30 62.85 46.70 1.230 1.33580 0.251 1.87
32 10 56.26 40.17 0.839 1.33642 0.255 1.64
20 59.22 42.43 0.837 1.33601 0.252 1.69
30 62.13 45.30 0.874 1.33569 0.249 1.64
S.Em. 0.521 0.401 0.013 6.6x10
-
5
8.6x10
-
5
0.030
CD @ 5 % 1.462 1.124 0.035 1.8 x10
-
4
0.002 0.084
Test NS Sig. Sig. Sig. NS NS
CFT= Centrifuge temperature, CFD= Centrifuge duration
79
Table 4.17 Combine effect of acetone, centrifuge speed and duration on gel
extraction process
Treatments Dependent Variables
Acetone
CFS
(rpm)
CFD
(min)
Crude gel
recovery
(%)
Pure gel
recovery
(%)
Viscosity
(Stokes)
Refractive
index
Optical
density
(abs)
TSS
(Brix)
Without
acetone
2000 10 44.02 31.89 0.580 1.33702 0.246 0.94
20 47.89 35.20 0.594 1.33630 0.241 0.92
30 50.76 38.35 0.578 1.33612 0.237 0.88
5000 10 50.44 36.39 0.595 1.33604 0.237 0.91
20 60.61 40.81 0.588 1.33600 0.231 0.96
30 62.91 43.98 0.586 1.33583 0.225 0.93
10000 10 64.02 44.43 0.591 1.33573 0.225 0.92
20 65.94 47.24 0.597 1.33564 0.224 0.98
30 69.26 50.07 0.611 1.33542 0.221 0.94
With
10 %
acetone
2000 10 47.67 33.87 1.336 1.33769 0.262 1.79
20 49.57 35.44 1.323 1.33728 0.258 1.73
30 54.52 39.39 1.313 1.33690 0.257 1.72
5000 10 55.67 40.52 1.420 1.33620 0.255 1.71
20 59.04 43.33 1.364 1.33607 0.252 1.84
30 63.65 45.96 1.382 1.33578 0.250 1.87
10000 10 64.30 48.19 1.526 1.33536 0.248 1.83
20 68.43 49.37 1.458 1.33512 0.245 1.86
30 70.37 53.02 1.455 1.33451 0.242 1.84
S.Em. 0.521 0.401 0.013 6.6x10
-
5
8.6x10
-
5
0.030
CD @ 5 % 1.462 1.124 0.035 1.8 x10
-
4
0.002 0.084
Test Sig. NS NS Sig. Sig. NS
CFS= Centrifuge speed, CFD= Centrifuge duration
4.6.4 Combine effect of centrifuge temperature, speed and duration on gel
extraction process
The three factors as centrifuge temperature; speed and duration on varying
proportions were studied and the resultant effect was recorded. The three factors
interaction of centrifuge temperature; speed and duration was tested for F value.
80
4.6.4.1 Crude gel recovery
Table 4.18 presents the combine effect of centrifuge temperature; speed and
duration on crude gel recovery and their statistical analysis are given in Appendix A.
The statistical analysis shows that the combine effect of centrifuge temperature;
speed and duration on, crude gel recovery is found to be significant.
Table 4.18 Combine effect of centrifuge temperature, speed and duration on crude
gel recovery (%)
4.6.4.2 Pure gel recovery
Table 4.19 presents the combine effect of centrifuge temperature; speed and
duration on pure gel recovery and their statistical analysis are given in Appendix B.
The statistical analysis shows that the combine effect of centrifuge temperature;
speed and duration on, pure gel recovery, is found to be non significant.
Centrifuge
temperature
(
0
C)
Centrifuge
speed
(rpm)
Centrifuge duration (min)
10 20 30
Crude gel recovery (%)
5 2000 45.33 47.81 54.19
5000 50.69 58.39 62.94
10000 63.67 68.25 69.92
10 2000 45.78 48.53 51.61
5000 53.36 59.42 64.31
10000 63.97 66.33 69.06
32 2000 46.42 49.86 52.11
5000 55.11 61.67 62.58
10000 64.83 66.97 70.47
S.Em. 0.638
CD @ 5 % 1.790
Test Sig.
81
Table 4.19 Combine effect of centrifuge temperature, speed and duration on pure gel
recovery (%)
4.6.4.3 Viscosity of gel
Table 4.20 presents the combine effect of centrifuge temperature; speed and
duration on viscosity of gel and their statistical analysis are given in Appendix C.
The statistical analysis shows that the combine effect of centrifuge temperature;
speed and duration on, viscosity of gel is found to be significant.
Centrifuge
temperature
(
0
C)
Centrifuge
speed
(rpm)
Centrifuge duration (min)
10 20 30
Pure gel recovery (%)
5 2000 32.11 35.61 38.06
5000 37.81 42.81 45.17
10000 46.92 48.81 51.94
10 2000 33.53 34.86 39.11
5000 38.56 41.61 45.03
10000 46.06 48.06 51.58
32 2000 33.00 35.50 39.44
5000 39.00 41.81 44.72
10000 45.94 48.06 51.11
S.Em. 0.491
CD @ 5 % 1.375
Test NS
82
4.6.4.4 Refractive index of gel
Table 4.21 presents the combine effect of centrifuge temperature; speed and
duration on refractive index of gel and their statistical analysis are given in
Appendix D. The statistical analysis shows that the combine effect of centrifuge
temperature; speed and duration on, refractive index of gel, is found to be
significant.
Table 4.20 Combine effect of centrifuge temperature, speed and duration on
viscosity of gel (Stokes)
Centrifuge
temperature
(
0
C)
Centrifuge
speed
(rpm)
Centrifuge duration (min)
10 20 30
Viscosity of gel (Stokes)
5 2000 1.367 1.327 1.233
5000 1.367 1.297 1.375
10000 1.495 1.427 1.434
10 2000 0.877 0.896 0.905
5000 0.954 0.939 0.889
10000 0.949 0.930 0.924
32 2000 0.629 0.653 0.699
5000 0.701 0.692 0.688
10000 0.733 0.726 0.743
S.Em. 0.150
CD @ 5 % 0.043
Test Sig.
83
4.6.4.5 Optical density of gel
Table 4.22 presents the combine effect of centrifuge temperature; speed and
duration on optical density of gel and their statistical analysis are given in Appendix
E. The statistical analysis shows that the combine effect of centrifuge temperature;
speed and duration on, optical density of gel, is found to be non-significant.