Title: Estimation of starch, dextran,

phosphate, silica, alanine, glycine, suplhur

and oxalic acid in cane juice using near
infrared spectroscopy

By: Aamarpali Ratna Puri

e-mail: aamarpali1@rediffmail.com
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 Sugarcane impurities studied.
• Sugarcane along with sugar contains numerous
other dissolved substances like reducing sugars,
organic matter other than sugar, inorganic
compounds (phosphates, chlorides, sulphates,
nitrates and silicates) and nitrogenous bodies
(albuminoids, amides and amino acids).
• Starch, dextran, phosphate, silica, alanine and
glycine are common impurities found in sugarcane.
In present research these are taken into
consideration. Lets have an idea about them in
sugarcane.
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 STARCH AND DEXTRAN
 Starch is natural constituent of cane juice and dextran enters in
cane juice during mechanical harvesting.
 Starch gets gelatinised by heating during juice clarification and
is removed to an extent of 30-35 % but the rest gets concentrated
in the process stream due to evaporation of the clear juice. The
harmful effects of it are the increase in viscosity and poor juice
filterability.
 Dextran formation causes sugar loss processing problems and
increases the viscosity manifold. This increase in the viscosity
decreases the clarification rate and gives poor clarification,
which in turn leads to crystal elongation.
 Thus quantitative estimation of starch and dextran in factories is
very essential before starting the process of clarification of cane
juice.

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 Phosphate
1. Phosphate is present in the form of
soluble phosphates in cane juice. The
phosphates are used to remove the
harmful action of iron, this has
beneficial effect upon the quality of
K the resulting white sugar.
O O 2. The optimum requirement of P2O5 is
300-500mg per liter of juice in the
H form of soluble phosphate.
P 3. Presence of phosphate in cane juice in
H optimum concentration is essential for
good clarification. Higher or lower
O O percentage of phosphate content in the
raw juice is equally undesirable.
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 Silica
• Cane juice is saturated with
respect to silica.
• Silica creates industrial
problems by the formation of
scales during sugar
crystallisation. Silica gets
deposited as calcium silicate
(CaSiO3) scales.
• Scales are hard deposits,
which stick very firmly to the
inner surface of boiler. They
cannot be removed even with
the help of hammer and chisel.

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 Amino acids in sugarcane
• The requirement of nitrogen in plants is dependent on the climatic
conditions, rainfall, drainage, and length of growing cycle and varietal
capacity for uptake of nitrogen. Most of nitrogenous compounds are found
in the leaves and shoot of the sugarcane plant.
• Amino acid and amides are present in raw cane juice and molasses. The
free Amino acids accumulate in molasses and contribute to sucrose loss
when amino acids are linked together to form proteins.
• Cane juice contains nitrogenous bodies such as albuminoids, ammonia,
amino acids (alanine and glycine) and amides varying from 0.5 to 1.0%.
• Amino acids are of importance as they along with other nitrogenous
bodies reacts with reducing sugars and form colored compounds.
• Also it is found from the static and dynamic analysis that different
colorants such as caramel, an iron-phenolic body complex, and the
browning reaction products from reducing sugars and amino acids, have
different affinities for the crystals and cause different coloration under the
same conditions.
• In industry, so many efforts are done to remove color. Analyzing amino
acids in a sample can help solving color problem in sugar to some extent.
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 Alanine
• Alanine (Molecular formula
C3H7NO2) is the major amino
acid present in cane juice. It is
hydrophobic, with a methyl
group side chain, and is the
second-smallest of the 20 after
glycine.
• As percentage dry solids, the
alanine is 0.06% free and is
0.05% as protein.
• It is an inhibitory or calming
neurotransmitter in brain.
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 Glycine
• Glycine (Molecular formula
C2H5NO2) which is the other
major amino acid present in
cane juice is polar,
uncharged, neutral and
genetically coded amino
acid.
• As percentage dry solids, the
glycine is 0.01% free and
0.04% as protein in cane
juice.

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 Sulfur
• Sulfur is a contaminant, which enters
sugar during refining.
• Sulfur in foodstuffs is found to be
linked to colon rectal cancer.
• Sulfur is believed to destroy Vitamin
A, as it promotes the oxidation of the
conjugated double bonds, and is also
known to destroy Vitamin B1.
• Sulfur and sulfur dioxide are also
known to promote allergies and
accelerate the onset of allergy attacks.

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 Oxalic acid
Oxalic acid and oxalates are mild
nephrotoxic acids that are
abundantly present in many plants

Oxalic acid binds vital nutrients

such as calcium, so long-term
consumption of foods high in
oxalic acid can lead to nutrient
deficiencies.

It is one of the major organic acid

Chemical structure of Oxalic acid.
present in sugarcane so its study in
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sugarcane is very important.
 Justification of the problem
 This research will lead to a set and definite technique to
produce quality sugar instead of following indefinite process
leading to increase in costs of the product.
 Undertaking NIR spectroscopic estimations of impurities in
cane juice will enable the sugar industry to check the
contents of cane juice sample before hand. The best-suited
sample will be selected and other one rejected without any
delay. This selection of sample will not only save time but
also reduce damages caused by the usage of improper cane
juice sample in sugar industry.
 Also Near-Infrared Spectroscopic estimation is an eco
friendly method, as it needs no hazardous chemicals.

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 Near-Infrared Spectroscopic
Estimations of impurities in cane juice

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The striking features of the Near-Infrared
technology are as follows:

Designed for all cane and beet juice, syrups and

molasses.
– It is environment friendly since no chemical reagents
are used for the tests.
– Easy to use, thus permitting analysis of clear to turbid
liquids without clarification or filtration.
– Frequency of response is online and continuous.
– Single and multiple-constituents analysis possible. it
has no consumables and operational costs are low, after
purchasing the information technology equipment and
the package there are no other costs involved.
12/27/19 05:07 AM 13
 Picture showing NIR Details:

  = 0.8m  = 2.5m  = 25m  = 400m


  = 12500 cm-1  = 4000 cm-1  = 400 cm-1  = 25 cm-1

 NIR MIR FIR

 IR region

  : wavelength
  : wavenumber
 NIR : near infrared
 MIR : mid infrared
 FIR : far infrared

12/27/19 05:07 AM 14
Importance of Near IR Spectroscopy.
 This region is of great  The estimation is not
importance in food time consuming and
Industry. does not involve the
 The sample can be use of Hazardous
scanned in minutes to chemicals.
know the concentration  The testing of Pol, Brix
of constituents and composition of a
(polysaccharides, silica, given sample without
phosphate and amino clarification can be
acids) in food products. done.
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 INSTRUMENT USED:
USED

• Near-Infrared spectrophotometer of Elico(India): Spectral

range 600-2500nm, bandwidth 10nm, accuracy +0.5nm,
repeatability +0.2nm and with advanced state of art MS
Windows® based software for data acquistion was used.

• The basic principle of the instrument is, that Near-Infrared

radiation when incident on a sample gets transmitted. In
transmission mode, the incident rediations get transmitted
through the sample with diminished energy. The amount of
transmitted energy from the sample is a measure of
concentration of the constituent molecules in the sample.

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Instrumental set up and method of measurement
For off line laboratory, the following set up can be done:

 NIR Source Monochromator sample cell Detector

 Source driver Current Read- out


 Amplifier Display


 Personal
 Computer

• Light Source is a 50 watt tungsten halogen lamp.

• For sugar syrup/juice i.e for liquid sample, the transmission mode of measurement
can be carried out.
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 Materials:

• Pure sucrose and Standard Starch (E.Merck),

Dextran (Himedia) Potassium dihydrogen
phosphate of Ranbaxy, Sodium meta silicate
(CDH), -Alanine of E-Merck (India) Ltd, glycine
of S. D. fine-chem Ltd and fresh cane juice of
local varieties were used.
• The initial concentration of starch, dextran,
phosphate, silica, alanine and glycine was found to
be 250, 42.1, 580, 250, 90 and 37ppm
respectively.

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 Method:
• Separate calibration models were prepared for these impurities in
cane juice (starch, dextran, phosphate, silica, alanine and glycine)
using PLSR. For setting calibration models, thirty samples each, for
these impurities were scanned in transmittance mode. Each
calibration file was having a particular concentration range i.e. for
starch 250-650 ppm, dextran 40-565 ppm, phosphate 200-800 ppm,
silica 150-720 ppm, alanine 5-150 ppm and glycine 2-60 ppm.
• Separate prediction files were prepared for starch, dextran,
phosphate, silica, alanine and glycine in cane juice. For the
preparation of each prediction file twenty samples of cane juice
having varying concentrations of the respective impurity were
scanned and their prediction was done using their respective
calibration models set up earlier.

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 RESULTS AND DISCUSSION

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 Partial Least - Squares Regression (PLS)
• This procedure involves simultaneously breaking
down the absorbance and concentration analytical
information derived from the calibration set.
• The statistical parameter used to select wavelengths
ensuring good fitting was the standard error of
calibration (SEC) for predicted samples used in the
calibration set and multiple correlation coefficient
‘R’. The SEC is given by:
 SEC = (Cexp. - CNIR)2
 n-m-1

– Where n is the number of samples used for calibration,

– m that of independent variables.
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Table.1
Summary statistics for the PLS calibration of phosphate, silica,

alanine and glycine (individually) from 30 samples of cane juice.

.

• S.No Analyte Factors Range of values Calibration

in cane juice (ppm) SEC MR

1. phosphate 9 200-800 0.003 1.000

2. silica 5 150-720 0.024 1.000
3. alanine 5 5-150 0.001 1.000
4. glycine 6 2-60 0.002 1.000

• Where: SEC = Standard Error of Calibration, MR =Multiple Regression

coefficient

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Table.2
Summary statistics for the prediction of phosphate, silica, alanine

and glycine (individually) from 20 samples of cane juice.

S.No Analyte SEP R

in cane juice

1. phosphate 0.615 0.999

2. silica 1.210 0.999
3. alanine 0.417 0.999
4. glycine 0.594 0.999

• Where: SEP = Standard Error of Prediction, R =correlation coefficient

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 800
700
600
500
actual
values 400
(ppm)
300
200
100
0
0 200 400 600 800
NIR predicted values (ppm)
.

Figure .1 Plot of actual values (ppm) vs. NIR

predicted values (ppm) of phosphate in cane juice.
12/27/19 05:07 AM 24
 800
700
600

actual 500
values 400
(ppm)
300
200
100
0
0 100 200 300 400 500 600 700 800

NIR predicted values (ppm)

Figure .2 Plot of actual values (ppm) vs. NIR

predicted values (ppm) of silica in cane juice.
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 160

140

120
actual 100
values
(ppm) 80

60

40

20

0
0 20 40 60 80 100 120 140 160

NIR predicted values (ppm)

Figure .3 Plot of actual values (ppm) vs. NIR

predicted values (ppm) of alanine in cane juice.
12/27/19 05:07 AM 26
 70

60

50

actual 40
values
(ppm) 30

20

10

0
0 20 40 60 80
NIR predicted values (ppm)

Figure .4 Plot of actual values (ppm) vs. NIR

predicted values (ppm) of glycine in cane juice.

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 Stepwise Multiple Linear Regression (SMLR)
– The SMLR procedure is based on the relationship between the
concentration and absorbance values obtained at several
selected wavelengths.
– In the SMLR wavelengths are selected by first choosing the
wavelengths which is most highly correlated with the
concentration and then adding terms, one at a time, until no
significantly improved standard error is obtained.

  = (Cexp. - CNIR)2
 p-1

 Where : p = number of selected wavelengths

  =Standard Error of Prediction

12/27/19 05:07 AM 28
Table.3
Regression coefficients and Standard Deviation () data for
phosphate, silica, alanine and glycine using Stepwise
Multivariate Linear Regression (SMLR) analysis.

S.No Analyte k0 k1 k2 k3 

1. phosphate 1066.17 -1081.83 -1525.84 1383.38 55.78

2. silica 1350.92 -1782.27 290.55 __ 35.29
3. alanine 175.48 -79.00 -84.48 18.97 14.12
4. glycine 86.73 -97.29 -23.39 -53.25 5.44

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Table.4

Wavelengths identified from Stepwise Multivariate Linear Regression

analysis that respond well to impurity under consideration.

Analyte Wavelengths (nm)

phosphate 1592 2275 2428

silica 943 1912 …..
alanine 1910 2037 2366
glycine 940 1590 2416

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 CONCLUSIONS:
• In the calibration models set up by the Partial Least Square
Regression analysis, SEC was found to be negligible which
confirms the reliability of calibration models set up for
phosphate, silica, alanine and glycine in cane juice.
• The SEP was found to be very less with best correlation in case
of all the four analytes, so NIR spectroscopic method can be
successfully used for accurate prediction of the concentration of
unknown samples of cane juice in few seconds.
• Stepwise Multivariate Linear Regression analysis was also
applied and with it the wavelengths affected by the content of
particular impurity in cane juice were selected.
• The partial least square analysis was found to give better results
than stepwise multi linear regression analysis.

12/27/19 05:07 AM 31
 References:
• Bassett, J.; Denny, R.C.; Jeffery, G.H. and Mendham, J. (1978). “Vogel’s textbook of quantitative inorganic analysis”. 4 th Edn.
ELBS with Longman. India., 757-758.

• Fiedler, F.M.; Edye, L.A and Watson, L.J. (2001). “The application of discriminant analysis to online near infrared spectroscopy
of prepared sugar cane”. Proc. Aust. Sug. Cane Tech., 23, 317-321.

• Lanza, E and Li, B.W. (1984). “Application of near Infrared spectroscopy for predicting the sugar content of fruit juices”, J.
Food. Sci., 49, 995-998.

• Mathur, R.L. (1986). “Handbook of cane sugar technology IInd Edition”. Oxford IBH Publishing Company., India, 26.

• Osborne, B.G.; Fearn, T and Hindle, P.H. (1993). “Practical spectroscopy with application in food and beverage analysis”. UK,
Longman Scientific & Technical.

• Salgo, A.; Nagy, J and Miko, E. (1998). “Application of near infrared spectroscopy in the sugar industry”. J. NIR. Spectrosc., 6,
101-106.

• Solid, H and Solberg, C. (1992). “Salmon fat content estimation by Near-Infrared transmission spectroscopy”. J Food Sci., 57,
792-93.

• Verma, N.C. (1988). “System of technical control for cane sugar factories in India”. Sug. Tech. Assoc. India., 34 -35.

• Vogel, A.I. (1992). “Elementary Practical Organic Chemistry Part-III”. Williams Clowes & Ltd, Eng. Language. Book. Soc.
Longman gp Ltd., 708-709.

12/27/19 05:07 AM 32
 List of publications
S.Kaur and Aamarpali. (2004) Quantitative estimation of
phosphate in cane juice using near infrared
Spectrophotometer, Indian Sugar, Vol (LIV) page 513-517.
Oct.
Aamarpali Ratna Puri and S. Kaur. (2006) Estimation of
silica in cane juice using near infrared spectroscopy,
Cooperative sugar, Vol (37): No 6 page 29-33, Feb.
Aamarpali Ratna Puri, Estimation of Alanine and
Glycine in Cane Juice Using Near Infrared Spectroscopy,
International Journal of food Engineering, accepted, in
press.

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12/27/19 05:07 AM Glycine microscopic 35