Separation of health compounds in Goji (Lyceum
barbarum) aqueous extracts by membrane technology
Carmela Conidi, Alfredo Cassano, Enrico Drioli
Institute on Membrane Technology, ITM-CNR,
c/o University of Calabria, via P. Bucci, 17/C
I-87030 Rende (CS), Italy
Phone: +39 0984 492067-492014
Fax: +39 0984 402103
E-mail: [email protected]
2nd International Conference and Expo on
Separation Techniques
September 26 - 28, 2016 Valencia, Spain
Outline
Goji berries and leaves: state of the art and health properties
Membrane processes for the treatment of Goji berries and
leaves aqueous extracts
Clarification of Goji aqueous extracts
Fractionation of clarified extracts
Analysis of membrane productivity
Analysis of membrane selectivity (antioxidant activity, total
polyphenols, soluble solids, total carbohydrates)
Conclusions
State of art
Goji berries are one of the top 10
“Super Food” cultivated in the world
Popular in traditional Chinese
medicine as well as Western parts of
the world
The plant is also cultivated in the
Mediterranean region (goji berries
grow very well in regions with semiarid
to
sub-tropical
climatic
conditions)
Global market of gojy extracts
Health benefits
H. Amagase , N. R. Farnsworth. A review of botanical characteristics, phytochemistry, clinical relevance in efficacy and safety of Lycium barbarum fruit (Goji), Food Research
International 44 (2011) 1702–1717
Bioactive compounds
b-carotene
Polyphenols
Ascorbic acid
Polysaccarides
D. Donno, G.L. Beccaro, M.G. Mellano, A.K. Cerutti, G. Bounous. Goji berry fruit (Lycium spp.): antioxidant compound fingerprint and bioactivity evaluation, Journal Functional Foods,
2015, 1070-1085
Recovery of bioactive compounds from vegetable sources
 Ultrasound assisted extraction
 Solvent extraction
Resin-based extraction
Supercritical fluid extraction
LIMITATIONS
more expensive than aqueous processing
residual solvents in the extracts (no
desiderable for food use)
easy denaturation of thermo-sensitive
compounds
reduction of nutritional value
 Ultrafiltration
Nanofiltration
Reverse Osmosis
ADVANTAGES
low energy requirement
additive free (do not require the use of
organic solvents)
mild operating conditions
preservation of thermo-sensitive compounds
Clarification/Fractionation with membranes processes
Driving force = DP
Retentate
Permeate
Pressure-driven membrane processes
Process
Feed
Membrane
DP range
Species retained
MF
Porous
0.1 – 5 mm
0.1 – 3 bar
Cells, bacteria,
yeasts, starch
granules, oil globules,
etc.
UF
Porous
5 – 100 nm
1 – 10 bar
Polysaccharides,
proteins, tannins,
virus, etc.
NF
Porous
1 – 5 nm
10 – 50 bar
Sugars, organic acids,
polyphenols, aroma
compounds, etc.
RO
Non porous
10 – 100 bar
Salts
Goji berries extracts / flow sheet of the investigated process
Goji berries
Aqueous extract
Washing
Sieving
(200 mm)
Milling
CLARIFICATION
(UF)
Retentate
b-carotene
Clarified extract
Extraction with water
(200 g/kg, 80 °C, 30 min)
FRACTIONATION
(UF/NF 0.5-2 kDa)
Separation of carbohydrates from polyphenols
Clarification of goji berries extracts
Manufacturer
Toray
China Blue Star Membrane
Technology Co. Ltd.
Membrane type
HFS
DCQ II-006 C-PS100
Hollow fiber
Hollow fiber
Polyvinylidene fluoride
Polysulphone
Max oper. pressure (bar)
2
1.5
Max oper. temperature (°C)
40
50
pH range
2-12
2-13
Membrane surf. area (m2)
0.4
0.16
0.02 mm
-
-
100
Configuration
Membrane material
Dimension of pores
MWCO (kDa)
Toray HFS
UF PS DCQ
Clarification of goji berries extracts
FEED
100
Permeate
Retentate
PS
PVDF
80
Jp(Kg/m2h)
60
40
20
Feed
0
1
2
3
4
5
VRF
Ultrafiltration of goji berries aqueous extracts. Permeate flux vs. VRF
(Operating conditions: TMP, 0.7 bar; Qf, 280 L/h; T, 24±2°C)
Permeate
Retentate
Clarification of goji berries extracts
PS membrane
Parameters
FEED
PERMEATE
RETENTATE
Total suspended solids (%)
3.2±1.1
-
Total soluble solids (°Brix)
4.5±0.4
4.0±0.2
b-carotene (mg/ml)
2.80±0.5
0.260±0.05
520.5±2.6
510.0±6.0
540.0±8.0
3.5±0.5
2.6±0.2
3.65±0.12
Total polyphenols (mg/L gallic acid)
TAA (mM Trolox)
Total carbohydrates (g/L glucose)
15.1±2.3
14.35±1.52
3.8±0.2
4.5±1.2
4.74 ±0.25
18.2±2.3
Clarification of goji berries extracts
PVDF membrane
Parameters
FEED
PERMEATE
RETENTATE
Total suspended solids (%)
3.1±0.2
-
3.8±0.4
Total soluble solids (°Brix)
4.2±0.1
4.1±0.2
4.2±0.1
b-carotene (mg/ml)
3.16±0.12
0.098±0.02
4.34±0.35
Total polyphenols (mg/L gallic acid)
448.5±2.6
400±6.0
480±6.0
TAA (mM Trolox)
2.8±0.3
2.15±0.25
2.6±0.4
Total carbohydrates (g/L glucose)
17.7±3.2
13.5±2.6
18.67 ±2.4
Clarification of goji berries extracts
Retention= (1-Cp/Cf)*100
120
PVDF
PS
100
Retention (%)
80
60
40
20
at
es
S
ar
bo
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TS
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A
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To
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To
To
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0
Retention of UF membranes towards analyzed compounds
Fractionation of clarified goji berries extracts
Clarified goji berries aqueous extracts
Membrane type
Desal GE
Desal GH
Desal GK
Manufacturer
GE Water & Process
GE Water & Process
GE Water & Process
Membrane material
thin film composite
with a cross-linked
aromatic polyamide
top layer
Molecular weight cut-off (Da)
500
1000
2000
Max. operating pressure (bar)
27
27
27
Max. operating temperature (°C)
90
90
90
Range of pH
1-11
1-11
1-11
thin film composite
with a cross-linked
aromatic polyamide
top layer
thin film composite
with a cross-linked
aromatic polyamide
top layer
Fractionation of clarified goji berries extracts
Steady state permeate flux (kg/m2h)
160
Desal GK
Desal GH
Desal GE
140
120
100
80
60
40
20
0
0
5
10
15
20
25
TMP (bar)
Treatment of clarified Goji aqueous extracts with UF and NF membranes.
Evolution of permeate flux as function of TMP
(Operating conditions: Qf, 300, L/h; T, 24±2°C)
Fractionation of clarified goji berries extracts
80
100
GK
GH
GE
GK
GH
GE
80
TAA retention (%)
Total polyphebnols retention (%)
100
60
40
20
60
40
20
0
0
5
10
15
20
5
10
TMP (bar)
15
20
TMP (bar)
Retention of tested membrane towards total polyphenols
Retention of tested membrane towards TAA
100
100
GK
GH
GE
Total carbohydrates retention (%)
TSS retention (%)
80
60
40
20
80
GK
GH
GE
60
40
20
0
5
10
15
20
TMP (bar)
0
5
10
15
20
TMP (bar)
Retention of tested membrane towards TSS
Retention of tested membrane towards total carbohydrates
Fractionation of clarified goji berries extracts
100
100
TSS
Total carbohydrates
Total polyphenols
TAA
80
60
60
Retention (%)
Retention (%)
80
TSS
Total carbohydrates
Total polyphenols
TAA
40
40
20
20
0
0
5
10
15
5
20
10
15
20
TMP (bar)
TMP (bar)
Retention of GK membrane towards analyzed compounds
Retention of GH membrane towards analyzed compounds
100
80
TSS
Total carbohydrates
Total polyphenols
TAA
60
40
20
0
5
10
15
20
TMP (bar)
Retention of GE membrane towards analyzed compounds
Diafiltration/concentration
20
3
Water
Jp
16
VRF
UF
permeate
WRF
Jp (Kg/m2h)
2
12
8
1
diafiltrate
Desal GE
Membrane
4
0
0
0
50
100
150
time (min)
Time evolution of permeate flux during
diafiltration/concentration processes
(Operating conditions: TMP, 10 bar; T, 27±1°C)
200
Diafiltration/concentration
Analyses of TSS, total polyphenols, total carboydrates and TAA in samples of gojy berries extracts coming
from the diafiltration/concentration processes
Diananofiltration*
Parameters
Initial
Total soluble solids (°Brix)
4.5±0.4
Final
Concentration in batch**
Final
0.4±0.02
0.8±1.2
Total polyphenols (mg/L gallic acid)
520.5±2.6
482.4±8.2
812.4±10.2
TAA (mM Trolox)
3.5±0.5
2.9±0.6
4.8±0.9
15.1±2.3
6.2±0.2
7.4±0.5
Total carboydrates (g/L glucose)
*After 4.5 volumes of diafitration
** Weight reduction factor (WRF) of 2
Goji leaves / Flow sheet of the investigated process
Goji leaves
Aqueous leaf extract
Washing
CLARIFICATION
(UF 100 kDa)
Retentate
Clarified extract
drying
FRACTIONATION
Extraction with water
Optimization
extraction parameters
(T, pH, solid/liquid ratio)
UP004
(Mycrodin Nadir)
flat-sheet
Polyethersulphone
MWCO = 4,000 Da
NP010
(Mycrodin Nadir)
flat-sheet
Polyethersulphone
MWCO = 1,000 Da
N30F
(Mycrodin Nadir)
flat-sheet
Polyethersulphone
MWCO = 0.300 Da
Optimization of extraction parameters
Effect of concentration (solid/liquid ratio) on the TSS, Total polyphenols and
TAA yields
12
2500
Total polyphenol retention (%)
10
6
4
2
1500
1000
500
0
0
50
100
150
200
250
0
300
0
50
Concentration (g/Kg)
100
150
200
Concentration (g/Kg)
16
14
12
TAA (mM Trolox)
TSS (°Brix)
8
2000
10
8
6
4
2
0
0
50
100
150
200
Concentration (g/Kg)
250
300
250
300
Optimization of extraction parameters
mAU
800
700
600
500
400
300
200
100
Ethanol/water (1:1) extraction: concentration 10 g/Kg
0 .
mAU 0
800
700
600
500
400
300
200
100
0
0
mAU
800
5
10
15
20
25
30
35
40
min
Aqueous extraction: concentration 10 g/Kg
5
10
15
20
25
30
35
40
min
700
600
500
400
Aqueous extraction: concentration 100 g/Kg
300
200
100
0
0
5
10
15
20
25
30
35
40
HPLC cromatograms of polyphenols in goji leaves extracts
min
Optimization of extraction parameters
Effect of temperature of the extraction on the TSS and Total polyphenols
yields
14
Total Polyphenols (mg/L gallic acid)
3000
TSS (°Brix)
12
10
8
6
2500
2000
1500
1000
500
4
30
40
50
60
70
Temperature (°C)
80
90
100
30
40
50
60
70
Temperature (°C)
80
90
100
Optimization of extraction parameters
Effect of the pH of the extraction solution on the TSS and Total polyphenols
yields
6
Total polyphenols (mg/L gallic acid)
1600
5
TSS (°Brix)
4
3
2
1
0
1400
1200
1000
800
600
400
0
2
4
6
8
pH
10
12
14
0
2
4
6
8
pH
10
12
14
Clarification of goji leaves extracts
20
DCQ II-006 C-PS100
Jp
VRF
Hollow fiber
15
4
100 kDa
VRF
2
Jp(Kg/m h)
Polysulphone
6
10
Ultrafiltration of goji leaves extracts.
Time course of permeate flux and VRF
(Operating conditions: TMP, 0.7 bar; Qf, 280 L/h;
T, 24±2°C)
2
5
0
0
0
50
100
150
200
time (min)
Parameters
Feed UF
Permeate UF
Retentate UF
pH
7.2±0.1
7.1±0.2
7.2±0.8
Total suspended solids
5.2±2.3
-
6.2±0.1
10.5 ±0.2
9.5±0.12
12±1.2
1870.7 ±4.5
1530.2 ±2.8
2570 ±2.6
13±1.2
11±2.6
14±1.6
6.33±1.6
5.43±0.12
9.65±2.8
Total soluble solids (°Brix)
Total polyphenols (mg/L gallic acid)
TAA (mM Trolox)
Total carbohydrates (g/L glucose)
Fractionation of clarified goji leaves extracts
Steady state permeate flux (kg/m2h)
100
UF 004
UP010
N30F
80
60
40
20
0
0
5
10
15
20
25
TMP (bar)
Treatment of clarified leaves aqueous extracts with UF and NF membranes .
Evolution of permeate flux as function of TMP
(Operating conditions: Qf, 300 L/h; T, 24±2°C)
Fractionation of clarified goji leaves extracts
100
100
Retention (%)
80
60
40
TSS
Total carbohydrates
Total polyphenols
TAA
60
40
20
20
0
5
0
5
10
15
10
20
20
Retention of NP010 membrane towards analyzed compounds
Retention of UP004 membrane towards analyzed compounds
100
15
TMP (bar)
TMP (bar)
TSS
Total carbohydrates
Total polyphenols
TAA
80
Retention (%)
Retention (%)
80
TSS
Total carbohydrates
Total polyphenols
TAA
60
40
20
0
5
10
15
20
TMP (bar)
Retention of N30F membrane towards analyzed compounds
Fractionation of clarified goji leaves extracts
Analyses of total polyphenols, TAA, total soluble solids and total carbohydrates in
goji leaves samples from NPO10 membrane at different WRF
Total
carbohydrates
(g/L glucose)
1520 ± 1.52
9.54 ± 1.47
10.2 ±0.2
5.61 ±0.01
Permeate WRF 1,5
525.59 ± 0.85
2.47 ± 0.23
5.6±0.05
4.84 ± 0.02
Permeate WRF 2
534.03 ± 0.20
3.02 ± 0.78
6.2±0.4
4.63 ± 0.05
Final permeate
528.60 ± 0.61
6.4±0.1
4.85 ± 0.4
Feed
Type of sample
2.80±0.1
100
80
Retention (%)
TAA
(mM Trolox)
Total
soluble solids
(%)
Total
polyphenols
(mg/L gallic acid)
60
40
20
le
ub
so
l
ta
l
To
TA
A
so
l
yd
bo
ca
r
ta
l
To
s
5.56 ± 0.2
ol
6.8±0.25
en
15.32 ± 0.99
ly
ph
2863.61 ± 0.42
po
Retentate WRF 2
ta
l
5.93 ± 0.8
s
6.5±0.3
To
10.49 ± 2.26
id
1961.73 ± 1.17
ra
Retentate WRF 1,5
te
0
Retention of NPO10 membrane towards analyzed compounds
Conclusions
A combination of aqueous extraction and membrane-based purification processes of
bioactive compounds from Goji berries and leaves has been studied and optimised.
The PVDF UF membrane allows to separate b-carotene in Goji berries aqueous
extracts with high efficiency in terms of retention (98%) and productivity (more than 40
kg/m2h).
Operating conditions (pH, temperature, concentration) for the aqueous extraction of
goji leaves have been optimized.
In the fractionation of both clarified extracts permeate fluxes increased linearly with
the operating pressure. At low pressure values a better separation of antioxidant
compounds (including polyphenols) from carbohydrates and TSS has been observed.
The sugar purification can be improved significantly if diafiltration is applied.
A strict correlation between the MWCO of selected membranes and the retention of
investigated compounds has been detected.
Tank you…for your kind attention!