evaluation of electrolyzed water for clean-in-place of ... water for nicma 1-17-2… · evaluation...
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EVALUATION OF ELECTROLYZED WATER FOR CLEAN-IN-PLACE OF DAIRY PROCESSING EQUIPMENT
Yun Yu and R. F. RobertsPresented at NICMA
1-17-2017
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Clean in place - CIP• The cleaning of items of equipment or pipeline
circuits without dismantling and with little orno manual involvement on the part of theoperator.
• CIP cleaning is accomplished by circulating solutions through the system.
(Tamime and Robinson, 1999)
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4-step CIP procedure• Pre-rinse
– to remove residual milk (soil)• Alkaline detergent circulation
– to remove bound protein and fat deposit• Intermediate rinse
– to remove all traces of detergent and entrained soil
– cool the system down for next step• Sanitizing
– to kill bacteria, and to retard bacterial growth
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Conventional CIP chemicals• Alkaline detergent
– NaOH• Saponification• Providing negative ions to disrupt soil
– Other components: sodium hypochlorite, surfactants
• Acidic Sanitizer– Chlorine
• Attacking bacterial cell wall and membrane– Other sanitizers: iodophors, peroxy acids
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Membrane
NaCl Solution Water
Acidic Alkaline
Na++ H- NaOHCl2+H2O HOCl+H++Cl-
Principle of electrolyzed water generation(Modified from www.kangenwater-asia.com) {Kim,2000} {Huang, 2008}
Electrolyzed water (EW)
Electrolyzed water generator
(www.ewatersystems.com)
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Conventional CIP Chemicals• Stored in
concentrated solutions
• Risk of burn on skin
• Cost
• Produced on-site
• Safe and easy to handle
• More cost effective– Water– Salt– Electricity
Electrolyzed Water
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Acidic EO water
• Electrolyzed oxidizing (EO) water pH: 2.3~2.7, ORP: >1100 mV, Free chlorine: 80-100 ppm
• Application (as disinfectant)– Medical, dental equipment– Food industry– Food service (restaurant, public eating
facilities)
Fabrizio & Cutter, 2003, 2004, & 2005; KIM, Huang, Brackett, & Frank,2001; Kim, Hung, & Brackett, 2000; Rahman et al., 2010; Venkitanarayananet al., 1999, Ozer and Demirci, 2006
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Alkaline ER water• Electrolyzed reducing (ER) water
– pH: 10.0~11.5– ORP: -800~ -900 mV
• Application in CIP (along with EOW)– for CIP of milking system on dairy farm
Walker, S.P., A. Demirci, R.E. Graves, S.B. Spencer, and R.F. Roberts. 2005. Response surface modelling for cleaning and disinfecting materials used in milking systems with electrolysed oxidizing water. Int. J. Dairy Technol. 58:65–73.
Walker, S., and A. Demirci. 2005. Cleaning milking systems using electrolyzed oxidizing water. Trans. ASAE. 48:1827–1834.
Dev, S.R.S., A. Demirci, and R. Graves. 2011. Mathematical modeling of CIP of milking systems using electrolyzed oxidizing water. Northeast Agric. Biol. Eng. Conf.
Wang, X., S.R.S. Dev, A. Demirci, R.E. Graves, and V.M. Puri. 2012. Electrolyzed oxidizing water for cleaning-in-place of milking systems on dairy farms – Performance evaluation and assessment. NABEC-CSBE/SCGAB 2012 Jt. Meet. Tech. Conf. Northeast Agric. Biol. Eng. Conf. Can.
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Hypothesis:Electrolyzed water can be used as aneffective cleaning and sanitizing agent formilk processing equipment.
Research question:Can electrolyzed water serve as analternative to conventional CIP chemicalsfor CIP of milk processing equipment?
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Objectives1. Construct a pilot-sale test system, and validate its performance and cleanability by CIP.
2. Develop CIP procedure using EW for cleaningof refrigerated milk storage tanks.
3. Optimize CIP procedure using EW for a tank used to heat milk.
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CONSTRUCTION, CHARACTERIZATION AND VALIDATION OF TEST SYSTEM
Objective 1
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Overview of pilot scale test system
Construction of test system
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Test Vessel
TE
Jacket media
Jacket media
Agitator
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Test Vessel
TE
Jacket media
Jacket media
Water
Watertank
Drain
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CIP tank
Coil heat exchanger
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Test Vessel
TE
Jacket media
Jacket media
Water
Watertank
CIP tank
Coil heat exchanger
Schematic of pilot CIP process test systemDrain
Construction of test system
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Characterization of test system• Determine flow rate of each pump
setting
• Check coverage of 360° static spray ball using riboflavin test
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Volumetric flow rate measurements
1.83.9
6.08.3
10.011.8
13.515.3
17.3
0.003.006.009.00
12.0015.0018.00
10 20 30 40 50 60 70 80 90
Mea
n flow
rat
e (L
/min)
VFD pump setting
Mean volumetric flow rates (L/min) at different VFD pump settings. The regression equation is: Flow Rate (L/min) = 0.18 + 0.19 (Pump Setting).
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196.0 L/min 8.3 L/min
3.9 L/min
1.8 L/min 10 L/minRiboflavin solution (0.2 gram/L)
Experiment: Riboflavin coverage test
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Before rinse 1.8 L/min for 1 min
3.9 L/min for 7 min 6.0 L/min for 17 min 8.3 L/min for 1 min
Residual riboflavin pattern at different flow rates
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To validate CIP performance of the test system, efficacy of conventional CIP was evaluated after using the tank to heat milk
Validation
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Pre-clean manually
Alkaline wash SanitizingPost-rinse
Soiling
Pre-rinse
Experiment: Validation of test system
Soiling:heat whole milk to 74°C (165°F) under agitation then hold for 15 min
Pre-rinse Post-rinse
Soiling
Pre-clean manually
Pre-clean manually:rinse -> wash -> rinse -> sanitize(water) (HC-10) (water) (XY-12)
Pre-rinse & Post-rinsewith water at room temperature for 3 min
Alkaline wash Sanitizing
Alkaline washwith Principal (3200 ppm)at 63°C for 15 min
Sanitizing:with XY-12 (100 ppm)at room temperaturefor 3 min
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Cleanliness assessments
( ) Cleanliness assessments– ATP bioluminescence assay– Residual protein detection
Pre-clean manually
Alkaline wash SanitizingPost-rinse
Soiling
Pre-rinse
1
432
0
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ATP Bioluminescence Method
– Quantitative result : relative light units (RLU)
www.charm.comPocket swab NovaLum ATP analyzer
50 cm2
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Protein Detection Method
– semi-quantitative result : 1~4 score
www.hygiena.net
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Result: Conventional CIP returned the test system to a cleaning condition
SamplingCommercial treatment
ATP Protein (μg)
After soiling Log10 RLU = 5.38 ± 0.27 Fail >100 Fail
After pre-rinse Log10 RLU = 5.60 ± 0.35 Fail >100 Fail
After post-rinse RLU = 0 ± 0 Pass 0~20 Pass
After sanitizing RLU = 0 ± 0 Pass 0~20 Pass
Cleanliness assessments of ATP and protein measurements after different steps of CIP using commercial chemicals following conventional procedure
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For swab sampling…• Each 50 cm2 inner surface could be
swabbed only once• Does sampling location effect the
variation of cleanliness measurements?
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Acid sanitizing with EOW
Test for swab sampling variability
Schematic of sampling locations
Soiling (heated)
Rinse
( ) ATP bioluminescence assayTo evaluate:– location effect,
swab three areas for each assessment
– repeatability between trials,repeated experiment in three times
– quench effect of EOW on ATPassay,
rinse with EOW before assessment
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log10 RLU comparison
Average log10 RLU comparison between trials and CIP steps
AB AB ABA A ABAB B AB
0.000
1.000
2.000
3.000
4.000
5.000
6.000
7.000
Trial #1 Trial #2 Trial #3
Log1
0 RL
U
After soilingAfter pre-rinseAfter sanitizing
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Summary of objective-1• A test system was constructed and characterized
to evaluate CIP of milk processing equipment.• Flow rate of 8.3 L/min was determined to provide
adequate coverage.• Sampling location was not a factor in RLU value
variations.• The repeatability of the test system was
acceptable.• The test system was able to be cleaned using a
conventional CIP procedure after soiling with milk under heating.
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CIP CLEANING USING ELECTROLYZED WATER FOR REFRIGERATED MILK STORAGE TANK
Objective 2
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To mimic raw milk silo…• Inoculated bacterial cocktail:
– 10 ml Escherichia coli (9.1 log10 CFU/ml)– 10 ml Enterococcus faecalis (9.4 log10 CFU/ml)– 10 ml Pseudomonas fluorescens (9.1 log10 CFU/ml)
into 11.4 L (3 gal.) of pasteurized whole milk
• Bacterial level in inoculated milk, 6.7 log10 CFU/ml
Milk
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Preparation of EW• pH and ORP • Chlorine
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EO Water ER Water
pHORP (mV)
Total [Cl-]
(ppm)
Free [Cl-]
(ppm) pHORP (mV)
[OH-](ppm)
Minimum 2.19 1157 64.1 60.5 11.58 -897 64.7
Maximum 2.51 1182 126.7 122.0 11.78 -844 102.5
Average 2.38 1169 90.5 85.8 11.66 -872 77.7
St.Dev. 0.06 6.08 12.76 12.27 0.06 13.21 10.21
Chemical properties of EW
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Alkaline wash SanitizingPost-rinsePre-rinse
Soiling
Experimental design
Soiling:Inoculated whole milk and hold at 2-4°C for 18 hours
Pre-rinse Post-rinse
Soiling
Pre-clean manually
Pre-clean manually:rinse -> wash -> rinse -> sanitize(water) (HC-10) (water) (XY-12) `0
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Washing temperature and time of four treatments
TreatmentsWash
Chemical Temp. (°C) Time (min)
Pos Ctrl Principal 63 15
EW Long ER water 40 15
EW Short ER water 40 5
Neg Ctrl water 40 15
Alkaline wash SanitizingPost-rinsePre-rinse
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Alkaline wash SanitizingPost-rinsePre-rinse
Sanitizing temperature and time of four treatments
TreatmentsSanitizing
Chemical Temp. (°C) Time (min)
Pos Ctrl XY 12 25 1
EW Long EO water 25 1
EW Short EO water 25 1
Neg Ctrl water 25 1
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Alkaline wash SanitizingPost-rinsePre-rinse
Soiling
Assess the cleanliness
Soiling:Inoculated whole milk to 2-4°C for 18 hours
( ) Cleanliness assessmentsATP bioluminescence assayResidual protein detectionMicrobial enrichment analyses
1
432
0
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Means of RLU values. Tukey’s comparison were conducted between 16 means of log10(RLU) measurements. Means that do not share a letter indicates significant difference (P < 0.05)
RLU resultsa
b
d d
a
b
d d
a
b
c
d
a
bb
b
1
10
100
1000
10000
100000
1000000
After Soiling After Pre-rinse After Post-rinse Sanitizing
RLU
Pos ctrlEW longEW shortNeg ctrl
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Protein (ug) After Soiling
After Pre-rinse
After Post-rinse
After Sanitizing
Pos ctrl 60 ~ 100 20 ~ 40 0 ~ 20 0 ~ 20EW long 60 ~ 100 20 ~ 40 0 ~ 20 0 ~ 20
EW short 60 ~ 100 20 ~ 40 0 ~ 20 0 ~ 20Neg ctrl 60 ~ 100 20 ~ 40 20 ~ 40 0 ~ 20
Protein results
Means of protein residue levels after different CIP steps
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Microbial enrichment analyses
Micro APC (log10cfu/ml)
Soiling Pre-rinse Post-rinse Sanitizing
Pos ctrl 6.4±0.01 + + - -EW long 6.5±0.08 + + - -EW short 6.6±0.11 + + - -Neg ctrl 6.6±0.09 + + - -
Microbiological enrichment results after different CIP steps
Incubate at 30°C for 48 h
Swab 50 cm2
using sterile alginate swab
+ -
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Summary of objective-2• A CIP procedure was developed for cleaning
of refrigerated milk storage tank:Pre-rinse Wash Post-rinse Sanitizing
Agent Water ERW Water EOWTemperature RT 40°C RT 25°CTime 1 min 15 min 3 min 1 min
• So far…– The use of test system for evaluation CIP
procedure was validated.– A refrigerated milk tank was able to be
cleaned using EW.
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CIP USING ELECTROLYZED WATER FOR HEATED MILK PROCESSING TANK
Objective 3
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Alkaline wash SanitizingPost-rinsePre-rinse
Soiling
Experimental design
Soiling:heat whole milk to 74°C (165°F) under agitation then hold for 15 min
Pre-rinse Post-rinse
Soiling
Pre-clean manually
Pre-clean manually:rinse -> wash -> rinse -> sanitize(water) (HC-10) (water) (XY-12) `
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Alkaline wash SanitizingPost-rinsePre-rinse
Soiling
Experimental designSoiling:
heat whole milk to 165 °F (74 °C) under agitation then hold for 15 min
Soiling
Alkaline wash Sanitizing
Code -1 0 1WashTemp (°C) 40 55 70WashTime (min) 5 15 25SaniTemp (°C) 25 35 45SaniTime (min) 1 5.5 10
4 factors, 3 levels
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Response surface model• Box-Behnken design matrix:
27 trials including 3 center points
Trials Run order WTemp WTime STemp STime
1 2 40°C 5 min 35°C 5.5 min2 10 70°C 5 min 35°C 5.5 min3 26 40°C 25 min 35°C 5.5 min4 6 70°C 25 min 35°C 5.5 min5 18 55°C 15 min 25°C 1 min6 13 55°C 25 min 45°C 5.5 min// // // // // //
25 27 55°C 15 min 35°C 5.5 min26 8 55°C 15 min 35°C 5.5 min27 23 55°C 15 min 35°C 5.5 min
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Alkaline wash SanitizingPost-rinsePre-rinse
Soiling
Cleanliness assessmentsSoiling:
heat whole milk to 165 °F (74 °C) under agitation then hold for 15 min
( ) Cleanliness assessmentsATP bioluminescence assay (RLU 1~4)Residual protein detection (Protein 1~4)
1 432
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RLU3 data analysis and modeling• Factors:
• WashTemp• WashTime
• Percentage RLU reduction (PRR): ((RLU2−RLU3)RLU2
)100%
• Transformed Response:(Natural logarithms transformation of PRR)
𝑙𝑙𝑙𝑙((RLU2−RLU3)RLU2
) (range: ≤0 )
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Regression model
ln(RLU2 − RLU3
RLU2)
= −0.6038 + 0.014045 × WashTemp+ 0.0205 × WashTime − 0.0001 × WashTemp 2
− 0.0002 × WashTime 2 − [0.0002 × WashTemp× WashTime]
Factor P-valueWashTemp 0.068
WashTime 0.011
WashTemp2 0.195
WashTime2 0.128
WashTemp × WashTime 0.068
(R2 = 48%)
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WashTemp
Was
hTim
e
656055504540
25
20
15
10
5
> – – – – – – – – – – <
-0.039 -0.027-0.027 -0.015
-0.015
-0.135-0.135 -0.123-0.123 -0.111-0.111 -0.099-0.099 -0.087-0.087 -0.075-0.075 -0.063-0.063 -0.051-0.051 -0.039
ln((RLU2-RLU3)/RLU2)
Contour Plot of ln((RLU2-RLU3)/RLU2) vs WashTime, WashTemp
After alkaline wash and post-rinse
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CurHighLow0.98160
DNew
d = 0.98160
Targ: 0.0ln((RLU2
y = -0.0092
0.98160DesirabilityComposite
5.025.0
40.069.40
WashTimeWashTemp
[54.6497] [20.5043]
𝑙𝑙𝑙𝑙(𝑅𝑅𝑅𝑅𝑅𝑅𝑅 − 𝑅𝑅𝑅𝑅𝑅𝑅𝑅
𝑅𝑅𝑅𝑅𝑅𝑅𝑅)
Optimization of ER water wash stepOptimum wash temperature = 54.6°C
Optimum wash time = 20.5 min
Predicted RLU reduction = 99.08%
Conventional ctrl:100% RLU reduction
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Conclusions• The test system developed was suitable for
evaluating CIP cleaning of milk tanks.
• It was possible to CIP clean a refrigerated milk storage tank using EW.
• It was possible to CIP clean a tank used to heat milk using the optimal condition, but the system was not as effective as when cleaned with conventional chemicals.
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Future research• Consider methods to improve the cleaning
efficacy of alkaline ER water – add surfactants– Increase alkalinity– Add other detergent builder
• To improve acidic EO water– Use neutral electrolyzed water with less corrosive
characteristic– Add corrosion inhibitors, such as silicates to acid
EO water to prevent corrosion