July 2006
NSF05/12b/EPADWCTR
EPA/600/R-06/131
Environmental Technology
Verification Report
Physical Removal of Microbial
Contaminants in Drinking Water
Watts Premier, Inc.
WP-4V Point-of-Use Drinking Water
Treatment System
Prepared by
NSF International
Under a Cooperative Agreement with
U.S. Environmental Protection Agency
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THE ENVIRONMENTAL TECHNOLOGY VERIFICATION
PROGRAM
&EPA
ET
V^lVl
V
U.S. Environmental Protection Agency
NSF International
ETV Joint Verification Statement
TECHNOLOGY TYPE: POINT-OF-USE DRINKING WATER TREATMENT
SYSTEM
APPLICATION: REMOVAL OF MICROBIAL CONTAMINANTS IN
DRINKING WATER
PRODUCT NAME: WATTS PREMIER WP-4V
VENDOR: WATTS PREMIER
ADDRESS: 1725 WEST WILLIAMS DR.
SUITE C-20
PHOENIX, AZ 85027
PHONE: 800-752-5582
INTERNET http://www.wattspremier.com
NSF International (NSF) manages the Drinking Water Systems (DWS) Center under the U.S.
Environmental Protection Agency's (EPA) Environmental Technology Verification (ETV) Program. The
DWS Center recently evaluated the performance of the Watts Premier WP-4V point-of-use (POU) reverse
osmosis (RO) drinking water treatment system. NSF performed all of the testing activities and also
authored the verification report and this verification statement. The verification report contains a
comprehensive description of the test.
EPA created the ETV Program to facilitate the deployment of innovative or improved environmental
technologies through performance verification and dissemination of information. The goal of the ETV
Program is to further environmental protection by accelerating the acceptance and use of improved and
more cost-effective technologies. ETV seeks to achieve this goal by providing high-quality, peer-
reviewed data on technology performance to those involved in the design, distribution, permitting,
purchase, and use of environmental technologies.
ETV works in partnership with recognized standards and testing organizations, stakeholder groups
(consisting of buyers, vendor organizations, and permitters), and with the full participation of individual
technology developers. The program evaluates the performance of innovative technologies by developing
test plans that are responsive to the needs of stakeholders, conducting field or laboratory tests (as
appropriate), collecting and analyzing data, and preparing peer-reviewed reports. All evaluations are
conducted in accordance with rigorous quality assurance protocols to ensure that data of known and
adequate quality are generated and that the results are defensible.
NSF 06/12b/EPADWCTR The accompanying notice is an integral part of this verification statement. July 2006
VS-i
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ABSTRACT
The Watts Premier WP-4V four-stage POU RO system was tested for removal of bacteria and viruses at
NSF's Drinking Water Treatment Systems Laboratory. Five systems were challenged with the
bacteriophage viruses fir and MS2, and the bacteria Brevundimonas diminuta. The virus challenges were
conducted at three different pH settings (6, 7.5, and 9) to assess whether pH influences the performance of
the RO membrane. The bacteria challenges were conducted only at pH 7.5.
The challenge concentrations ranged from 3.8 to 5.0 logs for the viruses, and 6.4 to 7.2 logs for the
bacteria. The log reductions ranged from 1.3 to 6.4 logio for B. diminuta, with an average of 2.1 Iogi0.
The virus log reductions ranged from 1.4 to 3.6 logic for fir, and 1.2 to 3.7 logic for MS2. The average
virus logic reductions were 2.5 and 2.7, respectively. The virus challenge data does not indicate that the
pH of the challenge water influenced removal by the RO membrane. See Table VS-2 below for the
complete log reduction data.
TECHNOLOGY DESCRIPTION
The following technology description was provided by the manufacturer and has not been verified.
The WP-4V is a four-stage POU drinking water treatment system using sediment filtration, activated
carbon filtration, and reverse osmosis. Treated water is stored in a three-gallon storage tank. The WP-4V
is certified by NSF to NSF/ANSI Standard 58 - Reverse Osmosis Drinking Water Treatment Systems. It
has a certified production rate of 9.06 gallons per day.
Incoming water first passes through a sediment filter to remove particulate matter, such as rust and silt,
and then through a carbon filter to remove chlorine or other contaminants. The third stage of treatment is
the reverse osmosis membrane, which removes a wide variety of inorganic and larger molecular weight
organic contaminants, and also protozoan cysts such as Cryptosporidium and Giardia. The permeate
water is sent to a 3-gallon maximum capacity storage tank. Upon leaving the storage tank, the water
passes through a second carbon filter to remove organic chemicals and other taste and odor causing
substances before dispensing through the faucet. The pre-membrane carbon and sediment filters were not
tested, because they are only designed to remove chlorine and particulate matter to protect the RO
membrane.
VERIFICATION TESTING DESCRIPTION
Test Site
The testing site was the Drinking Water Treatment Systems Laboratory at NSF in Ann Arbor, Michigan.
A description of the test apparatus can be found in the test/QA plan and verification report. The testing
was conducted in June and July of 2005.
Methods and Procedures
The testing methods and procedures are detailed in the Test/QA Plan for Verification Testing of the Watts
Premier WP-4V Point-of-Use Drinking Water Treatment System for Removal of Microbial
Contamination Agents. Five WP-4V systems were tested for bacteria and virus removal performance
using the bacteriophage viruses fir and MS2, and the bacteria Brevundimonas diminuta. The challenge
organisms were chosen because they are smaller than most other viruses and bacteria, and so provide a
conservative estimate of performance. NSF also used a genetically engineered strain of B. diminuta. The
NSF Microbiology Laboratory inserted into a culture of B. diminuta strain 19146 a gene conferring
resistance to the antibiotic kanamycin. This allowed the Microbiology Laboratory to use a growth media
NSF 06/12b/EPADWCTR The accompanying notice is an integral part of this verification statement. July 2006
VS-ii
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amended with 50 jog/mL of kanamycin to prohibit heterotrophic plate count (HPC) bacteria in the treated
water samples from growing along with the kanamycin resistant B. diminuta.
Five systems were evaluated. The systems were installed on a test rig and conditioned according to the
vendor's instructions (fill the storage tanks and dispensing the contents to a drain three times), and then
were conditioned for another five days. Prior to testing, the systems were evaluated for reduction of total
dissolved solids (TDS) to ensure that the systems undergoing testing were representative of the expected
performance of the system.
The test water for the bacteria challenges was set to pH 7.5 ± 0.5, while the virus challenges were
conducted at pH 6.0 ± 0.5, 7.5 ± 0.5, and 9.0 ± 0.5. The challenge schedule is shown in Table VS-1. The
virus challenges were conducted at different pH settings to evaluate whether the surface charges of the
viruses influenced their removal through electrostatic forces versus mechanical filtration. Viruses have
different surface charges, or different strengths of negative or positive charge, depending on their
isoelectric point and the pH of the water. The isoelectric point is the pH at which the virus surface is
neutrally charged. MS2's isoelectric point is pH 3.9, and fr's is pH 8.9. In solutions above the isoelectric
point, the virus is negatively charged. Below the isoelectric point, the virus is positively charged.
Table VS-1. Challenge Schedule
Day Surrogate Challenge pH
1 B. diminuta 7.5 ± 0.5
2 fr and MS2 6.0 ± 0.5
3 fr and MS2 7.5 ± 0.5
4 Kanamycin Resistant B. diminuta 7.5 ± 0.5
5 fr and MS2 9.0 ± 0.5
For each challenge, the systems were operated for one tank-fill period (approximately four to five hours).
The end of this period was evident through engagement of each system's automatic shutoff mechanism,
which causes the flow of reject water to cease. Influent water samples were collected at the beginning
and end of each challenge period. After each system ceased operation, the contents of the product water
storage tanks were emptied into sterile containers, and samples were collected for microbiological
analysis. All samples were enumerated in triplicate. Following each challenge period, the systems were
flushed by operating them for one tank-fill period using water without challenge organisms.
VERIFICATION OF PERFORMANCE
As discussed above, the systems were first subjected to a TDS reduction test to verify that the RO
membranes would perform as expected. The observed TDS reduction ranged from 89% to 96%. The
certified TDS reduction for the WP-4V is 97%.
The bacteria and virus logio reduction data is presented in Table VS-2. The logio reduction of B. diminuta
("normal" and kanamycin resistant B. diminuta combined) ranged from 1.3 to 6.4, with an average logio
reduction of 1.9. The challenge organisms were detected in the effluent samples for all test units but Unit
2 for the "normal" B. diminuta challenge. Since the Unit 2 effluent count for kanamycin resistant B.
diminuta was 4.3 logio, and all other effluent samples had bacteria counts greater than 4 logio (data not
shown), it is possible that there was a sampling or analytical error associated with the Unit 2 "normal" B.
diminuta sample. Therefore, that sample was not included in the mean logio reduction calculation for the
bacteria.
NSF 06/12b/EPADWCTR The accompanying notice is an integral part of this verification statement. July 2006
VS-iii
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The virus challenge data showed similar performance. The logic reduction of the fir virus ranged from 1.4
to 3.6, with an overall mean of 2.5. The logic reduction of MS2 ranged from 1.2 to 3.7, with an overall
mean of 2.6. A visual comparison of the logic reductions versus the challenge water pH shows the mean
logic reductions decreasing with increasing pH. However, an examination of the 95% confidence
intervals around the means (see verification report for data) shows that the decreases are not statistically
significant.
The minimum observed log reductions equal removal of 95% of B. diminuta, and 94% of the viruses.
Table VS-2. Bacteria and Virus Log Reduction Data
Initial Final Logio „ . ,f T _ ,
Measured Measured Challenge Influent Geometric Mean Loglo Reduction
Target pH pH pH Organisms Challenge Unit 1 Unit 2 Unit 3 Unit 4 Unit 5
7.5+0.5 7.6 7.8 B. diminuta 6.4 1.8
Kanamycin
7.5 + 0.5 7.5 7.8 Resistant 7.2 1.4
B. diminuta
6.0 ±0.5 6.1 6.5 fr 3.9 1.8
MS2 3.8 2.3
7.5 ±0.5 7.5 7.7 fr 4.5 1.9
MS2 4.2 1.7
9.0 ±0.5 8.9 9.0 fr 5.0 1.4
MS2 4.6 1.2
*Number not included in mean log reduction calculation.
6.4* 1.3 1.5 1.6
2.9 2.6 2.6 3.1
3.1 3.6 3.4 3.0
3.4 3.7 3.6 2.9
2.4 2.3 3.1 2.8
2.4 2.4 3.4 3.2
2.3 2.1 2.3 2.6
2.4 2.0 2.3 3.0
Overall Means: B. diminuta
fr
MS2
QUALITY ASSURANCE/QUALITY CONTROL (QA/QC)
NSF provided technical and quality assurance oversight of the verification testing as described in
verification report, including a review of nearly 100% of the data. NSF personnel also conducted
technical systems audit during testing to ensure the testing was in compliance with the test plan.
complete description of the QA/QC procedures is provided in the verification report.
Mean
1.5
2.4
2.9
3.1
2.5
2.5
2.1
2.1
1.9
2.5
2.6
the
a
A
NSF 06/12b/EPADWCTR
The accompanying notice is an integral part of this verification statement.
VS-iv
July 2006
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Original signed by Sally Gutierrez 08/11/06 Original signed by Robert Ferguson 08/23/06
Sally Gutierrez Date Robert Ferguson Date
Director Vice President
National Risk Management Research Laboratory Water Systems
Office of Research and Development NSF International
United States Environmental Protection Agency
NOTICE: Verifications are based on an evaluation of technology performance under specific,
predetermined criteria and the appropriate quality assurance procedures. EPA and NSF make no
expressed or implied warranties as to the performance of the technology and do not certify that a
technology will always operate as verified. The end-user is solely responsible for complying with
any and all applicable federal, state, and local requirements. Mention of corporate names, trade
names, or commercial products does not constitute endorsement or recommendation for use of
specific products. This report is not an NSF Certification of the specific product mentioned
herein.
Availability of Supporting Documents
Copies of the test protocol, the verification statement, and the verification report (NSF
report # NSF 06/12b/EPADWCTR) are available from the following sources:
(NOTE: Appendices are not included in the verification report. Appendices are available
from NSF upon request.)
1. ETV Drinking Water Systems Center Manager (order hard copy)
NSF International
P.O. Box 130140
Ann Arbor, Michigan 48113-0140
2. Electronic PDF copy
NSF web site: http://www.nsf.org/etv
EPA web site: http://www.epa.gov/etv
NSF 06/12b/EPADWCTR The accompanying notice is an integral part of this verification statement. July 2006
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July 2006
Environmental Technology Verification Report
Physical Removal of Microbial Contaminants in Drinking Water
Watts Premier Incorporated
WP-4V Point-of-Use Drinking Water Treatment System
Prepared by:
NSF International
Ann Arbor, Michigan 48105
Under a cooperative agreement with the U.S. Environmental Protection Agency
Jeffrey Q. Adams, Project Officer
National JAisk Management Research Laboratory
U.S. Environmental Protection Agency
Cincinnati, Ohio 45268
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Notice
The U.S. Environmental Protection Agency (USEPA), through its Office of Research and
Development, has financially supported and collaborated with NSF International (NSF) under
Cooperative Agreement No. R-82833301. This verification effort was supported by the Drinking
Water Systems (DWS) Center, operating under the Environmental Technology Verification
(ETV) Program. This document has been peer-reviewed, reviewed by NSF and USEPA, and
recommended for public release.
11
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Foreword
The U.S. Environmental Protection Agency (USEPA) is charged by Congress with protecting the
Nation's land, air, and water resources. Under a mandate of national environmental laws, the
Agency strives to formulate and implement actions leading to a compatible balance between
human activities and the ability of natural systems to support and nurture life. To meet this
mandate, USEPA's research program is providing data and technical support for solving
environmental problems today and building a science knowledge base necessary to manage our
ecological resources wisely, understand how pollutants affect our health, and prevent or reduce
environmental risks in the future.
The National Risk Management Research Laboratory (NRMRL) is the Agency's center for
investigation of technological and management approaches for preventing and reducing risks
from pollution that threaten human health and the environment. The focus of the Laboratory's
research program is on methods and their cost-effectiveness for prevention and control of
pollution to air, land, water, and subsurface resources; protection of water quality in public water
systems; remediation of contaminated sites, sediments and ground water; prevention and control
of indoor air pollution; and restoration of ecosystems. NRMRL collaborates with both public
and private sector partners to foster technologies that reduce the cost of compliance and to
anticipate emerging problems. NRMRL's research provides solutions to environmental problems
by: developing and promoting technologies that protect and improve the environment; advancing
scientific and engineering information to support regulatory and policy decisions; and providing
the technical support and information transfer to ensure implementation of environmental
regulations and strategies at the national, state, and community levels.
This publication has been produced as part of the Laboratory's strategic long-term research plan.
It is published and made available by USEPA's Office of Research and Development to assist
the user community and to link researchers with their clients.
Sally Gutierrez, Director
National Risk Management Research Laboratory
in
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Table of Contents
Verification Statement VS-i
Title Page i
Notice ii
Foreword iii
Table of Contents iv
List of Tables vi
List of Figures vi
Abbreviations and Acronyms vii
Acknowlegements viii
Chapter 1 Introduction 1
1.1 Environmental Technology Verification (ETV) Program Purpose and Operation 1
1.2 Purpose of Verification 1
1.3 Development of Test/Quality Assurance (QA)Plan 1
1.3.1 Bacteria and Virus Surrogates 2
1.4 Testing Participants and Responsibilities 3
1.4.1 NSF International 3
1.4.2 Watts Premier 4
1.4.3 U.S. Environmental Protection Agency 4
Chapter 2 Equipment Description 5
2.1 RO Membrane Operation 5
2.2 Equipment Capabilities 5
2.3 System Components 5
2.4 Same or Similar Models 5
2.5 System Operation 6
2.6 Rate of Waste Product!on 8
2.7 Equipment Operation Limitations 8
2.8 Operation and Maintenance Requirements 8
Chapters Methods and Procedures 9
3.1 Introduction 9
3.2 Verification Test Procedure 9
3.2.1 Test Rig 9
3.2.2 Test Rig Sanitization 9
3.2.3 Test Water 9
3.2.3.1 Base Water 9
3.2.3.2 Bacteria and Virus Challenges 11
3.2.4 System Operation 12
3.2.4.1 System Installation 12
3.2.4.2 TDS Reduction System Check 12
3.2.4.3 Additional Conditioning 12
3.2.4.4 Challenge Testing 12
iv
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3.3 Analytical Methods 13
3.3.1 Water Quality Analytical Methods 13
3.3.2 Microbiology Analytical Methods 14
3.3.2.1 Sample Processing, and Enumeration of Viruses 14
3.3.2.2 Bacteria Cultivation 14
3.3.2.3 Preparation of B. diminuta Challenge Suspensions 14
3.3.2.4 Sample Processing and Enumeration of B. diminuta 15
Chapter 4 Results and Discussion 16
4.1 IDS Reduction 16
4.2 Bacteria Reduction 16
4.3 Virus Reduction 17
Chapters QA/QC 18
5.1 Introduction 18
5.2 Test Procedure QA/QC 18
5.3 Sample Handling 18
5.4 Chemistry Analytical Methods QA/QC 18
5.5 Microbiology Laboratory QA/QC 18
5.5.1 Growth Media Positive Controls 18
5.5.2 Negative Controls 19
5.5.3 Bacteria Cell Size 19
5.6 Documentation 19
5.7 Data Review 19
5.8 Data Quality Indicators 20
5.8.1 Representativeness 20
5.8.2 Accuracy 20
5.8.3 Precision 20
5.8.4 Completeness 21
5.8.4.1 Completeness Measurements 21
5.9 Measurements Outside of the Test/QA Plan Specifications 22
Chapter 6 References 23
Appendices
Appendix A Bacteria and Virus Counts, and Water Chemistry Data
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List of Tables
Table 1-1. Virus and Host ATCC Designations .
Table 3-1. Challenge Schedule
Table 4-1. TDS Reduction Test Results
Table 4-2. Bacteria Log Reduction Data
Table 4-3. Virus Log Reduction Data
Table 5-1. Bacteria Cell Size Measurements
Table 5-2. Completeness Requirements
...3
.13
.16
.17
.17
.19
.21
List of Figures
Figure 2-1. Photograph of the WP-4V System...
Figure 2-2. Schematic Diagram of the WP-4V...
Figure 3-1. Schematic Diagram of Test Rig
Figure 3-2. Systems Installed on Test Rig
...7
.10
.11
VI
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Abbreviations and Acronyms
ANSI
ASTM
ATCC
°C
CPU
cm
DWS
ETV
°F
HPC
L
mg
mL
nm
NRMRL
NSF
PBDW
PFU
POE
POU
psig
QA
QC
QA/QC
RO
RPD
SLB
SOP
IDS
ISA
TSB
Hg
jiL
jam
|o,mho
liS
USEPA
American National Standards Institute
American Society of Testing Materials
American Type Culture Collection
Degrees Celsius
Colony Forming Unit
Centimeter
Drinking Water Systems
Environmental Technology Verification
Degrees Fahrenheit
Heterotrophic Plate Count
Liter
Milligram
Milliliter
Nanometer
National Risk Management Research Laboratory
NSF International (formerly known as National Sanitation Foundation)
Phosphate-Buffered Dilution Water
Plaque Forming Unit
Point-of-Entry
Point-of-Use
Pounds per Square Inch Gauge
Quality Assurance
Quality Control
Quality Assurance/Quality Control
Reverse Osmosis
Relative Percent Difference
Saline Lactose Broth
Standard Operating Procedure
Total Dissolved Solids
Tryptic Soy Agar
Tryptic Soy Broth
Microgram
Microliter
Micrometer
Micromho
MicroSieman
U. S. Environmental Protection Agency
vn
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Acknowledgments
NSF International was responsible for all elements in the testing sequence, including collection
of samples, calibration and verification of instruments, data collection and analysis, data
management, data interpretation and the preparation of this report.
The Manufacturer of the Equipment was:
Watts Premier Incorporated
1725 West Williams Drive
Suite C-20
Phoenix, AZ 85027
NSF wishes to thank the members of the expert technical panel for their assistance with
development of the test plan.
Vlll
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Chapter 1
Introduction
1.1 Environmental Technology Verification (ETV) Program Purpose and Operation
The U.S. Environmental Protection Agency (USEPA) has created the ETV Program to facilitate
the deployment of innovative or improved environmental technologies through performance
verification and dissemination of information. The goal of the ETV Program is to further
environmental protection by accelerating the acceptance and use of improved and more cost-
effective technologies. ETV seeks to achieve this goal by providing high-quality, peer-reviewed
data on technology performance to those involved in the design, distribution, permitting,
purchase, and use of environmental technologies.
ETV works in partnership with recognized standards and testing organizations; with stakeholder
groups consisting of buyers, vendor organizations, and permitters; and with the full participation
of individual technology developers. The program evaluates the performance of innovative
technologies by developing test plans that are responsive to the needs of stakeholders; by
conducting field or laboratory testing, collecting and analyzing data; and by preparing peer-
reviewed reports. All evaluations are conducted in accordance with rigorous quality assurance
protocols to ensure that data of known and adequate quality are generated and that the results are
defensible.
The USEPA has partnered with NSF International (NSF) under the ETV Drinking Water
Systems (DWS) Center to verify performance of drinking water treatment systems that benefit
the public and small communities. It is important to note that verification of the equipment does
not mean the equipment is "certified" by NSF or "accepted" by USEPA. Rather, it recognizes
that the performance of the equipment has been determined and verified by these organizations
under conditions specified in ETV protocols and test plans.
1.2 Purpose of Verification
The purpose of this verification was to evaluate treatment system performance under a simulated
intentional or non-intentional microbiological contamination event. Because any contamination
event would likely be short-lived, the challenge period for each chemical lasted only a few hours.
Long-term performance over the life of the membrane was not investigated.
1.3 Development of Test/Quality Assurance (QA) Plan
USEPA's "Water Security Research and Technical Support Action Plan" (USEPA, 2004)
identifies the need to evaluate point-of-use (POU) and point-of-entry (POE) treatment system
capabilities for removing likely contaminants from drinking water. As part of the ETV program
NSF developed a test/QA plan for evaluating POU reverse osmosis (RO) drinking water
treatment systems for removal of microbial contaminants. To assist in this endeavor, NSF
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assembled an expert technical panel, which gave suggestions on a protocol design prior to
development of the test/QA plan. Panel members included experts from USEPA, United States
Army, and United States Centers for Disease Control and Prevention, Division of Parasitic
Diseases, as well as a water utility microbiologist, a university professor, and an independent
consultant in the POU drinking water treatment systems industry.
The product-specific test/QA plan for evaluating the WP-4V was entitled Test/QA Plan for
Verification Testing of the Watts Premier WP-4V Point-of-Use Drinking Water Treatment
System for Removal ofMicrobial Contamination Agents. This test/QA plan uses surrogate
bacteria and viruses in place of testing with the actual agents of concern. The test organisms
serve as surrogates not only for bacteria and viruses, but also protozoa, such as Cryptosporidium
oocysts. Please note that this test plan does not cover chemical agents derived from
microorganisms, such as ricin or botulism toxin.
By participating in this ETV, Watts Premier has obtained USEPA and NSF verified independent
test data indicating potential user protection against intentional or unintentional biological
contamination of drinking water. POU RO systems are not typically marketed as
microbiological water purifiers that remove bacteria and viruses from drinking water, but they
may still remove significant numbers of the microorganisms, thus offering the user a significant
level of protection. Verifications following an EPA approved test/QA plan serve to notify the
public of the possible level of protection against biological contamination agents afforded to
them by the use of a verified system.
1.3.1 Bacteria and Virus Surrogates
The expert technical panel recommended that NSF and USEPA use the bacteria Brevundimonas
diminuta (American Type Culture Collection (ATCC) strain 19146, formerly Pseudomonas
diminuta), as the surrogate for bacterial agents. This surrogate was chosen based on its small
size, as the smallest identified bacterium of concern can be as small as 0.2 |j,m in diameter. B.
diminuta has a minimum diameter of 0.2 to 0.3 |j,m (see section 5.5.3 for discussion about the
bacteria cell sizes measured in the cultures used for this verification). B. diminuta is the accepted
bacteria of choice for testing filters and membranes designed to remove bacteria. It is used in the
American Society of Testing Materials (ASTM) "Standard Test Method for Retention
Characteristics of 0.2-|o,m Membrane Filters Used in Routine Filtration Procedures for the
Evaluation of Microbiological Water Quality" (2001).
NSF also used a genetically engineered strain of B. diminuta. The NSF Microbiology
Laboratory inserted into a culture of B. diminuta strain 19146 a gene conferring resistance to the
antibiotic kanamycin. This allowed the Microbiology Laboratory to use a growth media
amended with 50 |j,g/mL of kanamycin to prohibit heterotrophic plate count (FtPC) bacteria in
the treated water samples from growing along with the kanamycin resistant B. diminuta.
The virus surrogates were the bacteriophages MS2, and fr. The ATCC designation and host E.
coli strain for each virus is given Table 1-1.
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Table 1-1. Virus and Host ATCC Designations
Virus ATCC Designation Host Bacteria ATCC Strain
MS2 ATCC 15597-B1 E. coli ATCC 15597
fr ATCC 15767-B1 E. coli ATCC 19853
The expert technical panel recommended these viruses based on their small sizes and isoelectric
points. The isoelectric point is the pH at which the virus surface is neutrally charged. MS2 is 24
nm in diameter with an isoelectric point at pH 3.9, and fr is 19 nm in diameter with an isoelectric
point at pH 8.9. With varying isoelectric points, the viruses have different surface charges, or
different strengths of negative or positive charge, depending on the pH. In solutions above the
isoelectric point, the virus is negatively charged. Below the isoelectric point, the virus is
positively charged. Using different pH settings for the virus challenges allowed an evaluation of
whether electrostatic forces enhance virus retention in mechanical filtration scenarios. The pH 6
and 9 settings were chosen because they just are beyond the upper and lower boundaries for
allowable pH in the USEPA National Secondary Drinking Water Regulations. The pH 7.5
setting was chosen because it is the midpoint between the boundaries.
The bacteria reduction challenges were performed only at pH 7.5, because the expert panel
believed that bacteria cell size and mass are too large for electrostatic interactions to play a
significant role in retention by the RO membrane
1.4 Testing Participants and Responsibilities
The ETV testing of the Watts Premier WP-4V was a cooperative effort between the following
participants:
NSF
Watts Premier, Inc.
USEPA
The following is a brief description of each of the ETV participants and their roles and
responsibilities.
1.4.1 NSF International
NSF is a not-for-profit organization dedicated to public health and safety, and to protection of the
environment. Founded in 1946 and located in Ann Arbor, Michigan, NSF has been instrumental
in the development of consensus standards for the protection of public health and the
environment. The USEPA partnered with NSF to verify the performance of drinking water
treatment systems through the USEPA's ETV Program.
NSF performed all verification testing activities at its Ann Arbor location. NSF prepared the
test/QA plan, performed all testing, managed, evaluated, interpreted, and reported on the data
generated by the testing, and reported on the performance of the technology.
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Contact Information:
NSF International
789 N. Dixboro Road
Ann Arbor, MI 48105
Phone: 734-769-8010
Fax: 734-769-0109
Contact: Bruce Bartley, Project Manager
Email: bartley@nsf.org
1.4.2 Watts Premier
The verified system is manufactured by Watts Premier, a division of Watts Water Technologies.
Watts Premier manufactures industrial, food service, POE, and POU water treatment systems
The manufacturer was responsible for supplying the test units, and for providing logistical and
technical support as needed.
Contact Information:
Watts Premier Incorporated
1725 West Williams Drive
Suite C-20
Phoenix, AZ 85027
Phone: 800-752-5582
Fax:623-931-0191
Contact Person: Mr. Bob Maisner
Email: maisnerr@wattsind.com
1.4.3 U.S. Environmental Protection Agency
USEPA, through its Office of Research and Development, has financially supported and
collaborated with NSF under Cooperative Agreement No. R-82833301. This verification effort
was supported by the DWS Center operating under the ETV Program. This document has been
peer-reviewed, reviewed by USEPA, and recommended for public release.
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Chapter 2
Equipment Description
2.1 RO Membrane Operation
Membrane technologies are among the most versatile water treatment processes with regard to
their ability to effectively remove a wide variety of contaminants. RO membranes operate by the
principal of cross-flow filtration. In this process, the influent water flows over and parallel to the
filter medium and exits the system as reject water. Under pressure, a portion of the water
diffuses through the membrane becoming "permeate". Membrane pore sizes are small enough to
reject bacteria and viruses by size exclusion, but they may still pass through imperfections in the
membrane, or go around the membrane due to microscopic seal leaks.
2.2 Equipment Capabilities
The WP-4V is certified by NSF to NSF/ANSI Standard 58 - 2006, Reverse Osmosis Drinking
Water Treatment Systems (NSF 2006). It has a certified production rate of 9.06 gallons per day
This measurement is based on system operation at 50 pounds per square inch (psi) inlet pressure,
a water temperature of 25°C, and a total dissolved solids (TDS) level of 750 + 40 mg/L. The
amount and quality of treated water produced varies depending on the inlet pressure, water
temperature, and level of TDS. These measurements were not subject to verification during this
study.
2.3 System Components
The WP-4V is a four-stage treatment system. Incoming water first passes through a sediment
filter to remove particulate matter, such as rust and silt, and then through a carbon filter to
remove chlorine or other contaminants. The third stage of treatment is the reverse osmosis
membrane, which removes a wide variety of inorganic and larger molecular weight organic
contaminants, protozoan cysts such as cryptosporidium and Giardia, and also bacteria and
viruses to some degree. The permeate water is sent to a 3-gallon maximum capacity storage
tank. Upon leaving the storage tank, the water passes through a second carbon filter to remove
organic chemicals and other taste and odor causing substances before dispensing through the
faucet. A photograph of the system is shown in Figure 2-1, and a parts diagram is shown in
Figure 2-2.
Please note that this description, and the system operation description in section 2.4 are given for
informational purposes only. This information was not subject to verification.
2.4 Other Same or Similar Models
Watts Premier markets other models that are either identical to the WP-4V, except in name, or
that are identical to the WP-4V except for different pre-membrane or post-membrane filters. The
-------
WP-4V was tested without any pre-membrane or post-membrane filters in place (see Section 3.1
for further discussion), so the results of this verification also apply to the following models:
WP-5
KP-5
RO-5M
KP-4
RO-4M
WP-4
WP-4BVC
WP—BVC-5
NSF has verified that the RO membranes in these models are identical, and function identically,
to the RO membrane in the WP-4V.
Figure 2-1. Photograph of the WP-4V System
A
WAITS
2.5 System Operation
When the flow of water into the system is started, treated water will be continually produced
until the storage tank is nearly full. At that time, the water pressure in the tank causes an
automatic shut-off valve to stop the flow of water through the system. After a portion of the
water is dispensed from the tank, the shut-off valve deactivates, allowing water to once again
flow through the RO membrane into the storage tank.
-------
The operational storage tank capacity will vary slightly from unit to unit, and may also be
affected by the inlet water pressure. The storage tank capacity was measured to be 2.64 gallons
when the system was tested for NSF/ANSI Standard 58 certification.
Figure 2-2. Parts Diagram of the WP-4V
500109(WP-4V)
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-------
2.6 Rate of Waste Production
The rate of reject water production was measured during the certification process for NSF/ANSI
Standard 58 certification. The efficiency rating, as defined by Standard 58 is the percentage
measure of the amount of influent water delivered as permeate under a closed permeate
discharge set of actual use conditions. The efficiency rating of the WP-4V is 8.4%, which means
the system produces approximately 11 gallons of reject water for each gallon of product water
produced. The efficiency rating was not verified as part of this evaluation.
2.7 Equipment Operation Limitations
Watts Premier gives the following limitations for the drinking water to be treated by the system:
• temperature of 40 - 100°F;
• pressure of 40 - 100 psig;
• pHof3-ll;
• maximum TDS level of 1,800 mg/L;
• maximum water hardness of 10 grains per gallon (1 grain per gallon equals 17.1 mg/L of
hardness, expressed as calcium carbonate equivalent) may reduce membrane life; and
• no iron present.
2.8 Operation and Maintenance Requirements
The following are the operation and maintenance requirements specified in the product owner's
manual:
• Replacement of the pre-membrane sediment and pre-membrane carbon filter every 12
months;
• Replacement of the RO membrane every 2 to 5 years (Watts Premier offers free treated
water TDS analysis for monitoring membrane operation, or the user can purchase a TDS
monitor);
• Replacement of the post-membrane carbon filter every 12 months or after 600 gallons
have been treated;
• Annual sanitization of the system with hydrogen peroxide or bleach is recommended; and
• The flow restrictor plug must be cleaned each time the RO membrane is replaced.
The WP-4V system relies on the user to determine when the filters and RO membrane need to be
replaced. There are no on-line monitors or indicators built into the system to track the volume of
water treated. However, to compensate for this, for NSF/ANSI Standard 58 certification the
post-membrane carbon filter was tested out to 200% of the claimed capacity, as opposed to 120%
of capacity for systems with volume-based monitors.
-------
Chapter 3
Methods and Procedures
3.1 Introduction
The challenge tests followed the procedures described in the Test/QA Plan for Verification
Testing of the Watts Premier WP-4V Point-of-Use Drinking Water Treatment System for
Removal of Biological Contamination Agents.
Five WP-4V systems were tested. As described in Section 2.3, the WP-4V employs an RO
membrane, a sediment filter, and carbon filters to treat drinking water. However, the systems
were tested with only the RO membrane in place. The sediment and carbon filters do not have
pore sizes small enough to remove bacteria or viruses, but they could temporarily retain
significant numbers of the organisms through electrostatic interactions, giving a positive bias to
the performance data. Otherwise the systems were operated as sold to the consumer.
3.2 Verification Test Procedure
3.2.1 Test Rig
The five systems to be tested were plumbed to a single test station such that they were all
attached to the same influent feed line. The test station used a 500-gallon polyethylene tank to
hold the influent challenge water. See Figure 3-1 for a schematic diagram of the test rig. Figure
3-2 shows the systems installed on the test rig.
3.2.2 Test Rig Sanitization
The test apparatus was sanitized prior to the installation of the test systems to keep the
heterotrophic bacteria population to a minimum. After sanitization, the test apparatus was
flushed until a less-than-detectable concentration of sanitizing agent was present.
3.2.3 Test Water
3.2.3.1 Base Water
Ann Arbor, Michigan municipal drinking water was deionized to make the base water for the
tests. The base water had the following constraints:
• Conductivity < 2 nS/cm at 25°C;
• Total chlorine < 0.05 mg/L;
• TOC< 100|^g/L;and
• Heterotrophic bacteria plate count < 100 colony forming units (CFU)/mL.
-------
The base water was then adjusted to meet the following characteristics:
• Addition of sodium bicarbonate to achieve an alkalinity (expressed as calcium carbonate)
of 100 ± 5 mg/L prior to pH adjustment;
• pH adjustment with hydrochloric acid or sodium hydroxide to reach a value of 6.0 ± 0.5,
7.5 ± 0.5, or 9.0 ± 0.5 as required by challenge protocol; and
• Temperature of 25 ± 2°C.
Sodium chloride was also added to the water to achieve a target level of 750 mg/L of TDS.
Please note that the test/QA plan did not specify that any TDS be added to the base water, and it
also specified a water temperature of 20°C instead of 25°C. The water specifications were
changed so that the testing laboratory could use the NSF/ANSI Standard 58 TDS reduction test
water, which was already being produced for other testing activities. These deviations are
discussed further in Section 5.9 on page 21.
The test water was made fresh for each challenge in 200-gallon volumes. Each batch was
analyzed for alkalinity, pH, temperature, total chlorine, total hardness, TDS, and turbidity.
Figure 3-1. Schematic Diagram of Test Rig
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10
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3.2.3.2 Bacteria and Virus Challenges
The viruses were purchased from Biological Consulting Services of North Florida, and the
bacteria from ATCC. The viruses were purchased in adequate volumes so that volumes of the
suspensions received were added directly to the base test water. The bacteria were cultivated at
NSF to obtain the challenge suspensions. Section 3.3.2.3 describes the method used to create the
bacteria challenges.
The targeted influent challenge concentrations were IxlO5 CPU per 100 milliliters, or greater, for
B. diminuta, and IxlO4 plaque forming units (PFU) per milliliter, or greater, for the fr and MS2
viruses. See Appendix A for influent challenge data.
Figure 3-2. Systems Installed on Test Rig
Separate challenges were conducted for the "normal" B. diminuta and kanamycin resistant B.
diminuta, but both viruses were mixed together into one challenge. After addition of the
challenge organism(s) to the base test water, the resultant challenge water was mixed for a
minimum of 30 minutes using a recirculation pump prior to beginning the test.
11
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3.2.4 System Operation
3.2.4.1 System Installation
The test systems were installed on a test rig by a NSF laboratory technician following the
instructions in the WP-4V owner's manual. After installation, the systems were conditioned
according to the vendor's instructions (filling the storage tanks and dispensing the contents to
drain three times) using the base test water in Section 3.2.3.1 at pH 7.5 ± 0.5. At the end of the
conditioning procedure, treated water samples were collected from each system as negative
controls and analyzed for the challenge organisms.
3.2.4.2 TDS Reduction System Check
After completion of the vendor's conditioning procedure, the membranes underwent a TDS
reduction test using the test protocol in NSF/ANSI Standard 58, modified so that the systems
were operated for only one tank-fill period. Influent water samples and treated water samples
from each system were collected and analyzed for TDS. Each system had to remove at least
75% of the TDS (the pass/fail point for NSF/ANSI Standard 58 certification) to be used for
testing. This test ensured that the products undergoing verification testing were representative of
the expected performance of the system, and that there were no membrane integrity or membrane
seal problems. All systems passed this test, see Section 4.1 for the test data.
3.2.4.3 Additional Conditioning
After the TDS reduction system check test was complete, the RO membranes were operated
using the base test water in Section 3.2.3.1 at pH 7.5 + 0.5 for 5 more days prior to challenge
testing. On each day the systems were operated continuously at a dynamic inlet pressure of 60 ±
3 psig for one tank-fill period. The systems then sat idle overnight under pressure, and the tanks
were emptied the next morning to resume system operation.
Previous POU RO system ETV tests indicated that perhaps membrane performance does not
stabilize until after four or five days, or four or five tank fills, of conditioning. Five extra days of
conditioning ensured that the membranes were performing optimally prior to the chemical
challenges.
3.2.4.4 Challenge Testing
Following the conditioning period, the RO membranes were challenged according to the
schedule in Table 3-1. The test plan called for both the normal B. diminuta and kanamycin
resistant B. diminuta challenges to be conducted prior to beginning the virus challenges, but the
kanamycin resistant B. diminuta challenge was delayed to give the Microbiology Lab more time
to grow the challenge suspensions.
12
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Table 3-1. Challenge Schedule
Day Surrogate Challenge pH
1
2
o
J
4
5
B. diminuta
frandMS2
frandMS2
Kanamycin resistant
B. diminuta
frandMS2
7.5 ±0.5
6.0 ±0.5
7.5 ±0.5
7.5 ±0.5
9.0 ±0.5
At the end of the workday before each challenge, or the morning of the challenge, a tank of the
test water without challenge organisms was prepared as described in section 3.2.3.1. Prior to
beginning each challenge, the pH was checked and adjusted, if necessary, and the bacteria or
viruses were added as described in section 3.2.3.2.
Influent samples were collected at the start and end of each challenge for bacteria or virus
enumeration, and for water chemistry analysis. Each system was operated continuously for one
tank-fill period (approximately 4 to 5 hours).
After all systems shut off, the storage tanks were emptied into separate sterile containers, and
samples were collected in sterile polypropylene bottles for challenge organism enumeration. The
sample volumes were 1 L for the bacteria challenges, and 150 mL for virus challenges. All
samples for bacteria or virus enumeration were enumerated in triplicate.
Following each challenge, the systems were flushed for one tank-fill period using the base test
water without the test organism(s) included. The systems rested under pressure overnight, and
the morning of the next challenge the storage tanks were emptied into sterile containers, and
negative control samples were collected for analysis of that day's challenge organism(s). The
negative control samples for the first B. diminuta challenge were collected after the last day of
the conditioning period.
3.3 Analytical Methods
3.3.1 Water Quality Analytical Methods
The following are the analytical methods used during verification testing. All analyses followed
procedures detailed in NSF Standard Operating Procedures (SOPs).
• Alkalinity was measured according to EPA Method 310.2 with the SmartChem Discrete
Analyzer. Alkalinity will be expressed as mg/L CaCO3.
• pH measurements were made with a Beckman 350 pH meter. The meter was operated
according to the manufacturer's instructions, which are based on Standard Method 4500-
H+.
• Water temperature was measured using an Omega model HH11 digital thermometer, or
equivalent.
13
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• IDS for the IDS reduction system check test was measured through conductivity
according to Standard Method 2510 using a Fisher Scientific Traceable™ Conductivity
Meter. This method has been validated for use with the test water; NSF uses this method
for analysis of samples from TDS reduction tests under NSF/ANSI Standard 58.
• Total chlorine was measured according to Standard Method 4500-C1 G with a Hach
Model DR/2010 spectrophotometer using AccuVac ampules.
• Total Hardness was measured according to USEPA method 310.1 using the SmartChem
Discrete Analyzer.
• Turbidity was measured according to Standard Method 2130 using a Hach 2100N
turbidimeter.
3.3.2 Microbiology Analytical Methods
3.3.2.1 Sample Processing, and Enumeration of Viruses
The viruses were enumerated using a double agar layer method published in NSF/ANSI Standard
55 - 2004, Ultraviolet Microbiological Water Treatment Systems for enumerating MS2. This
method is similar to the double agar layer method in USEPA Method 1601.
Four to eighteen hours prior to sample processing, 100 |jL of the appropriate host E.coli
suspension was pipetted into tubes containing 10 mL of fresh Tryptic Soy Broth (TSB), and
incubated at 35 °C. After incubation, 100 |jL volumes of the resulting E. coli culture were
transferred to sterile, capped test tubes.
All samples were enumerated in triplicate. All samples were serially diluted for enumeration,
and the effluent samples were also enumerated directly. One milliliter volumes of the sample or
dilution were pipetted into the E. coli suspension test tubes. The tubes were vortexed for a
minimum of 30 seconds to "mate" the bacteria and virus, and then 4 mL of molten, tempered
TSB plus 1% agar was added to each tube. These mixtures were then poured over Tryptic Soy
Agar (TSA) plates, and allowed to solidify. The plates were incubated at 35°C for 18-24 hours.
Virus plaques were counted using a Quebec Colony Counter.
3.3.2.2 Bacteria Cultivation
The bacteria were purchased from ATCC and rehydrated with nutrient broth. After 48 hours of
incubation at 30°C, tubes containing 10 mL of TSB were inoculated with 100 |jL of the nutrient
broth suspension. These tubes were incubated for 48 hours at 30°C. After this incubation
period, 100 |jL of these suspensions were pipetted into new tubes containing 10 mL of fresh
TSB. These tubes were then also incubated for 48 hours at 30°C. This process was repeated at
least three times, up to a maximum of 30 times.
3.3.2.3 Preparation of B. diminuta Challenge Suspensions
To obtain the challenge suspensions, 1 mL of a 48-hour TSB culture was pipetted into an
appropriate volume of Saline Lactose Broth (SLB). The SLB culture was incubated in a shaking
14
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water bath at 30 °C for 48 hours. Cells were harvested after centrifugation at 3,000 revolutions
per minute for 10 minutes. The supernatant was discarded and the pellet was resuspended in 100
mL of phosphate buffered dilution water (PBDW). The resulting challenge suspensions were
refrigerated and added to the tank of test water within one hour. Samples of the challenge
suspension were collected and enumerated according to the method in section 3.3.2.4.
The challenge preparation procedure was identical for both the normal B. diminuta and the
kanamycin resistant B. diminuta, the only difference was that for the kanamycin resistant
bacteria, the SLB was amended with 50 ng/L of kanamycin, and 10 [ig/L of tetracycline.
3.3.2.4 Sample Processing and Enumeration of B. diminuta
All samples were enumerated in triplicate using a membrane filtration method based on Standard
Method 9215 D. All samples were serially diluted for enumeration with sterile PBDW, and the
effluent samples were also enumerated directly. For the influent samples, 1 mL volumes of
either the straight sample or dilutions were pipetted into sterile glass vacuum filtration funnels,
and 25 mL of PBDW was also poured into the funnels. For the effluent samples, 100 mL of the
straight sample and the dilutions were pipetted into the funnels. The contents were then vacuum
filtered through sterile 0.1 jam membrane filters. The funnels were rinsed three times with
approximately 5 mL of PBDW, and the rinse water was also suctioned through the filters. The
membrane filters were aseptically removed from the apparatuses and placed onto R2A agar
plates. The plates were incubated at 30°C for 48 hours. Characteristic B. diminuta colonies were
counted with a Quebec Colony Counter.
The sample processing and enumeration procedures were identical for both the normal B.
diminuta and the kanamycin resistant B. diminuta, the only difference was that the R2A agar was
amended with 50 (ig/L of kanamycin, and 10 (ig/L of tetracycline for enumeration of the
kanamycin resistant bacteria.
15
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Chapter 4
Results and Discussion
4.1 TDS Reduction
The performance data from the TDS reduction system check test described in 3.2.4.2 are
presented in Table 4-1. The certified TDS reduction for the WP-4V is 97%. The five units did
not meet that percent reduction, but they did all reduce the TDS of the challenge water by greater
than 75%, thus meeting the requirement in the test/QA plan for use of each unit in the bacteria
and virus challenges..
Table 4-1. TDS Reduction Test Results
TDS (mg/L) Percent Reduction
Influent
Effluents:
Unitl
Unit 2
Units
Unit 4
Unit5
770
66
44
40
86
30
91
94
95
89
96
4.2 Bacteria Reduction
Presented in Table 4-2 are the logic reduction data for the B. diminuta challenges. The influent
and effluent triplicate bacteria counts are presented in Appendix A. The triplicate influent and
effluent counts were averaged by calculating geometric means. The means were then logio
transformed and Iog10 reduction values were calculated for each test unit.
The challenge organisms were detected in the effluents for all units in both challenges except for
Unit 2 in the "normal" B. diminuta challenge. Since the Unit 2 effluent count for kanamycin
resistant B. diminuta was 4.3 logs, and all other effluent samples had bacteria counts greater than
IxlO4, it is possible that there was a sampling or analysis error associated with the Unit 2
"normal" B. diminuta sample. Therefore, that sample was not included in the mean logic
reduction calculation for the B. diminuta challenge.
The minimum log reduction observed was 1.3, which equates to a 95% removal of the bacteria.
The maximum observed log reduction, excluding the "non-detect" 6.4 logic removal, was 3.1,
equaling 99.9% removal. The geometric mean logic reduction for both challenges combined was
1.9.
All negative control samples were non-detect for the bacteria.
16
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Table 4-2. Bacteria Log Reduction Data
Log10 Geometric Mean Logio Reductions
Influent Geometric Mean of
Challenge Organism Challenge Unit 1 Unit 2 Unit 3 Unit 4 Unit 5 the 5 Units
B. diminuta 6.4 1.8 6.4* 1.3 1.5 1.6
Kanmycin Resistant ?2 J4 ^ ^ ^ 31
B. diminuta
1.5
2.4
Overall Geometric Mean: 1.9
*Number not included in mean log reduction calculation.
4.3 Virus Reduction
The virus Iog10 reduction data are presented in Table 4-3. The influent and effluent triplicate
PFU counts are presented in Appendix A. As was done for the bacteria, the triplicate influent
and effluent counts were averaged by calculating geometric means. The means were then Iog10
transformed and logio reduction values calculated for each test system.
The minimum observed logio reduction was 1.2, and the maximum observed logio reduction was
3.7. These logio reductions correspond to percent reductions of 94% and 99.98%, respectively.
The overall geometric mean logio reductions were 2.5 for fir and 2.6 for MS2.
A visual comparison of the logio reductions versus the challenge water pH shows the mean logio
reductions decreasing with increasing pH. However, an examination of the 95% confidence
intervals around the means (see Appendix A for data) shows that the decreases are not
statistically significant.
All negative control samples were non-detect for the viruses.
Table 4-3. Virus Log Reduction Data
Initial Final Logio Geometric Mean Logio Reduction
Measured Measured Challenge Influent Geometric Mean
Target pH pH pH Organisms Challenge Unit 1 Unit 2 Unit 3 Unit 4 Unit 5 of the 5 Units
6.0 ±0.5 6.1
7.5 ±0.5 7.5
9.0 ±0.5 8.9
6
7
9
* The mean influent challenj
IxlO4 PFU/mL requirement,
.5
.7
.0
fr
MS2
fr
MS2
fr
MS2
3
3
4
4
5
4
9*
.8*
.5
.2
.0
.6
1.8
2.3
1.9
1.7
1.4
1.2
3.1
3.4
2.4
2.4
2.3
2.4
3.6
3.7
2.3
2.4
2.1
2.0
3.4
3.6
3.1
3.4
2.3
2.3
Overall
3.0
2.9
2.8
3.2
2.6
3.0
Means: fr
MS2
;e did not meet the 4 logio requirement. The start-up influent samples were
but the end-of-challenge influent samples were not.
2.9
3.1
2.5
2.5
2.1
2.1
2.5
2.6
above
17
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Chapter 5
QA/QC
5.1 Introduction
An important aspect of verification testing is the QA/QC procedures and requirements. Careful
adherence to the procedures ensured that the data presented in this report was of sound quality,
defensible, and representative of the equipment performance. The primary areas of evaluation
were representativeness, precision, accuracy, and completeness.
Because the ETV was conducted at the NSF testing lab, all laboratory activities were conducted
in accordance with the provisions of the NSF International Laboratories Quality Assurance
Manual (NSF 2004).
5.2 Test Procedure QA/QC
NSF testing laboratory staff conducted the tests by following a USEPA-approved test/QA plan
created specifically for this verification. NSF QA Department Staff performed an informal audit
during testing to ensure the proper procedures were followed. The audit yielded no significant
findings.
5.3 Sample Handling
All samples analyzed by the NSF Chemistry Laboratory were labeled with unique ID numbers.
These ID numbers appear in the NSF laboratory reports for the tests. All samples were analyzed
within allowable holding times.
5.4 Chemistry Analytical Methods QA/QC
The calibrations of all analytical instruments, and the analyses of all parameters complied with
the QA/QC provisions of the NSF International Laboratories Quality Assurance Manual.
The NSF QA/QC requirements are all compliant with those given in the USEPA method or
Standard Method for the parameter. Also, every analytical instrument has an NSF SOP
governing its use.
5.5 Microbiology Laboratory QA/QC
5.5.1 Growth Media Positive Controls
All media were checked for sterility and positive growth response when prepared and when used
for microorganism enumeration. The media was discarded if growth occurred on the sterility
check media, or if there was an absence of growth in the positive response check. Both E. coli
hosts for the viruses were plated on TSA and incubated with the virus enumeration plates during
18
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sample enumeration as a second positive growth control. B. diminuta from the stock cultures
was plated on R2A agar and incubated with the bacteria enumeration plates as a positive control.
5.5.2 Negative Controls
All samples were enumerated in triplicate. For each sample batch processed, an unused
membrane filter and a blank with 100 mL of PBDW filtered through the membrane were also
placed onto the appropriate media and incubated with the samples as negative controls. No
growth was observed on any blanks.
5.5.3 Bacteria Cell Size
The theoretical minimum size for B. diminuta cells is 0.2 to 0.3 jam in diameter. Using the
accepted method of growth in SLB media to obtain smaller cell sizes, the NSF Microbiology lab
was able to achieve cells less than 0.5 jam in diameter. Samples from the stock cultures of both
the normal B. diminuta and kanamycin resistant B. diminuta were examined microscopically
with a Zeiss Axioskop 2 Plus, and measurements were taken using the accompanying Axiovision
computer program. The measurements for each culture are presented below in Table 5-1.
Table 5-1. Bacteria Cell Size Measurements
Normal^, diminuta Kanamycin Resistant B.
Diameter (|J.m) diminuta Diameter (|J.m)
Average:
Standard Deviation:
0.47
0.38
0.34
0.44
0.34
0.39
0.06
0.34
0.34
0.32
0.30
0.24
0.31
0.04
5.6 Documentation
All laboratory activities were documented using specially prepared laboratory bench sheets and
NSF laboratory reports. This documentation can be found in the appendices. Data from the
bench sheets and laboratory reports were entered into Excel spreadsheets. These spreadsheets
were used to calculate average influents and effluents, and logic reductions for each challenge.
One hundred percent of the data entered into the spreadsheets was checked by a reviewer to
confirm all data and calculations were correct.
5.7 Data Review
NSF QA/QC staff reviewed the raw data records for compliance with QA/QC requirements.
NSF ETV staff checked 100% of the data in the NSF laboratory reports against the lab bench
sheets.
19
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5.8 Data Quality Indicators
The quality of data generated for this ETV is established through four indicators of data quality:
representativeness, accuracy, precision, and completeness.
5.8.1 Representativeness
Representativeness refers to the degree to which the data accurately and precisely represent the
expected performance of the RO system under normal use conditions. The test protocol was
designed to be a conservative evaluation of product performance. The test water was of very low
turbidity to minimize the potential for microbial adhesion to suspended particles, which could
enhance apparent log reduction. The surrogates were chosen because of their small size. The
virus surrogate challenges were carried out at pH 6, 7.5, and 9 to assess whether pH affects the
performance of the RO membrane.
Representativeness was ensured by consistent execution of the test protocol for each challenge,
including timing of sample collection, sampling procedures, and sample preservation.
Representativeness was also ensured by using each analytical method at its optimum capability
to provide results that represent the most accurate and precise measurement it is capable of
achieving.
5.8.2 Accuracy
Accuracy was quantified as the percent recovery of the parameter in a sample of known quantity.
Accuracy was measured through use of both matrix spikes of a known quantity, and certified
standards during calibration of an instrument. The following equation was used to calculate
percent recovery:
Percent ReCOVery = 100 X [(Xknown - Xmeasured)/Xknown]
where: Xkn0wn = known concentration of the measured parameter
= measured concentration of parameter
Accuracy of the benchtop chlorine, pH, TDS, and turbidity meters was checked daily during the
calibration procedures using certified check standards. Alkalinity and total hardness were
analyzed in batches. Certified QC standards and/or matrix spikes were run with each batch.
The percent recoveries of all matrix spikes and standards were within the allowable limits for all
analytical methods.
5.8.3 Precision
Precision refers to the degree of mutual agreement among individual measurements and provides
an estimate of random error. One sample per batch was analyzed in duplicate for the TDS
measurements. Duplicate municipal drinking water samples were analyzed for pH, total
20
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chlorine, and turbidity as part of the daily calibration process. One out of every ten samples for
alkalinity and total hardness was analyzed in duplicate. Precision of duplicate analyses was
measured by use of the following equation to calculate relative percent difference (RPD):
RPD =
:200
where:
Sl = sample analysis result; and
^2 = sample duplicate analysis result.
All RPDs were within NSF's established allowable limits for each parameter. Please note that
samples from this evaluation for alkalinity, IDS, and total hardness were batched with other
non-ETV samples. The duplicate analysis requirements apply to the whole batch, not just the
samples from this ETV.
5.8.4 Completeness
Completeness is the proportion of valid, acceptable data generated using each method as
compared to the requirements of the test/QA plan. The completeness objective for data
generated during verification testing is based on the number of samples collected and analyzed
for each parameter and/or method.
Table 5-2. Completeness Requirements
Number of Samples per Parameter _ ^ „ , ^
j / A , 5 j Percent Completeness
and/or Method
(MO 80%
11-50 90%
> 50 95%
Completeness is defined as follows for all measurements:
%C = (V/T)X100
where:
%C = percent completeness;
V = number of measurements judged valid; and
T = total number of measurements.
5.8.4.1 Completeness Measurements
• Five systems were tested, as called for in the test/QA plan, giving a completeness
measurement of 100% for this category.
21
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• All planned water chemistry samples were collected and analyzed, except that total
chlorine was not analyzed on day three of the conditioning period. A total of 15 samples
were to be collected for total chlorine over the course of the evaluation. The one missed
sample gives a completeness of 93%, which is acceptable.
• All scheduled bacteria and virus samples were collected and analyzed with acceptable
results.
5.9 Measurements Outside of the Test/QA Plan Specifications
As discussed in section 3.2.3.1, the test water used for this evaluation was the NSF/ANSI
Standard 58 TDS reduction test water. This water differed from the water called for in the
test/QA plan in that it had sodium chloride added for TDS, and the temperature was 25 °C
instead of 20° C. These changes did not significantly affect the viability of the challenge
organisms, since there was no significant decrease in organism concentrations from the first to
the second influent samples (see Appendix A).
All other water chemistry measurements were within the allowable ranges.
The second influent samples for both viruses during the pH 6 virus challenge were below the
minimum target level of IxlO4 PFU/mL (see Appendix A). However, the first influent samples
were above the target level. The low second influent sample levels caused the overall mean
influent to be below IxlO4 PFU/mL, but the low influents did not limit the logic reduction
numbers, since there were no effluents with virus counts <1 PFU/mL.
22
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Chapter 6
References
American Society of Testing Materials (2001). D 3862-80, Standard Test Method for Retention
Characteristics of 0.2-|j,m Membrane Filters Used in Routine Filtration Procedures for the
Evaluation of Microbiological Water Quality.
NSF International (2004). NSF International Laboratories Quality Assurance Manual. Ann
Arbor, NSF International.
NSF International (2004). NSF/ANSI55-2004, Ultraviolet microbiological water treatment
systems. Ann Arbor, NSF International.
NSF International (2006). NSF/ANSI 58 - 2006, Reverse osmosis drinking water treatment
systems. Ann Arbor, NSF International.
USEPA (2004). Water Security Research and Technical Support Action Plan. EPA/600/R-
04/063.
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Appendix A
Bacteria and Virus Counts, and Water Chemistry Data
-------
Bacteria Challenges Data
Brevundimonas diminuta
Sample
First Influent
Second Influent
Influents Combined
Effluents:
Unitl
Unit 2
Units
Unit 4
Unit5
*Number not
Influent/Effluent Triplicate
Counts (CFU/lOOmL)
2.9xl06, 2.3xl06, 1.9xl06
2.0xl06, 2.4xl06, 2.3xl06
8xl04, 4xl04, 2xl04
-------
Virus Challenges at pH 6
Sample
First Influent
Second Influent
Influents Combined
Effluents:
Unitl
Unit 2
Units
Unit 4
Unit5
Influent/Effluent Triplicate
Counts (PFU/mL)
LlSxlO4, 1.21xl04, 1.16x10
4.2xl03, 5.1xl03, 4.6xl03
98, 142, 111
6,4,9
1,3,5
5,3,2
12, 5, 6
fr
Log10
Influent/Effluent Influent/
Geometric Mean (PFU/mL) Effluent
4 1.17xl04
4.6xl03
7.3xl03
117
6
2
o
J
7
95%
3.9
2.1
0.8
0.3
0.5
0.9
Overall Mean
Standard Deviation
Confidence Interval
Log10
Reduction
1.8
3.1
3.6
3.4
3.0
2.9
0.7
1.7-4.1
Sample
First Influent
Second Influent
Influents Combined
Effluents:
Unitl
Unit 2
Units
Unit 4
Unit5
Influent/Effluent Triplicate
Counts (PFU/mL)
LlSxlO4, 1.02xl04, 1.08x10
4.3xl03, 3.9xl03, 3.7xl03
29, 34, 39
3,4,2
1,2,1
4, 1,1
10, 6, 8
MS2
Log10
Influent/Effluent Influent/
Geometric Mean (PFU/mL) Effluent
4 1.09xl04
4.0xl03
6.6xl03
34
3
1
2
8
95%
3.8
1.5
0.5
0.1
0.2
0.9
Overall Mean
Standard Deviation
Confidence Interval
Log10
Reduction
2.3
3.4
3.7
3.6
2.9
3.1
0.6
2.1-4.2
A-2
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Virus Challenges at pH 7.5
fr
Sample
First Influent
Second Influent
Influents Combined
Effluents:
Unitl
Unit 2
Units
Unit 4
Unit5
Influent/Effluent Triplicate
Counts (PFU/mL)
3.2xl04, 7.1xl04, 2.9xl04
2.1xl04,2.6xl04,2.4xl04
302,416,512
131,121, 142
112,143, 190
17, 28, 35
54, 49, 38
Influent/Effluent
Geometric Mean (PFU/mL)
4.0xl04
2.4xl04
3.1xl04
401
131
145
26
47
Log10
Influent/
Effluent
4.5
2.6
2.1
2.2
1.4
1.7
Overall Mean
Standard
Deviation
95% Confidence Interval
Log10
Reduction
1.9
2.4
2.3
3.1
2.8
2.5
0.5
1.7-3.3
MS2
Sample
First Influent
Second Influent
Influents Combined
Effluents:
Unitl
Unit 2
Units
Unit 4
Unit5
Influent/Effluent Triplicate
Counts (PFU/mL)
1.21xl04, 1.37xl04, 1.61xl04
9.1xl03,2.0xl04,2.9xl04
322,260,310
67, 75, 60
45,75,78
6, 4, 12
5, 12, 16
Influent/Effluent
Geometric Mean (PFU/mL)
1.39xl04
1.7xl04
1.6xl04
296
67
64
7
10
Log10
Influent/
Effluent
4.2
2.5
1.8
1.8
0.8
1.0
Overall Mean
Standard
Deviation
95% Confidence Interval
Log10
Reduction
1.7
2.4
2.4
3.4
3.2
2.5
0.7
1.3-3.7
A-3
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Virus Challenges at pH 9
fr
Sample
First Influent
Second Influent
Influents Combined
Effluents:
Unitl
Unit 2
Units
Unit 4
Unit5
Influent/Effluent Triplicate
Counts (PFU/mL)
1.39xl05, 1.05xl05, 1.12xl05
7.7xl04, 6.3xl04, 8.2xl04
5.3xl03, 3.5xl03, 3.2xl03
443, 409, 600
990, 636, 572
672, 440, 492
290,220,216
Log10
Influent/Effluent Influent/
Geometric Mean (PFU/mL) Effluent
l.lSxlO5
7.4xl04
9.3xl04 5.0
3.9xl03 3.6
477 2.7
711 2.9
526 2.7
240 2.4
Mean
Standard Deviation
95% Confidence Interval
Log10
Reduction
1.4
2.3
2.1
2.3
2.6
2.1
0.5
1.3-2.9
MS2
Sample
First Influent
Second Influent
Influents Combined
Effluents:
Unitl
Unit 2
Units
Unit 4
Unit5
Influent/Effluent Triplicate
Counts (PFU/mL)
9.4xl04, 5.7xl04, 6.5xl04
2.9xl04, 1.5xl04, 3.7xl04
2.9xl03, 2.1xl03, 2.8xl03
182, 116,210
348, 500, 308
110,220,244
44, 28, 42
Log10
Influent/Effluent Influent/
Geometric Mean (PFU/mL) Effluent
7.0xl04
2.5xl04
4.2xl04 4.6
2.6xl03 3.4
164 2.2
377 2.6
181 2.3
37 1.6
Mean
Standard Deviation
95% Confidence Interval
Log10
Reduction
1.2
2.4
2.0
2.3
3.0
2.1
0.7
0.9-3.3
A-4
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RO Membrane Conditioning Water Chemistry Data
Sample Day 1 Day 2 Day 3 Day 4 Day 5
Alkalinity (mg/L CaCO3)
pH
Temperature (°C)
Total Chlorine (mg/L)
Total Hardness (mg/L CaCO3)
TDS (mg/L)
Turbidity (NTU)
# Sample not collected
100 100
8.0 7.5
25 26
ND(0.05) ND(0.05)
6 6
870 830
0.1 ND(O.l)
100
7.7
25
#
8
670
0.1
97 100
7.9 7.5
26 25
ND(0.05) ND(0.05)
6 ND(2)
780 880
ND(O.l) ND(O.l)
RO Membrane Challenges Water Chemistry Data
Sample
Kanamycin
Resistant B.
B. diminuta diminuta
pH6
Viruses
pH7.5
Viruses
pH9
Viruses
Start-up Influent
Alkalinity (mg/L CaCO3) 100 100 61 97 120
pH 7.6 7.5 6.1 7.5 8.9
Temperature (°C) 25 26 25 25 26
Total Chlorine (mg/L) ND (0.05) ND (0.05) ND (0.05) ND (0.05) ND (0.05)
Total Dissolved Solids (mg/L) 850 870 880 860 870
Total Hardness (mg/L CaCO3) 8 ND (2) ND (2) ND (2) ND (2)
Turbidity (NTU) ND(O.l) ND(O.l) ND (0.1) ND (0.1) ND (0.1)
2nd Influent
pH 7.82 7.8 6.5 7.7 9.0
Temperature (°C) 25 24 24 24 25
Turbidity (NTU) (XI (XI (XI (XI 0.8
A-5
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