Method 8506

COPPER (0 to 5.00 mg/L) For water, wastewater and seawater*

Bicinchoninate Method** (Powder Pillows or AccuVac Ampuls);
USEPA approved for reporting wastewater analysis (digestion needed; See Section 2)***
Using Powder Pillows

1. Enter the stored 2. Press: 20 ENTER 3. Fill a sample cell 4. Place the blank into
program number for The display will show with 10 mL of sample the cell holder. Tightly
bicinchoninate copper mg/L, Cu and the (the blank). cover the sample cell
(Cu)- powder pillows. ZERO icon. Note: Adjust the pH of with the instrument cap.
Press: PRGM Note: Determination of acid-preserved samples to
total copper needs a prior 4-6 with 8 N KOH before
The display will show: analysis. Do not exceed pH
digestion (see Digestion in
PRGM ? Section 2). 6 or copper may precipitate.
Note: For most accurate
results, perform a Reagent
Blank Correction using
deionized water (see
Section 1).

* Pretreatment required; see Interferences (Using Powder Pillows)

** Adapted from Nakano, S., Yakugaku Zasshi, 82 486-491 (1962) [Chemical Abstracts, 58 3390e (1963)]
*** Powder Pillows only: Federal Register, 45 (105) 36166 (May 29, 1980)

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COPPER, continued

5. Press: ZERO 6. Fill another sample 7. Add the contents of 8. Press:

The cursor will move to cell with 10 mL of one CuVer 1 Copper TIMER ENTER
the right, then the the sample. Reagent Powder Pillow
A two-minute reaction
display will show: to the sample cell (the
period will begin.
prepared sample). Swirl
0.00 mg/L Cu Note: Accuracy is not
the cell to mix.
Note: If Reagent Blank affected by
Note: If copper is present,
Correction is on, the undissolved powder.
A purple color will develop.
display may flash “limit”.

9. Within 30 minutes 10. Press: READ

after the timer beeps, The cursor will move to
place the prepared the right, then the result
sample into the cell in mg/L copper will be
holder. Tightly cover the displayed.
sample cell with the
Note: Standard Adjust may
instrument cap. be performed using a
prepared standard (see
Section 1).

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COPPER, continued
Using AccuVac Ampuls Method 8026

1. Enter the stored 2. Press: 20 ENTER 3. Fill a sample cell 4. Place the blank into
program number for The display will show with at least 10 mL of the cell holder. Tightly
bicinchoninate copper mg/L, Cu and the sample (the blank). cover the sample cell
(Cu)- AccuVac ampuls. ZERO icon. Collect at least 40 mL of with the instrument cap.
Press: PRGM sample in a 50-mL
Note: Determination of
total copper needs a prior beaker.
The display will show:
digestion (see Digestion in
PRGM ? Section 2).
Note: For most accurate Note: Adjust the pH of
results, perform a Reagent stored samples before
Blank Correction using analysis.
deionized water (see
Section 1).

5. Press: ZERO 6. Fill a CuVer 2 7. Quickly invert the 8. Press:

The cursor will move to Copper Reagent ampul several times to TIMER ENTER
the right, then the AccuVac Ampul with mix. Wipe off any
A two-minute reaction
display will show: sample. liquid or fingerprints.
period will begin.
0.00 mg/L Cu Note: Keep the tip Note: A purple color will
immersed while the ampul form if copper is present.
Note: If Reagent Blank fills completely.
Correction is on, the Note: Accuracy is not
display may flash “limit”. affected by undissolved
powder

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COPPER, continued

9. After the timer 10. Press: READ

beeps, place the The cursor will move to
AccuVac ampul in the the right, then the result
cell holder. Tightly in mg/L copper (Cu) will
cover the sample cell be displayed.
with the instrument cap.
Note: Standard Adjust may
Note: Step 10 must be be performed using a
completed within 30 prepared standard (see in
minutes after the Section 1).
timer beeps.

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COPPER, continued
Sampling and Storage
Collect samples in acid-cleaned glass or plastic containers.
Adjust the pH to 2 or less with nitric acid (about 2 mL per liter).
Store preserved samples up to six months at room temperature.
Before analysis, adjust the pH to 4 to 6 with 8 N potassium
hydroxide. Do not exceed pH 6, as copper may precipitate.
Correct the test result for volume additions; see Correction for
Volume Additions in Section 1 for more information. If only
dissolved copper is to be determined, filter the sample before acid
addition using the labware listed under Optional Apparatus.
Accuracy Check
Standard Additions Method
a) Fill three 25-mL graduated mixing cylinders with 25 mL of
sample.

b) Snap the neck off a Copper Voluette Ampule Standard, 75

mg/L
as Cu.

c) Use the TenSette Pipet to add 0.1, 0.2, and 0.3 mL of

standard, respectively, to the mixing cylinders. Stopper
and mix thoroughly.

d) For analysis with AccuVac Ampuls, transfer the solutions

to dry, clean 50-mL beakers to fill the ampules. For
analysis with powder pillows, transfer only 10 mL of the
solution to 10-mL sample cells.

e) Analyze each sample as described in the procedure. The

copper concentration should increase about 0.3 mg/L for
each 0.1 mL of standard added.

f) If these increases do not occur, see Standard Additions in

Section 1 for more information.
Standard Solution Method
Prepare a 1.00 mg/L copper standard by pipetting 1.00 mL of
Copper Standard Solution, 100 mg/L as Cu, into 100-mL
volumetric flask. Dilute to volume with deionized water and mix
well. Prepare this solution daily. Using this solution as the
sample, perform the copper procedure as described above.
Method Performance
Precision
In a single laboratory, using a standard solution of 2.25 mg/L Cu

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COPPER, continued
and two representative lots of reagent with the instrument, a
single operator obtained a standard deviation of ±0.02 mg/L Cu.

In a single laboratory, using a standard solution of 2.25 mg/L Cu

and two representative lots of AccuVac Ampuls with the
instrument, a single operator obtained a standard deviation of
±0.02 mg/L Cu.

Estimated Detection Limit (EDL)

The EDL for program 20 (Powder Pillows and AccuVac Ampuls)
is 0.02 mg/L Cu. For more information on derivation and use of
Hach’s estimated detection limit, see Section 1.

Interferences
Interfering Substances and Suggested Treatments for Powder
Pillows
Interfering
Interference Level and Treatment
Substance

Acidity If the sample is extremely acidic (pH 2 or less) a

precipitate may form. Add 8 N Potassium Hydroxide
Standard Solution drop-wise while swirling to dissolve the
turbidity. Continue with Step 3.
Aluminum, Al3+ Follow the powder pillow procedure above, but substitute a
CuVer 2 Copper Reagent Powder Pillow for the CuVer 1
Pillow used in Step 4. Results obtained will include total
dissolved copper (free and complexed).
Cyanide, CN- Prevents full color development. Add 0.2 mL of
formaldehyde to the 10-mL sample. Wait 4 minutes before
taking the reading. Multiply the test results by 1.02 to
correct for sample dilution by the formaldehyde.
Hardness Follow the powder pillow procedure above, but substitute a
CuVer 2 Copper Reagent Powder Pillow for the CuVer 1
Pillow used in Step 4. Results obtained will include total
dissolved copper (free and complexed).
Iron, Fe3+ Follow the powder pillow procedure above, but substitute a
CuVer 2 Copper Reagent Powder Pillow for the CuVer 1
Pillow used in Step 4. Results obtained will include total
dissolved copper (free and complexed).
Silver, Ag+ If a turbidity remains and the precipitate turns black, silver
interference is likely. Add 10 drops of saturated Potassium
Chloride Solution to 75 mL of sample, followed by filtering
through a fine or highly retentive filter. Use the filtered
sample in the procedure.

To differentiate free copper from that complexed to EDTA or

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COPPER, continued
other complexing agents, use a Free Copper Reagent Powder
Pillow in place of the CuVer 1 pillow in Step 4. Results in Step
10 will be free copper only. Add a Hydrosulfite Reagent Powder
Pillow to the same sample and
re-read the result. This result will include the total dissolved
copper (free and complexed).
Interfering Substances and Suggested Treatments for AccuVac
Ampuls
Interfering
Interference Level and Treatment
Substance

Acidity If the sample is extremely acidic (pH 2 or less) a

precipitate may form. Add 8 N Potassium Hydroxide
Standard Solution drop-wise until sample pH is above 4.
Continue with Step 3.
Aluminum, Al3+ Reagents accommodate high levels.
Cyanide, CN- Prevents full color development. Add 1.0 mL of
formaldehyde to a 50-mL sample. Wait 4 minutes before
taking the reading. Multiply the test results by 1.02 to
correct for sample dilution by the formaldehyde.
Hardness Reagents accommodate high levels
3+
Iron, Fe Reagents accommodate high levels
Silver, Ag+ If a turbidity remains and the precipitate turns black, silver
interference is likely. Add 10 drops of saturated Potassium
Chloride Solution to 75 mL of sample, followed by filtering
through a fine or highly retentive filter. Use the filtered
sample in the procedure.

Unlike CuVer 1 Reagent, CuVer 2 Reagent reacts directly with

copper which is complexed by chelants such as EDTA. If free
copper is to be determined separately from complexed copper,
see the Powder Pillow Interference section above.

Summary of Method
Copper in the sample reacts with a salt of bicinchoninic acid
contained in CuVer 1 or 2 Copper Reagent to form a purple
colored complex in proportion to the copper concentration. This
method includes procedures for both powder pillow and AccuVac
reagents.

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COPPER, continued
REQUIRED REAGENTS & APPARATUS (Using Powder Pillows)
Quantity Required
Description Per Test Unit Cat. No.
CuVer 1 Copper Reagent Powder Pillows ...............1 pillow ............ ...100/pkg.......... 21058-69
Sample Cell, 10-20-25 mL, w/cap ................................ 2......................... 6/pkg.......... 24019-06

REQUIRED REAGENTS & APPARATUS (Using AccuVac Ampuls)

CuVer 2 Copper Reagent AccuVac Ampuls ............1 ampul ................. 25/pkg.......... 25040-25
Beaker, 50 mL............................................................... 1........................... each........... 500-41H
OPTIONAL REAGENTS
Copper Standard Solution, 100 mg/L ....................................................100 mL.............. 128-42
Copper Standard Solution, Voluette Ampule, 75 mg/L Cu, 10 mL........ 16/pkg.......... 14247-10
CuVer 2 Reagent Powder Pillows ........................................................ 100/pkg.......... 21882-99
Formaldehyde, 37%, ACS .........................................................100 mL* MDB............ 2059-32
Free Copper Reagent Powder Pillows .................................................. 100/pkg.......... 21186-69
Hydrochloric Acid Solution, 6.0 N........................................................500 mL.............. 884-49
Hydrosulfite Reagent Powder Pillows.................................................. 100/pkg.......... 21188-69
Metals Drinking Water Standard, LR for Cu, Fe, Mn ...........................500 mL.......... 28337-49
Metals Drinking Water Standard, HR for Cu, Fe, Mn ...........................500 mL.......... 28336-49
Nitric Acid, ACS....................................................................................500 mL.............. 152-49
Nitric Acid Solution, 1:1 .......................................................................500 mL............ 2540-49
Potassium Chloride Solution, saturated .................................................100 mL .............. 765-42
Potassium Hydroxide Standard Solution, 8.0 N ........................ 100 mL* MDB........... 282-32H
Sodium Hydroxide Standard Solution, 5.0 N ............................ 100 mL* MDB............ 2450-32
Water, deionized............................................................................................4 L.............. 272-56

OPTIONAL APPARATUS
Description Unit Cat. No.
AccuVac Snapper Kit.................................................................................. each.......... 24052-00
Ampule Breaker Kit.................................................................................... each.......... 21968-00
Cylinder, graduated, mixing, 25 mL........................................................... each.......... 20886-40
Cylinder, graduated, polypropylene, 25 mL ............................................... each............ 1081-40
Cylinder, graduated, 100 mL ...................................................................... each.............. 508-42
Filter Paper, folded, 12.5 cm................................................................. 100/pkg............ 1894-57
Filter Pump ................................................................................................. each............ 2131-00
Flask, volumetric, 100 mL, Class A ........................................................... each.......... 14547-42
Funnel, polypropylene, 65 mm ................................................................... each............ 1083-67
Hot Plate, 4" diameter, 120 V ..................................................................... each.......... 12067-01
Hot Plate, 4" diameter, 240 V ..................................................................... each.......... 12067-02
pH Indicator Paper, 1 to 11 pH ......................................................... 5 rolls/pkg.............. 391-33
pH Meter, sension 1, with electrode .......................................................... each.......... 51700-10
Pipet, TenSette, 0.1 to 1.0 mL..................................................................... each.......... 19700-01

* Contact Hach for larger sizes.

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COPPER, continued
Pipet Tips, for 19700-01 TenSette Pipet .................................................50/pkg ..........21856-96
Pipet Tips, for 19700-01 TenSette Pipet .............................................1000/pkg ..........21856-28
Pipet, volumetric, Class A, 1.00 mL ...........................................................each ..........14515-35
Pipet Filler, safety bulb ..............................................................................each ..........14651-00

For Technical Assistance, Price and Ordering

In the U.S.A.—Call 800-227-4224
Outside the U.S.A.—Contact the Hach office or distributor serving you.

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