**SOP 16**

**Standard Operating Procedure for**

**Calibration of Measuring Flasks**

**Volume Transfer Method**

1.  Introduction

    1.  Purpose of Test

> This procedure describes a method for volume transfer calibration of
> the \"to deliver\" volume of measuring flasks from calibrated
> volumetric standards that, in turn, are used as volumetric measurement
> standards, often for packaged commodity verification. If "to contain"
> calibrations are to be performed, appropriate modifications are needed
> in the procedure to ensure complete drying of the flask between
> calibration runs.

2.  Prerequisites

    1.  Verify that valid calibration certificates are available for the
        standards used in the test.

    2.  Verify that the standards to be used have sufficiently small
        standard uncertainties for the intended level of calibration.

    3.  Verify that the standard burets and pipets are clean, free of
        leaks, and in good operational condition.

    4.  Verify the availability of an adequate supply of pure distilled
        or deionized water (See GLP 10).

    5.  Verify that the operator has had specific training and is
        proficient in SOP 16, GMP 3, GMP 7, and is familiar with the
        operating characteristics and conditioning of pipets and burets.

    6.  Verify that the laboratory facilities meet the minimum
        conditions shown in Table 1 to meet the expected uncertainty
        that is achievable with this procedure.

# Table 1. Laboratory environmental conditions

+-----------+------------------------------------+--------------------+
| Procedure | Temperature                        | #                  |
|           |                                    |  Relative Humidity |
+-----------+------------------------------------+--------------------+
| Volume    | 18 C to 27 C,\                     | 40 % to 60 %       |
| Transfer  | maximum change 1 C/h               |                    |
|           |                                    | ± 20 % max change  |
|           |                                    | / 4 h              |
+-----------+------------------------------------+--------------------+

2.  Methodology

    1.  Scope, Precision, Accuracy

> The method is applicable for the calibration of any size of measuring
> flask for which standard pipets of comparable volume are available.
> Typical flasks have volumes in the range of 100 mL to 5 L (1 gill to 1
> gallon). The precision of calibration depends on the care exercised in
> adjusting the various volumes and strict adherence to the various
> steps of the procedure. The accuracy will depend on the accuracy of
> calibration of the volumetric standards used, accuracy in reading the
> meniscus, together with the precision of the comparison. Clean
> glassware and strict adherence to the drainage instructions are
> essential for precise and accurate results.

2.  Summary

The flask to be calibrated is given an initial wet‑down using the
correct drain procedure, then nearly filled with water delivered from a
calibrated pipet. (Calibrated pipets generally deliver slightly less
than the full nominal volume.) Additional water is added from a
calibrated buret until the meniscus in the test item coincides with the
calibration graduation mark. The sum of the volumes delivered into the
wetted flask is equivalent to its \"to deliver\" volume. The process
described assumes that the flask is marked with a 10-second drain time.
If the flask is marked with a drain time other than 10 s, the specified
drain time should be used rather than the 10 s drain time described. A
30-second drain time should be used when a flask has no drain time
specified.

3.  Equipment

    1.  Calibrated standard pipet(s) of suitable volume with recent
        calibration certificates and values traceable to NIST. These
        standards are typically made of borosilicate glass.

    2.  Calibrated standard buret(s) of 10 mL (or 120 minim[^1])
        capacity with recent calibration certificates and values
        traceable to NIST. These standards are typically made of
        borosilicate glass.

    3.  Calibrated thermometer, accurate to 0.1 C with recent
        calibration certificates and values traceable to NIST.

    4.  Supply of pure distilled or deionized water (See GLP 10) at
        thermal equilibrium with the calibration environment.

    5.  Meniscus reading device (See GMP 3).

    6.  Stopwatch or other suitable timing device (does not need to be
        calibrated).

4.  Procedure

    1.  Cleanliness check

> Verify that all glassware, including the standards used and the
> vessels to be calibrated, are internally clean, as evidenced by
> uniform drainage of water. No water droplets should remain on the
> internal surfaces as the water drains from the vessels. If this
> occurs, the glassware must be cleaned with suitable agents and rinsed
> with pure water until uniform drainage is obtained. (See GMP 7).

2.  Wet-down of pipet and flask

    1.  Fill the standard pipet to until water flows from the top tip,
        then completely drain into an empty vessel. Check for uniform
        drainage. Touch off the pipet outlet tip against the con­tainer
        wall to remove excess droplets and to establish a constant tip
        retention volume. The pipet and bore of the delivery side of the
        stopcock should appear to be \"empty\" and the delivery tip
        should contain a small volume of water retained in it. This is
        the \"wet-down\" condition of the pipet. It must be done at the
        beginning of each test sequence. Ensure that the outside of the
        standard pipet is free of water droplets that may fall into the
        unknown flask.

    2.  Refill the pipet and use it to fill the flask to be calibrated.
        Empty the flask in a 30 s 5 s period by gradually inclining the
        flask so as to avoid splashing of the walls as much as possible.
        When the main drainage stream has ceased, the flask will be
        nearly vertical. Allow an additional 10 second drainage after
        discharge of the flask contents while holding the flask at a
        10 - 15 angle from vertical, then touch off the rim of the flask
        to remove any drops adhering to it. At the same time, check that
        uniform drainage has been achieved. This establishes the \"to
        deliver\" condition of the flask. Note that some small amount of
        water will remain in the flask. Each flask to be calibrated must
        be given this \"wet-down\" treatment immediately before the
        start of the test.

2.4.3 Conditioning the buret

1.  Fill the standard buret to overflow and drain several times to
    verify uniform drainage. Refill with water. Note that the stopcock
    bore and delivery tip should be filled with water at all times, in
    contrast to the condition for the pipet.

2.  Touch off the delivery tip against the wall of the receiving vessel,
    to remove any droplet adhering to the external surface of the tip.
    This is an operation that must be done every time a measured volume
    of water is delivered from the buret to ensure consistent volume
    delivery.

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3.  Calibration

    1.  Run 1. Fill the standard pipet to overflow. The delivery bore
        and the delivery tip will be empty, except for the small volume
        retained in the tip. Place the inside of the flask neck in
        contact with the tip of the pipet or buret, to avoid splashing
        but in a manner that does not block the flow of water.

    2.  Deliver the contents of the pipet into the \"wet-down\" flask.
        Repeat as necessary to nearly fill the flask to the calibration
        line. The value recorded for the standard pipet will be the
        total of the deliveries used.

    3.  Add water to the flask from the buret until the meniscus
        coincides with the calibration mark. (See GMP 3 for instructions
        on how to read a meniscus.) Read the volume of water delivered
        from the buret and record on a suitable data sheet such as the
        one in the Appendix. Note: If multiple deliveries of the
        complete buret contents are required, ensure that at no time the
        buret water level drops below the lowest graduation mark, or the
        entire process must be started over. The value recorded for the
        standard buret will be the sum of the delivered volumes.

    4.  Empty the measuring flask as described in 2.4.2.2 to
        re‑establish \"wet-down.\"

    5.  Record all data using the form given in the Appendix or a
        similar format.

4.  Run 2. Replicate Measurement. Repeat the procedure described in
    2.4.4. The test measure must be capable of repeating to 0.01 % of
    the test volume during calibration. Repeatability problems may be
    due to contamination, poor meniscus readings, lack of cleanliness,
    or when calibration is conducted in an unstable environment.
    Repeatability problems must be corrected before calibration can be
    completed.

5.  Calibration Note: If the vessel has a graduated neck (in addition to
    the nominal mark) additional graduations should be inspected for
    proper marking and calibrated as well.

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2.  Computations

    1.  Compute the individual \"to deliver\" volumes, *V~TD~*, for Run
        1 and Run 2.

$V_{\text{TD}} = V_{P} + V_{B}$ Eqn. 1

## 

## 

## Table 2. Variables for volume equation

  ---------- ------------------------------------------------------------
  Variable   Description

  *V~TD~*    "To deliver" Volume

  *V~P~*     \"to deliver\" calibrated volume of the standard pipet

  *V~B~*     volume of water delivered from the buret, corrected for any
             calibration values
  ---------- ------------------------------------------------------------

2.  Temperature correction

> For calibration of glassware that is not made of borosilicate glass,
> not only should the above temperature conditions be realized but the
> water temperature must also be known. The water used must be stored in
> the laboratory until its tempera­ture is equilibrated with its
> surroundings and the temperature of the discharged water is measured,
> using the calibrated thermometer. The \"to deliver\" volume of the
> flask, corrected to the reference temperature of 20 C, *V~TD~*~20~, is
> computed, using the expression:

$V_{\text{TD}\text{20}}\text{  } = \text{  }V_{\text{TD}}\left\lbrack 1\text{  } + \text{  }\left( \alpha\text{  } - \text{  }\beta \right)\left( t\text{  } - \text{  }\text{20} \right) \right\rbrack$
Eqn. 2

### Table 3. Variables for volume temperature correction equation

  ------------- ------------------------------------------------------------
  Variable      Description

  *V~TD~*       observed delivered volume as computed in 3.1

  *V~TD~*~20~   "To deliver" volume at the reference temperature of 20 C

  *t*           temperature of water at time of calibration

                cubical coefficient of expansion of glass of standard (e.g.,
                0.000010 °C^-1^ for borosilicate glass)

                cubical coefficient of expansion of glass of flask (e.g.,
                0.000025 °C^-1^ for soda-lime glass)
  ------------- ------------------------------------------------------------

> The volumetric standards are typically made of borosilicate glass. If
> borosilicate flasks are calibrated, no temperature correction is
> required. This procedure relies on the fact that since deliveries are
> made over a short period of time, the temperature remains the same in
> the standard as in the unknown. *When standards and flasks are not of
> the same material, the water temperature change must be less than 0.5*
> C *while the water is in the standards and the flask.*

3.  Report the average volume *V~TD~*~20~ as the value for the flask
    after the temperature correction has been applied to each Run.

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3.  Measurement Assurance

    1.  Duplicate this process with a suitable check standard or have a
        suitable range of check standards for the laboratory. See SOP 17
        and SOP 30.

    2.  Plot the check standard volume and verify that it is within
        established limits. Alternatively a *t*-test may be incorporated
        into the process to check the observed value against an accepted
        reference value. (See NISTIR 6969, Section 8).

    3.  The mean of the check standard value is used to evaluate bias
        and drift over time.

    4.  Check standard observations are used to calculate the standard
        deviation of the measurement process.

4.  Assignment of Uncertainties

The limits of expanded uncertainty, *U*, include estimates of the
standard uncertainty of the laboratory volumetric standards used,
*u~s~*, plus the standard deviation of the process, *s~p~*, at the 95
percent level of confidence. See SOP 29 for the complete standard
operating procedure for calculating the uncertainty.

1.  The standard uncertainty for the standard, *u~s~*, is obtained from
    the calibration report. The combined standard uncertainty, *u~c~*,
    from the calibration report must be used and not the expanded
    uncertainty, *U*. Therefore, the reported uncertainty for the
    standard will usually need to be divided by a coverage factor, *k*.
    Multiple standards are used in this procedure; however, they are
    typically calibrated independently. Multiple standard uncertainties
    are handled according to SOP 29.

2.  The standard deviation of the measurement process is obtained from
    control chart performance (See SOP 17 or 20, and SOP 30).

    1.  The value for *s~p~* is obtained from the control chart data of
        the check standard using the same volume transfer procedures on
        flasks of comparable size and neck dimensions.

3.  Other standard uncertainties usually included at this calibration
    level include uncertainties associated with the ability to read the
    meniscus, only part of which is included in the process variability,
    the cubical coefficient of expansion for the flask under test, use
    of proper temperature corrections, the accuracy of temperature
    measurements, round robin data showing reproducibility,
    environmental variations over time, and bias or drift of the
    standard.

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5.  Report

    1.  Report results as described in SOP 1, Preparation of
        Calibration/Test Results, with the addition of the following
        items:

        1.  Volume, uncertainty, reference temperature, material,
            thermal coefficient of expansion (assumed or measured),
            construction, any identifying markings, tolerances (if
            appropriate), laboratory temperature, water temperature at
            time of test, barometric pressure, relative humidity, and
            any out-of-tolerance conditions.

6.  Alternative Reference Temperatures

    1.  Reference temperatures other than 20 ºC (68 ºF) may occasionally
        be used. Common reference temperatures for other liquids follow:

  ------------------------------------------------- ---------------------
  Commodity                                         Reference Temperature

  Frozen food labeled by volume (e.g., fruit juice) \- 18 ºC (0 ºF)

  Beer                                              3.9 ºC (39.1 ºF)

  Food that must be kept refrigerated (e.g., milk)  4.4 ºC (40 ºF)

  Distilled spirits or petroleum                    15.56 ºC (60 ºF)

  Petroleum (International Reference)               15 ºC (59 ºF)

  Wine                                              20 ºC (68 ºF)

  Unrefrigerated liquids (e.g., sold unchilled,     20 ºC (68 ºF)
  like soft drinks)                                 

  Petroleum (Hawaii)                                26.67 ºC (80 ºF)
  ------------------------------------------------- ---------------------

2.  When alternative reference temperatures are used, at the request of
    the user of the field standard, and when the glassware is *not*
    marked with a reference temperature, the calibrated value of the
    standard may be reported at a different reference temperature,
    *t~ref~*, by using this equation:

$V_{\text{TD}t\text{ref}}\text{  } = \text{  }V_{\text{TD}}\left\lbrack 1\text{  } + \text{  }\left( \alpha\text{  } - \text{  }\beta \right)\left( t\text{  } - \text{  t}_{\text{ref}} \right) \right\rbrack$

Eqn. 3

**Additional References:**

Bean, V. E., Espina, P. I., Wright, J. D., Houser, J. F., Sheckels, S.
D., and Johnson, A. N., NIST Calibration Services for Liquid Volume,
NIST Special Publication 250-72, National Institute of Standards and
Technology, Gaithersburg, MD, (2006).

**Appendix**

**Calibration of Measuring Flask**

**(Volume Transfer Method)**

**Laboratory data and conditions**:

  ----------------- ----------------- ----------------- -----------------
  Date                                Temperature       

  Operator                            Pressure          

  Test Number                         Relative Humidity 

  Vessel Calibrated                   Vessel ID         

  Process standard                                      
  deviation from                                        
  control chart,                                        
  *s~p~*                                                
  ----------------- ----------------- ----------------- -----------------

**Volume standard(s) data:**

  --------- --------- --------- -------- -------- --------- ------------ --------
  ID        Nominal   Calib.    Coeff.   ID       Nominal   Volume       Coeff.
                      Volume    of Exp                      Correction   of Exp

  Pipet                                  Buret                           

  Unknown                                                                
  Flask                                                                  

  *u~s~*,                                                                
  and *k*                                                                
  factor                                                                 
  --------- --------- --------- -------- -------- --------- ------------ --------

**Observations:**

  ----------------------------------------------------------------------------------------------------------------------- ----------------------- ----------------------
                                                                                                                          TRIAL NO. 1             TRIAL NO. 2

  Calibrated Pipet Volume: *V~p~*                                                                                                                 

  Buret Volume:                                                                                                                                   

  Final Reading                                                                                                                                   

  Initial Reading                                                                                                                                 

  Difference                                                                                                                                      

  Calibration Correction (if required)                                                                                                            

  Corrected Buret Volume, *V~B~*                                                                                                                  

  *V~TD~* = *V~P~* + *V~B~*                                                                                                                       

  Water Temperature                                                                                                                               

  $$V_{\text{TD}\text{20}} = V_{\text{TD}}\left\lbrack 1 + (\alpha - \beta)\left( t - \text{20} \right) \right\rbrack$$                           

  Average *V~TD~*~20~                                                                                                                             
  ----------------------------------------------------------------------------------------------------------------------- ----------------------- ----------------------

[^1]: 1 \[US\] minim = 0.061615496 milliliter (mL). 120 minim standards
    were provided to the States as reference standards for use in volume
    transfer calibrations. These standards have higher resolution than
    metric standards provided at the same time.