Precise Determination of Protein in Meat Products

Protein is one of the most important nutrient components. The correct and accurate determination of protein has a vital role in characterizing the dietary or nutritional value in food products. It is also significant to the economic value of the products. Usually, the amount of protein in meat products is calculated with the nitrogen measured from the sample and a multiplier (6.25 in general). The amount of nitrogen is determined with the help of a combustion method or the classical wet chemical method (Kjeldahl).

The Kjeldahl method can be applied for handling samples with macro sizes of greater than 1 g, used in general for heterogeneous meat product samples. The sample mass is often restricted to ~500 mg or less by physical restrictions in sample encapsulation and circumstances of dealing with the ash build-up inside a vertical furnace combustion nitrogen instrument, rendering it difficult to accurately analyze heterogeneous meat products. The innovative design of the LECO TruMac N combustion nitrogen determinator enables it to handle macro sample sizes, of greater than 1 g, while rapidly performing the analysis with a low cost per analysis.

The LECO TruMac N is a macro combustion protein/nitrogen determinator that performs the macro sample combustion process by using a pure oxygen environment in a ceramic horizontal furnace and large ceramic boats. The moisture in the combustion gas is eliminated by a thermoelectric cooler without the use of chemical reagents. A combustion gas collection and handling system is used for taking up a 3- or 10-cc volume of combustion gas. The combustion gas collection and handling system reduces the quantity of the chemical reagents used for scrubbing and transforming the nitrogen oxide combustion gas into nitrogen, thereby achieving a low cost per analysis. Nitrogen in the combustion gas is detected by using a thermal conductivity (TC) cell.

LECO TruMac N

Sample Preparation

To achieve appropriate results, the consistency of the samples should be uniform. For additional information related to sample preparation, one can refer to AOAC 992.15.

Accessories

528-203 Crucibles

Calibration Samples

502-092 EDTA, 502-642 phenylalanine and 501-050 nicotinic acid

Analysis Parameters*

. .
Furnace Temperature 1100 ºC
TE Cooler Temperature 5 °C
Dehydration Time 0 seconds
Purge Cycles 2 seconds

Instrument Model and Configuration

The operation principle of thermal conductivity detectors is the detection of variations in the thermal conductivity of the analytical gas in comparison with the constant thermal conductivity of the reference gas. The sensitivity of the detector is directly proportional to the difference between the thermal conductivity of the carrier gas and that of the analyte gas. The LECO TruMac N comes in various models that support the use of argon or helium as the carrier gas of the instrument for the thermal conductivity cell.

When helium is used as the carrier gas, it offers the highest sensitivity, ensuring the optimal performance at the lower end of the nitrogen range. An added benefit of helium models is the replacement of the 10-cc aliquot loop with a 3-cc loop inside the gas collection and handling system of the instrument. The instrument is optimized for the best precision and lowest nitrogen range when the 10-cc aliquot loop is used. The 3-cc aliquot loop increases the reagent life expectancy by nearly three times than the 10-cc aliquot loop and also ensures the lowest cost per analysis with very less influence on practical application performance.

The recent shortage in the supply and general availability of helium gas led to the development of the argon model, where argon is used as the carrier gas. Due to the fact that the difference in thermal conductivity between nitrogen and argon is not as high as the difference in thermal conductivity between nitrogen and helium, the detector is intrinsically less sensitive when argon is used as the carrier gas. The argon model (only 10-cc aliquot) has a practical application performance comparable to that of the helium model, when operated with equivalent method and instrument configurations.

Note: For changing aliquot loop size and carrier gas, hardware changes should be made within the instrument.

Element Parameters

Helium
10 cc and 3 cc
Argon
10 cc
Baseline Delay Time 6 seconds 6 seconds
Minimum Analysis Time 35 seconds 55 seconds
Endline Time 2 seconds 2 seconds
Conversion Factor 1.00 1.00
Significant Digits 5 5
TC Baseline Time 10 seconds 10 seconds

Burn Profile

Burn Cycle Lance Flow Purge Flow Time
1 Off On 5 seconds
2 On On 35 seconds
3 On Off END

Ballast Parameters

. .
Equilibrate Time 30 seconds
Not Filled Timeout 300 seconds

Aliquot Loop

. .
Equilibrate Pressure Time 4 seconds
High Precision Yes
High Speed No

*Refer to TruMac Operator’s Instruction Manual for Method for Parameter definitions.

Procedure

  1. The instrument should be set up for operation as described in the operator’s instruction manual.
  2. The system should be conditioned by analyzing three to five blanks (no need for crucible).
  3. Determine blank.
    1. 1.0000 g mass should be entered into Sample Login (F3) with Blank as the sample name.
    2. A 528-203 crucible should be placed in the suitable position of the autoloader.
    3. Steps 3a and 3b should be repeated at least three times.
    4. The analysis sequence (F5) should be initiated.
    5. The blank must be set according to the procedure described in the operator’s instruction manual.
  4. Calibrate.
    1. About 0.75 g of EDTA calibration sample should be weighed into a 528-203 crucible, and mass and sample identification should be entered into Sample Login (F3).
    2. The crucible should be transferred to the suitable position of the autoloader.
    3. Steps 4a and 4b should be repeated at least three times.
    4. The analysis sequence (F5) should be initiated.
    5. The instrument should be calibrated by following the procedure outlined in the operator’s instruction manual. Single standard calibration should be used.

Note: According to the requirements, multi-point (multiple or fractional weight calibration samples) can be used to calibrate. Studies have demonstrated that it is possible to calibrate a properly functioning TruMac using a number of replicates of a single mass range (nominal 0.75 g) of EDTA with a single standard calibration. This is a simple and cost-efficient process. Verification of the calibration can be performed through the analysis of different compounds such as phenylalanine (0.5–0.75 g) and/or nicotinic acid (0.25–0.5 g).

  1. Analyze samples.
    1. About 1 g of the meat product sample should be weighed into a 528-203 crucible; mass and sample identification should be entered into Sample Login (F3).
    2. The crucible should be transferred to the suitable position of the autoloader.
    3. Steps 5a and 5b should be repeated for each sample to be analyzed.
    4. The analysis sequence (F5) should be initiated.

Notes

  • Upon observing soot, or carbon black, in the primary filter, which is a steel wool filter, the sample mass has to be reduced to prevent accumulation of soot in this filter. Soot is formed while analyzing larger masses of high-fat samples.
  • Evident combustible residue and/or low or erratic results in the crucible might hint at incomplete combustion. A more complete combustion of the sample can be achieved through alternative analysis options.
  1. The mass of the sample might be reduced from 1.0 to 0.5 g, and/or
  2. Increase in the analysis time could alter the burn profile. This will still enable the use of 1.0 g samples; however, this will increase the analysis time from ~5.5 to ~7.5 minutes. This is achieved by minimizing the flow rate into the ballast to increase the combustion time.

Burn Profile (Extended)

Burn Cycle Lance Flow Purge Flow Time
1 Off On 5 seconds
2 On On 5 seconds
3 On Off END

Typical Results

3 cc Helium 10 cc Helium 10 cc Argon
Mass(g) % N % Protein Mass(g) % N % Protein Mass(g) % N % Protein
Pre-Cooked Bacon 1.0545 5.58 34.9 1.1018 5.60 35.0 1.0943 5.52 34.5
1.0177 5.47 34.2 1.0768 5.53 34.6 1.0578 5.59 34.9
1.1380 5.50 34.4 1.0537 5.59 34.9 1.0147 5.59 35.0
1.0283 5.57 34.8 1.0068 5.53 34.5 1.0918 5.66 35.4
1.0000 5.48 34.3 1.1606 5.56 34.7 1.0875 5.62 35.1
Avg = 5.52 34.5 Avg = 5.56 34.7 Avg = 5.60 35.0
s = 0.05 0.3 s = 0.03 0.2 s = 0.05 0.3
Roast Beef 1.1097 3.15 19.7 1.0906 3.15 19.7 1.0900 3.17 19.8
1.1058 3.14 19.6 1.0535 3.15 19.7 1.0036 3.13 19.6
1.0644 3.13 19.6 1.1204 3.14 19.6 1.0123 3.12 19.5
1.0504 3.14 19.6 1.0787 3.13 19.5 1.0716 3.12 19.5
1.0725 3.14 19.6 1.1208 3.13 19.6 1.0581 3.14 19.6
Avg = 3.14 19.6 Avg = 3.14 19.6 Avg = 3.14 19.6
s = 0.01 0.04 s = 0.01 0.08 s = 0.02 0.12
Chicken 1.1216 3.10 19.4 1.1232 3.12 19.5 1.0792 3.09 19.3
1.0976 3.10 19.4 1.0916 3.09 19.3 1.0893 3.11 19.4
1.0629 3.06 19.2 1.0094 3.11 19.5 1.0917 3.12 19.5
1.1191 3.08 19.3 1.1038 3.08 19.3 1.0679 3.09 19.3
1.0837 3.07 19.2 1.0031 3.12 19.5 1.0001 3.08 19.3
Avg = 3.08 19.3 Avg = 3.10 19.4 Avg = 3.10 19.4
s = 0.02 0.1 s = 0.02 0.1 s = 0.02 0.1
Smoked Turkey 1.0245 2.83 17.7 1.0843 2.84 17.8 1.0280 2.86 17.9
1.1015 2.80 17.5 1.0463 2.79 17.4 1.0913 2.80 17.5
1.0716 2.80 17.5 1.0778 2.85 17.8 1.0859 2.83 17.7
1.0880 2.78 17.4 1.0739 2.81 17.6 1.0180 2.83 17.7
1.0262 2.80 17.5 1.0649 2.81 17.5 1.0636 2.85 17.8
Avg = 2.80 17.5 Avg = 2.82 17.6 Avg = 2.83 17.7
s = 0.02 0.1 s = 0.02 0.2 s = 0.02 0.1
Ham 1.0595 2.96 18.5 1.1332 2.94 18.4 1.0265 2.89 18.1
1.0273 2.97 18.6 1.0955 2.92 18.2 1.0407 2.96 18.5
1.0567 2.95 18.5 1.0335 2.93 18.3 1.0090 2.91 18.2
1.1205 2.93 18.3 1.0591 2.96 18.5 1.0487 2.89 18.1
1.0636 2.96 18.5 1.0750 2.97 18.6 1.0103 2.94 18.4
Avg = 2.95 18.5 Avg = 2.94 18.4 Avg = 2.92 18.3
s = 0.02 0.1 s = 0.02 0.2 s = 0.03 0.2

This information has been sourced, reviewed and adapted from materials provided by LECO Corporation.

For more information on this source, please visit LECO Corporation.

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