Work plan for Reserva de los González´s proprietary yeast strain propagation

feasibility study

Project Aims and Deliverables

This document describes a demonstration study to evaluate the pilot scale production of
Reserva de los González’s proprietary yeast strain. This will include determining the
feasibility of its propagation at laboratory scale (Chemap 3000; 20L) and conduciveness to
the drying process. The work plan includes the following steps:

1. Propagation, separation, and drying of Reserva de los González’s proprietary yeast

strain at laboratory scale. Samples of the propagated yeast will be provided to the
customer for evaluation. Samples will include 1 L of stabilized liquid yeast (SLY)
and 100 grs of instant dry yeast (IDY).

Methods Overview

Objective 1:

Validate purity of the strain sample provided by the customer.

a. Upon reception, the proprietary strain will be inoculated on YPD agar media.
Five isolated colonies will be selected and processed by RAPD-PCR to insure
product purity.
b. RAPD-PCR will be performed using primers targeting inter-δ sequences within
the yeast genome according to a method adapted for routine quality
measurements at Lallemand. PCR products will be resolved using standard gel
electrophoresis.
c. Purity of the sample will be assessed by comparing the DNA fingerprint of the
five isolated colonies as well as the total yeast biomass.

Objective 2:

Propagate Reserva de los González’s proprietary strain in standard molasses medium

under usual conditions taking into consideration information on growth conditions
provided by the customer.
a. Following standard yeast propagation protocol, the proprietary yeast strain
will be propagated in a series of stages culminating in fed-batch propagation
using molasses medium with nitrogen supplement. Calculations of yield and
productivity, as well as all standard quality tests, will be included.
b. Yeast will be recovered from the propagation medium in cream and cake
forms, with all applicable quality testing.
c. (Optional) The genetic profile of the strain will be determined by PCR at the
end of fermentation to confirm strain stability.

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Objective 3:

Determine the drying feasibility of Reserva de los González’s proprietary

strain under typical process conditions.
a. Following standard yeast propagation protocol, the proprietary yeast strain
will be propagated in a series of stages culminating in fed-batch propagation
using molasses medium with nitrogen supplement. Calculations of yield and
productivity, as well as all standard quality tests, will be included.
b. Yeast will be recovered from the propagation medium and subjected to
standard drying process including press, extrusion and drying to produce
instant dry yeast (IDY).
c. (Optional) The genetic profile of the strain will be determined by PCR at the
end of fermentation to confirm strain stability.

Timing

1. Objective 1 will be completed within 4 weeks of receipt of proprietary yeast

strain.
2. Objective 2 will be completed within 10 weeks of receipt of proprietary
yeast strain.
3. Objective 3 will be completed within 16 weeks of receipt of proprietary yeast
strain.
.
Total cost – CAD $15,000 / $ 12,000 USD

Labor - CAD $5,200 / $4,160 USD

• 1 full time technician (2 weeks)
• 1 partial time Research Scientist (1 week)
• 1 partial time Research Manager (1 day)

Equipment – CAD $7,000 / $5,600 USD

• 20L Bioreactor (2 weeks) 3 stage propagation
• Separator (2 days)
• Fluidized bed dryer 2 days)

Materials and overhead – CAD $2,800 / $2,240 USD

• Propagation ingredients
• Strain preparation and conservation
• Quality control on finished product (1 sample)
• Overhead

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