1. QA at Two Roads: 0 to 60 in 2 years flat Presentation to MBAA Summer Technical Meeting Middlebury, VT August 8, 2015 Kendra Frederick, Ph.D. Director of Quality Assurance, Two Roads Brewing Co - Stratford, CT

2. An Outline 1.Intro 1.Two Roads’ story, challenges, strengths 2.Growth of QA 1.Focus 2.Equipment 3.Personnel 3.Current Focus / Testing 4.Future

3. Setting the Stage Two Roads Brewing (TRB) -Dual Business model: contract & own brand -Contract customers (13) -Large number of recipes (89 in 2014) -Numerous Yeast Strains (12 total; 7 in regular use) -Rapid growth -Volume -# of recipes -Personnel -Location -converted old factory next to train tracks; warehousing outdoors -Lab space OK at first…quickly became small

4. Growth since inception YEAR BBLS TRB brand Total % growth – total 2012¹40100 20136,64324,196 201416,886 69,849154% 201525,000*100,000*34% *projected BBLS ¹ Launched in Dec

5. Challenges Can we slow down for a sec? Rapid Growth – volume -Increasing testing throughput -heavy reliance on technology/automation -Prioritizing testing -Everything tested for micro -Some tests only every X batches, # packages -TRB brand-only tests: IBU, Color, Sensory -Recipes (large #) -Takes a longer time to assess consistency, detect brand trends

6. Challenges What’s your name again? Rapid Growth – Personnel -Training -Split & 3 rd shift -Method creep -SOPs! (words vs pictures) -Learning to Manage -Adjusting to supervising instead of doing -Learning to Trust someone else doing -I’m hands off…I hope you are too! -Agile Leadership

7. Challenges Would you like fries with that? Customers -Matching brand flavor across breweries -Communication -One customer changed their yeast & forgot to tell us! -I’m 2 months behind on reports -Numbers: just because it’s on the label, doesn’t mean it’s true -We’d hit target OG & FG…but ABV would be out of compliance with label -Special requests ->20 beers sent out for external testing for a single client

8. Strengths -Funding -Start-up & subsequent phases have been well-funded -Debt OK if permits growth, improves quality, reduces risk -Experience -Brewmaster 25 years, CEO & CMO >15 years each -Past negative experience w/ contract brewing = desire to do it right -Support from Owners -Quality is important to Owners -Regional/National craft breweries serve as models

9. My background / philosophy -Education -Degree in Biophysical Chemistry -Lots of pipetting! -Research, reading literature -Show me the Data -Biopharmaceuticals -Product = Process -Documentation, SOPs -Cleanliness -Method development / assay validation QA = Quality Assurance -Preventative measures to ensure quality -SOPs, Process Monitoring, Raw material testing, Quality CCPs QC = Quality Control -Measurements taken after the fact to check for quality / prevent the release of inferior product -Finished goods testing

10. HOW QA HAS GROWN at TRB

11. QA Focus PROGRESSION 1.Basic Data (gravity, pH), Micro, DO 2.Compliance & package integrity (fill volumes, seam checks) 3.Process Consistency (wort aeration, cell counts, sensory) 4.Add capabilities (IBU, Color, Haze, ABV, sensory training) 5.Reduce risk / time to test result (PCR, TPA) 6.Shelf Life (TPA) ALWAYS AT THE FOREFRONT A.Throughput (digital meters, automation, filtration manifold) B.Precision (digital, automation) C.Documentation

12. Equipment - growth Approx Date Approx Annualized BBLS InstrumentsUse Start-up Dec 2012: Getting off the ground n/a Microscope pH meter Hydrometers Scale Laminar Flow Hood Micropipettors ATP swabs / luminometer Basic measurements, micro Automated Cell counter (Nucleocounter) Yeast counts & viability Dissolved Oxygen Meter + package piercer DO, TPO

13. Equipment - growth Approx Date Approx Annualized BBLS InstrumentsUse early 2013: Increasing throughput 15,000 Handheld density meter (Anton Paar DMA 35) Increase throughput / precision of gravity measurements Filtration Manifold* Increase throughput of micro testing Headspace airs tester (Zahm-Nagel) TPO

14. Equipment - growth Approx Date Approx Annualized BBLS InstrumentsUse late 2013 – early 2014: Increase capabilities, consistency 24,000 Seam Check equipment* Package integrity High-range DO meter Wort aeration UV-VisIBU, Color, et al *at recommendation of Packaging Manager

15. Equipment - growth Approx Date Approx Annualized BBLS InstrumentsUse Mid 2014: Increase capabilities, throughput, reduce risk 50,000 PCR equipment* Rapid detection of micro contamination 2 nd handheld DMA Increase throughput of gravity measurements AlcolyzerABV, density *decision driven by need to supply timely results to contract customers; got a good deal on instrument

16. Equipment - growth Approx Date Approx Annualized BBLS InstrumentsUse Late 2014: Increase capabilities, precision / consistency 70,000 Turbidity meter*Haze Cell counter (Cellometer) Improve precision / accuracy of cell counts. Add capabilities (vitality) Digital carbonation tester (CboxQC) CO2, DO Mid 2015 Increase capabilities, thruput 90,000 Total Package analyzer (Hach Orbisphere)* TPO *purchase driven by issue with particular contract brand & desire to match measurements across 2 sites

17. Lab Growth: Personnel Date Approx Annual BBLS # Empl. BBLS / Employee before -- after new hire Notes 11/120 1 0 06/1317,280 Intern ~10 hrs/wk starting 03/13 2 8,640 09/1450,400 25,200Intern ~10 hrs/wk starting 09/13 3 16,800 12/1456,348 18,763 3.5 16,1000.5 FTE dedicated to sensory 07/1590,933 25,981 4.7519,144A real QA Department!

18. CURRENT STATE OF QA at TRB

19. Current brewery set-up -Brewhouse: -100-bbl, 4 pieces -Fermentor Tanks: 34, >200k capacity -12 x 100-bbl -18 x 300-bbl -4 x 600-bbl (new as of 06/12) -Yeast Brinks: 4 x 15-bbl -Centrifuge: HP2 GEA (150 bbl/hr) -Brite Tanks: -1 x 50-bbl -2 x 100-bbl -1 x 200-bbl -3 x 300-bbl -Bottle Filler -40-head, 300 bpm (750 cases/hr) -Formats: Heritage, long-neck, 22- oz, 375-mL, 750-mL -Crown, cork & cage -Can filler -32-head, 280 bpm (700 cases/hr) -12-oz & 16-oz -High-cone; 4-, 6-, 12-pack wraps -Keg line -1 keg/min (60 kegs / hr) -4-station Shifts Brew: 24/5 Package: 12/5

20. Current Quality Focus minor major x Kegging x Centrifuge

21. Micro testing FermentationBright Tank Packaged Beer Yeast Timing: 24 -72 hours post-KO once per week 2 from each run; >12-24 hrs post-fill When harvested to brink Media LMDA+ / aeroXXEvery 3 rd New prop WLD / anaeroXXEvery 3 rd New prop NBBXXXNew prop LCSM¹X 1/wkNew prop LWYM¹,²X 1/wkNew prop WLNX RDM³X In case of positive: Retest at 5-7 days PCR on package Retest if BS; PCR if NBB is pos WLN: test beer; dump yeast ¹Saisons not tested ²Hefs not tested ³YPD + TTC overlay BS = beer spoiler

22. Micro “Clean room” was an old bathroom…so hood was a must!

23. Yeast Testing Strain cross-contamination: -Currently: Plate on WLN 4-5 days. Look for colony homogeneity -Coming soon: PCR assay?! Ale Lager French Saison

24. Micro: PCR -Real-time PCR thermocycler (AB StepOne Plus) -Kits available to detect Lacto/Pedio and Dekkera -Sensitive enough to detect after 1-2 days of pre-incubation -“Open source”: can develop own assay -Currently collaborating to develop assay to detect yeast cross-contamination *PCR = Polymerase Chain Reaction; detect DNA from target organism / gene

25. Cell Counts Previously: Nucleocounter -Cassettes – expensive. Images: low resolution. No opportunity to manually adjust counts Now: Nexcelom Cellometer -Higher magnification, brightfield image, manual adjustment possible, several “staining” options including vitality. -Required fair degree of method development & user training (pipetting!)

26. Finished Beer Testing Package Testing TestFrequencyTestFrequency GravityEvery batchTPOEvery run pHEvery batch Fill Volume Verification Every TRB; contract: > every 4000 cases ABVEvery batchCarbonationOccasionally Haze Nearly every batch Seam Check At least every 4000 cases ColorTRB only IBUTRB only

27. UV-Vis Spectrophotometer Turbidimeter Phys-Chem Orbital shaker for IBU sample prep

28. Seam Check Equipment Date  Adjustment

29. Alcolyzer -Have had issues keeping in “calibration” from the get-go. -We use to perform daily gravity checks. Requires more vigorous cleaning, but worth it. -Peristaltic pump broke. Service tech of 17 years had never seen one break.

30. Dissolved Gasses o-DGM -DO into/out of centrifuge Previously: -DO in BT’s -DO in packages CboxQC -Carb & DO in BT’s -Required significant training, despite seeming simple to use!

31. Our Newest Toy: TPO analyzer -Broke within the first week!

32. Other Testing -ATP swab (after every CIP) -Water testing (weekly) -Swabs on fillers -Sieve testing of grist (as “needed”)

33. Data Management -Lots of work / attention / time devoted to documentation -measurements, volumes, dates/times, etc -Series of Excel spreadsheets “linked” via Macros (saving, copying / pasting automated) -Leveraging Dropbox to share data/files with customers -Will investigate more elegant data management solution in 2016

34. Data Management

35. Primary DataPrimary SourceData Transfer 1-->2.1 Secondary Destination 1 Density, ABV, FG Alcolyzer/DMA output copy raw data to 'Alcolyzer - data manipulations' file. Then use macros to export to 2 dest QA Batch Data Package Date Batch Listuse macro in 2 dest to pull from 1° CRS Batch Sheet - Testing tab Cell Counts Control Room Sheetuse macro in 2 dest to pull from 1°Yeast Tracking Sheet OG, pH Control Room Sheetuse macro in 2 dest to pull from 1°QA Batch Data Brew Date, Sep Date Control Room Sheet ues 'Log Book - reports' macro to generate report; copy manually into 2 / auto copy to 2 Batch List Dry hop date Control Room Sheetformulas in-file CRS Batch Sheet - Testing tab DO - BT Control Room Sheet**need procedureDO data log Carbonation Control Room Sheetformulas in-file / manual CRS Batch Sheet - Testing tab TPO - PKG DO loguse macro in 2 dest to pull from 1° CRS Batch Sheet - Testing tab Fill Volumes Fill Volume data files**need procedure CRS Batch Sheet - Testing tab IBU, Color IBU/Color data filesmacro in 1 sends data to 2°QA Batch Data Final pH Weekly Data SheetmanualQA Batch Data Micro Micro monthly log filesmanual CRS Batch Sheet - Testing tab Haze Weekly Data SheetmanualQA Batch Data

36. Sensory A work in progress… -Began formal sessions in March 2013 -Differing visions for how to approach, begin, conduct sensory -Me: start with training. Others: dive right into descriptive analysis -Different groups with different goals/needs -Brewers/Cellar: Off-flavors in wort, green beer, and finished beer -Sensory Panel: trueness to brand, off-flavors in finished beer -Sales – Internal: brand recognition, off-flavors in the “field” (oxidation, dirty lines) -Sales – External: as above, plus “Cicerone” (general beer IQ)

37. Sensory Struggles… -Schedules: “9-5’ers”, 2 nd & 3 rd shift, sales -Attendance -11am ideal palate-wise, but we’ve found mid to late afternoon better schedule-wise -Lots of management “too busy” for sensory -Space -(currently) No dedicated sensory space. Use event room, second bar, meeting room.

38. Sensory A compromise… -2 sessions a week, focusing on consistency & stability of TRB brands. Occasional trainings & external comparisons -Brand Attribute forms – to guide tasters. -Brand-specific list of desirable (or not) attributes + target strength.

39. Sensory Righting the course… -Hired a dedicated employee for sensory! (0.5 FTE) -More availability to accommodate different groups’ schedules -Training -Flavor Recognition: Using Sigma & Aroxa spikes -Sales: 3 sessions so far, seeing progress (!), giving them ideas -Brewers & General Panel getting trained as well. -Methods: -Introducing & using triangle -Space -Coming soon!

40. THE FUTURE

41. Expansion! +

42. QA Focus 1.Basic Data (gravity, pH), Micro, DO 2.Compliance & package integrity (fill volumes, seam checks) 3.Process Consistency (wort aeration, cell counts, sensory) 4.Add capabilities (IBU, Color, Haze, ABV, sensory training) 5.Reduce risk / time to test result (PCR, TPA) 6.Sensory (trained & validated panel!) 7.Kegs 8.Shelf Life (TBARS, TPO, chill haze) 9.Consistency (wort & green beer IBU & color, GC) 10.Raw Materials