Innovation and creativity 18 April 2018

Anne Grobler and teamsDST/NWU Preclinical Drug Development PlatformPheroid Cluster Incubator

The world must produce as much food in the next 40 – 80 years as it has in the past 12,000 years combined, and on less land.

1 out of 6 people suffers from hunger globally.Need a minimum of 0.5 ha cropland per capita.1961 = 0.41 ha of cropland per capita.2002 = 0.23 ha of cropland /capita.

±30% of the world's cropland nowunproductive1.5 billion hectares (nearly ⅓) of the world's

cropland abandoned over the past 40 years

The problem of food security requires real innovation

2

Is nanotechnology the solution?

Nanoscale“Nano” – Greek work for dwarf1 m = 1 billion nanometers (nm)Various types of nano

Nanotechnology refers to the use of structures between 0,1 and 100nm

Management of pathogensPhysical tracking of crops & productsUse of materials Environmental uses

Ulbrich, Holá, Šubr†, Bakandritsos, Tuček, Zbořil. Chem. Rev., 2016, 116 (9), pp 5338–5431

"The Scale of Things - Nanometers and More”. 2012. https://nanohub.org/resources/13842.

Nano-particles

Nano-composites

Nano-emulsions

Nano-structured materials

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Delivery

Formulation

Packaging

Heat/mass transferReaction

engineering

Bio-processes

Molecular synthesis

Nano-sensorsNano-tracers

Materials

Processing

Product

Biosecurity

Food & nutritional

security

The relevancy of nanotechnology

Nano in food & agricultural

science

http://www.nano-and- society.org/commentaries/commentary_willow02.html www.euronanotrade.com/news/Sources_of_Information.dochttp://www.azonano.com/Industries.asp http://www.nanotechforest.org/dwnlds/Nano%20Workshop_Nano-Art%5B1%5D.ppt#357,13http://nanoandcommodities.wordpress.com/tag/agricultural-and- food-commodities/agricultural-inputs-for-nanotechnology/ http://www.azonano.com/SearchResults.aspIndKeyWord=Agriculture+forestry+and+fishing http://www.sciencedirect.com/science? _ob=ArticleURL&_udi=B6VHY-4KFV3B4 http://www.nano-and-society.org/commentaries/commentary_willow02.html www.euronanotrade.com/news/Sources_of_Information.dochttp://www.azonano.com/Industries.asp http://www.nanotechforest.org/dwnlds/Nano%20Workshop_Nano-Art%5B1%5D.ppt#357,13, http://nanoandcommodities.wordpress.com/tag/agricultural-and- food-commodities/agricultural-inputs-for-nanotechnology/ http://www.azonano.com/SearchResults.aspIndKeyWord=Agriculture+forestry+and+fishing http://www.sciencedirect.com/science? _ob=ArticleURL&_udi=B6VHY-4KFV3B4

A plethora of nanoagric websites

A plethora of nanoagric products

• Reduces phytotoxicity• Increases productivity• Stimulates plant growth• Improves stability of AIs• Protection from leaching• Increases root formation• Pest control at lower doses• Increases the product weight• Improves bioavailability of AIs• Delivery of water insoluble AIs• Improves the quality of the soil• Reduces cyto- and genotoxicity

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A plethora of nanoproduct claims

Conceptual claims

• Improved leaf adhesion• Protection from photolysis &

volatilization• Uniform spreading• Controlled release• Improved translocation• Provide smart monitoring

And more claims

Contextual claims

What has been proven & how &

by whom?• In the field?• Single or as tank mix?• Stability on the shelf, in

the tank?• Is it safe?• Is it green?• Is it financially viable?• What should be

regulated?

Innovation = the implementation of creativity. It requires certain preconditions in order to occur fluidly.

Wishful thinking

Innovation is cyclical: the more it happens, the more it happens again – it inspires.

Diplomatic Courier and World Affairs Council in Washington, DC.

Innovation ≠ just about a creative idea.

12

Liquid fertilizer

Macro-elements

Micro-elements

Deficiencies

Specialized studies

Pellet Fertilizer

Coatgro

Field studies

Stability trials

Anngro Investments

Insecticide

Fungicide

Herbicide

Formulation & administration

Pheroid transporter

development

Combination Pheroid & other nano-materials

Improved efficacy & safety

Manufacturing

An integrated developmental approach

Compound (AI) does not reach its target.

New biological molecules are unstable.

Minimize phytotoxicity and chemical use.

Address compound resistance.

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Pheroid

Flexible & safeDiverse entrapment

Inexpensive

Diverse administration forms

Pheroid as base for bio-nanotransporter

Form

ulat

ion

Pilo

t Pla

nt

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PCDDP Facilities

The PCDDP is a service to the scientific community in SA, not owned by an institution or an individual.Established in 2012, it is aligned with the Bioeconomy Strategy (DST), the NDP 2030, the health priorities of DoH & the industrial development plan (IPAP) of SA.

Laser-based analysis

15

PRODUCT QUALITY CONTROL

Risk/ benefit ratio: formulations

16

Risk of systemic absorption

Absence of “nano effects”

100 – 400 nm

Malvern particle size analysis

Safe & green nano-transporter & crop enhancer

Based on Pheroid drug delivery system & customized to plant use;

NWU retains IP; Biopher develops & manufactures & distributes in SA;

International companies distributes in the rest of the world.

Bio-nano-transporter

Plant growth

regulators

Insecticides

Antifungals

Macro-elements

Micro-elements

Herbicides

dticluster

The Pheroid constituents & properties

1316 5730

500

1000

1500

2000

2500

Dea

th C

ompa

rison

Tim

e (S

econ

ds)

Treatments

The efficacy of insecticide to control blowfly without and with ANNGRO™

FASTAC FASTAC + Anngro0

2

4

6

8

10

12

Control Mancozep+Flint+Oil Mancozep+Flint+Anngro

Dis

ease

inde

x

Treatments

Disease incidence of citrus blackspot (Guignardia citricarpa)

b

Fertilizers & pesticidesA. Improved uptake by

plantsB. Enhanced transportC. Cellular uptakeD. Metabolism

Results from trials

Photos of sunflower plants treated with the 4:2:1 (18) + 3% S clear liquid fertilizer (Left) and the 4:2:1 (18) + 3% S clear liquid fertilizer + AnnGro™ combination treatment (Right) on 23/3/2011.

Improved uptake!!!

PILOT TRIAL

Zinc uptake in macadamias Statistical – Nelspruit SA

0

10

20

30

40

50

60

70

Control ZnO ZnO+Anngro

Zn (m

g/kg

)

Treatments

Zinc content in the leaf of Macadamias 4h after application

+317%

+555%+238%

9000

11000

13000

15000

17000

Hartswater Orania Prieska

Yiel

d (k

g/ha

)Impact of N-carriers on maize yield under irrigation - 2010

UAN Opti-N

8.3%

7.2%

11.1%

N: 40 kg N/ha; applied 3 weeks after emergenceHartswater: CV = 14.7%; Orania: CV = 7.6%; Prieska: CV = 4.5%With complements of Sidi Parani Fertilizer South Africa

Opti-N (Nitrogen + AnnGro™)

Test Site：Lianshui JianSu

Rice: direct seeding, 3-4 leaf stage

1、Penoxsulam(Dow ArgoSciences®)27g+20% Cyhalofop-Butyl 25ml+50% Quinclorac 40g +Fenoxaprop10ml(BAYER®)+3ml AnnGroTM/15L water

2、Penoxsulam(Dow ArgoSciences®)27g+20% Cyhalofop-Butyl 25ml+50% Quinclorac 40g + Fenoxaprop10ml(BAYER®) +3ml AnnGroTM + 1g VitaCat/15Lwater

LSD oh(T≥0.05) = 0.196 LSD 4h(T≥0.05) = 0.145

0,4

0,42

0,44

0,46

0,48

0,5

0,52

0,54

Control Ca(NO3)2 Ca(NO3)2+Anngro

Cal

cium

con

tent

in %

DW

bas

is

Treatments

B. TRANSPORT: Calcium content in potato leaves at tuber initiation - Bloemhof 08/09, RSA

Ca 0h LCa 4h L

Before application

LSD 0h (T≥0.05) = 0.1632 / LSD 0h (T≥0.05) = 0.1364

Treatments – Ca(NO3)2 =1%, AnnGro™ = 120ml/haCultivar – FianaPopulation – 44 000 seeds/haFertilizer – Before plant: 2 ton diplomatic lime, 1 ton Super P/ha, At plant: 500kg 5:1:1 (30) + 500kg Super PWater volume – 500L/haTime of application – 20 mm tuber sizeLeaf analysis – youngest matured leaves o hours and 4 hoursTuber analysis – at harvest

0,001

0,00105

0,0011

0,00115

0,0012

0,00125

0,0013

0,00135

Control Ca(NO3)2 Ca(NO3)2+Anngro

Cal

cium

con

tent

in %

DW

bas

is

Treatment

Calcium content of potato tubers at harvest- Bloemhof 08/09, RSA

Control

Ca(NO3)2

Ca(NO3)2+Anngro

+16%

LSD 0h (T≥0.05) = 0.0003

> Ca into tuber

LSD(T≥0.05) = 0.0004

TRIAL INFORMATION

• Fertilizer applied @ 1% solution and AnngroTM @ 20ml/100L• A complete randomized block design was used and each treatment was replicated

three times. • Spraying commenced at the GS.5 – GS.7 growth stage as illustrated by the Joubert

wheat classification system. • Distilled water was used with the spray volume set at 300L/ha• Twenty four hours after treatment application, plants were taken out with a spade to a

depth of 30 cm at three spots randomly selected in each plot.• Statistical analysis: NCSS 2000 statistical program at a 95% significance level.

Differences between treatments were calculated using the Tukey-Kramer multi-comparison test.

RH(%) 25.3

T (°C) 20.1°C

Delta T (°C)

10.5

Cu (mg/kg) 0.47Mn (mg/kg) 4.8Zn (mg/kg) 9.3

20304050607080

Control CuSO4 CuSO4+AnnGro

Cu

mg/

kg D

W b

asis

Treatments

Amount of Cu in the wheat leaf after 24 hours - Bloemfontein -2009

20304050607080

Control CuSO4 CuSO4+AnnGroCu

mg/

kg D

W b

asis

Treatments

Amount of Cu in the wheat root after 24hours - Bloemfontein - 2009

+84%

LSD(T≥0.05) = 11.89

LSD(T≥0.05) = 14.62

CuSO4 & AnngroTM

• Fertilizer applied @ 1% solution and AnngroTM @ 20ml/100L• A complete randomized block design was used and each treatment was replicated

three times. • Spraying commenced at the GS.5 – GS.7 growth stage as illustrated by the Joubert

wheat classification system. • Distilled water was used with the spray volume set at 300L/ha• Twenty four hours after treatment application, plants were taken out with a spade to a

depth of 30 cm at three spots randomly selected in each plot.• Statistical analysis: NCSS 2000 statistical program at a 95% significance level.

Differences between treatments were calculated using the Tukey-Kramer multi-comparison test.

RH(%) 25.3

T (°C) 20.1°C

Delta T (°C) 10.5

Cu (mg/kg) 0.47Mn (mg/kg) 4.8Zn (mg/kg) 9.3

50

100

150

Control MnSO4 MnSO4+AnnGro

Mn

(mg/

kg) D

W b

asis

Treatments

Amount of Mn in the wheat leaf 24 hours after application - Bloemfontein - 2009

+40%

50

100

150

Control MnSO4 MnSO4+AnnGro

Mn

(mg/

kg) D

W b

asis

Treatments

Amount of Mn in the wheat root 24 hours after application - Bloemfontein - 2009

LSD(T≥0.05) = 24.94

LSD(T≥0.05) = 62.23

MnSO4 & AnngroTM

C. CELLULAR UPTAKE: AnnGro™ and lipid rafts

Pheroid®

Cell membrane ruffles

* Micrograph from an optical section* Krypton argon laser & helium neon laser

Iswanto, A.B.B.; Kim, J.-Y. Lipid Raft, Regulator of Plasmodesmal CalloseHomeostasis. Plants 2017, 6, 15.

Grobler, A. 2009. Applications of Pheroid technology.

D. METABOLISM: yield effect of foliar applied fertilizer on rain fed maize

2010 – Bultfontein – South Africa

Control Zn Chel ZnSO4 ZnSO4+A ZnSO4+A2Series1 0 479 370 504 839

0

100

200

300

400

500

600

700

800

900

Diff

per

ha

in k

g

Yield of dry land maize - 2009/2010 - Bultfontein - P Ferreira

9000

11000

13000

15000

17000

Hartswater Orania Prieska

Yiel

d (k

g/ha

)Impact of N-carriers on yield of maize under irrigation - 2010

8:1:3(20) Opti-Top

1.9%8.7%

7.4%

N: 40 kg N/ha; applied 3 weeks after emergenceHartswater: CV = 14.7%; Orania: CV = 7.6%; Prieska: CV = 4.5%With complements of Sidi Parani Fertilizer South Africa

Independent trials: Opti – TOP (NPK + AnnGro™)

Maize trials Sidi Parani 2010 season

12711

17921

16229

1520014844

18605 18438

16990

16089

19474

20495

18210

10000

12000

14000

16000

18000

20000

22000

Vaalharts Hopetown Prieska Orania

Yiel

d (k

g/ha

)

Areas

ControlUANOPTI-N

5

7

9

11

13

15

17

19

21

UAN Opti-N

kg g

rain

/kg

N

Treatments

Grain produced/kg N applied - Sidi ParaniFertilizer in the Northern Cape, SA

+36%

Economical advantage for 2013 maize season

3000

3200

3400

3600

3800

4000

4200

Control ComCat ComCat +AnnGroTreatments

Yield of rainfed maize - Bultfontein 2009+513 kg/ha+448 kg/ha

700007200074000760007800080000820008400086000

Control ComCat ComCat + AnnGro

Yiel

d (k

g/ha

)

Treatments

Yield of potatoes - Petrusburg 2008

+5.9ton/ha

NWU home –grown IP

IP portfolio:(i) 6 patents; 1 of which is

Necsa/NWU.(ii) Trade marks (Pheroid, etc).(iii) Designs (2 in process).(iv) Know-how, including

proprietary models & methods.

(v) Trade secrets (NDA-based).(vi) Copyright (publications, book

chapters, websites, etc).(vii)Products packages. (viii)Markets & marketing

material.

CONFIDENTIALAnnGro USA LLC

BACKGROUND

AnnGro™ is based upon the Pheroid® adjuvant technology developedat North-West University in South Africa.

The Pheroid delivery system was initially developed as an adjuvant for drugsand vaccines.

AnnGro and Pheroids are covered by multiple patent filings as well asmanufacturing/formulation trade secrets.

3

Manufacturing, distribution

36

Current capacity: 91 200 L/year/day shift

Green technology; no waste; water re-used0

1000000

2000000

3000000

4000000

5000000

6000000

7000000

8000000

2011 2012 2013 2015

TUR

NO

VER

(R

AND

)

YEAR

Contribution to agriculture

37

0

100

200

300

400

500

600

2011 2012 2013 2015

Milli

ons

of R

and

Year

Total turnover of farmers

1 litre of product per 1 ton of liquid fertilizerCarbon footprint/ tonnage decreases; trade improvement

Pheroid® products: From the lab to pilot plant to factory

Current capacity: 91 200 L/year/day shift

Green technology; no waste; water re-used

Pheroid® in agriculture: From the lab to pilot plant to factory

Outputs

Students• 67 PhDs & MSc's delivered• 7 PhD/MSc at UFS; 1 at UP, 2 at

UCT, 1 at Basel, 2 at ETH, 1 at CATUC

• 4 NWU PhDs & 11 MSc in process• 5 Postdocs 3 new

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Articles :Peer reviewed high impact, international

Patents:International & SA

18 Jobs in industry

3 CompaniesNPC & for profit

Impact: growing more than agriculture

Use-inspired research that fuses separate areas of knowledge

• Economic impact by impact on health, energy, food security environment, HCD

• Scientific impact– smart renewable materials, – bioengineering for agriculture,– bioengineering for food & nutritional security

The challenge is to translate research into real-world impact

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42

ControlSprayed with Anngro & Comcat

“It is through science that we prove, but through intuition that we discover.” - Henri Poincaré

Is nanotechnology usable in agriculture? You decide. Questions?